WO2007120762A2 - Fluorescent particles bound to multifunctional scaffolds and their uses - Google Patents

Abstract

Fluorescent particles including quantum dots and fluorescent beads bound to multifunctional scaffolds can be used as taggants. The taggants can be further bound to a substrate of interest through binding sites on the multifunctional scaffolds.

Description

FLUORESCENT PARTICLES BOUND TO MULTIFUNCTIONAL SCAFFOLDS AND THEIR USES

BACKGROUND OF THE INVENTION

Field of the Invention This invention relates to fluorescent particles bound to multifunctional scaffolds and applications thereof, such as their use as taggants.

Description of the Related Art

Taggants are overt and/or covert markers that provide unique identity information. Security features based on taggants have been widely developed as brand-protection and anti-counterfeiting measures.

Fluorescent materials are particularly useful as covert taggants due to their sensitivity, ease-of-use and compatibility with many substrates.

They are typically invisible under normal lighting and are spectrally detectable when exposed to ultraviolet (UV) light. Fluorescent material-based taggants can therefore encode various substrates, for example, by incorporating verifiable image patterns on the substrates. The image patterns can be authenticated or "decoded" through direct visualization or a reading device under an ultraviolet light source. The identity information encoded in the image patterns can be presented in the form of serial numbers, barcodes, full-color . pictures, etc. Such information can further be stored in a database and retrieved for authentication purposes.

Microscopic fluorescent taggants have the advantage of being both physically and spectrally undetectable under normal circumstances, such as normal lighting condition. The microscopic dimensions of the taggants also allow for their easy incorporation into many carrier materials such as paint, ink, binder or adhesives. Given the immense potential of fluorescent taggants as an effective measure against counterfeiting, there remains a need in the art to provide microscopic taggant systems based on fluorescent particles having robust environmental stability and enhanced light intensity.

BRIEF SUMMARY OF THE INVENTION

In one embodiment, a composition comprises: a multifunctional scaffold having one or more first binding sites having a binding affinity for a surface of a substrate; and one or more fluorescent particles bound to the multifunctional scaffold, wherein the fluorescent particles emit light in a range of wavelength from about 400nm to about 1400nm when exposed to an excitation energy.

In another embodiment, a security device comprises: a substrate; and an invisible image indicia comprising a plurality of taggants positioned on the substrate, each taggant comprising one or more fluorescent particles bound to a multifunctional scaffold, the fluorescent particles emitting light in a range of wavelength from about 400nm to about 1400nm when exposed to an excitation energy, the multifunctional scaffold further including one or more binding sites bound to the substrate, wherein the invisible image indicia is substantially invisible when exposed to visible light but emits detectable fluorescence when exposed to the excitation energy.

In another embodiment, a method of forming a security device comprises: depositing a plurality of taggants on a substrate to form an invisible image indicia, each taggant comprising one or more fluorescent particles bound to a multifunctional scaffold, the multifunctional scaffold including one or more binding sites bound to the substrate, wherein the fluorescent particles emit light in a range of wavelength from about 400nm to about 1400nm when exposed to an excitation energy; irradiating the invisible image indicia to display a signature image of a fluorescence pattern; and storing the signature image in a readable format in a database In another embodiment, a security system is described herein, comprising: a security device comprising an invisible image indicia, the invisible image indicia having a plurality of taggants positioned on a substrate, each taggant comprising one or more fluorescent particles bound to a multifunctional scaffold, the multifunctional scaffold including one or more binding sites bound to the substrate, wherein the invisible image indicia is substantially invisible when exposed to light within a visible range but displays a signature image of a fluorescence pattern when exposed to an excitation energy; a database storing the signature image in a readable format; and a reading device for reading the invisible image indicia.

In another embodiment, a method of authenticating an article is described herein, comprising: marking an authentic article with an invisible image indicia, the invisible image indicia having a plurality of taggants bound to a surface of the authentic article, each taggant comprising one or more fluorescent particles bound to a multifunctional scaffold, the multifunctional scaffold further including one or more binding sites bound to the surface, wherein the invisible image indicia emit light in a range of wavelength from about 400nm to about 1400nm and displays a signature image of a fluorescence pattern when exposed to an excitation energy; irradiating the invisible image indicia to display a signature image of a fluorescence pattern; storing the signature image in a readable format in a database; irradiating the article to be authenticated; detecting the presence of an image of a fluorescent pattern; and matching the image to the signature image of the authentic article in the database. In another embodiment, a method is described herein, comprising: encapsulating a nanocluster of metal atoms in a matrix; and conjugating the matrix to a multifunctional scaffold, the multifunctional scaffold having a binding affinity for a substrate.

In another embodiment, a method is described herein, comprising: depositing a layer of taggants on a substrate according to a preselected pattern, the taggants comprising one or more fluorescent particles bound to a multifunctional scaffold, the fluorescent particles emitting light in a range of wavelength from about 400nm to about 1400nm when exposed to an excitation energy, the multifunctional scaffold further including one or more first binding sites bound to the substrate and one or more second binding sites having an affinity for a functional material; irradiating the substrate to display a fluorescent pattern, wherein the taggants act as an internal control to indicate any non-binding of the multifunctional scaffold to the substrate; and binding a functional material to the multifunctional scaffold via the second binding site thereof to form a functional material layer in the pre-selected pattern.

BRIEF DESCRIPTION OF THE SEVERAL VIEWS OF THE DRAWINGS In the drawings, identical reference numbers identify similar elements or acts. The sizes and relative positions of elements in the drawings are not necessarily drawn to scale. For example, the shapes of various elements and angles are not drawn to scale, and some of these elements are arbitrarily enlarged and positioned to improve drawing legibility. Further, the particular shapes of the elements as drawn, are not intended to convey any information regarding the actual shape of the particular elements, and have been solely selected for ease of recognition in the drawings.

Figure 1 illustrates schematically an embodiment in which a taggant composition comprises four fluorescent particles bound to a multifunctional scaffold, and a binding site having an affinity for a substrate.

Figure 2 illustrates schematically another embodiment in which a taggant composition further comprises a binding site having an affinity for a functional material. Figure 3 illustrates the results of tape tests conducted to evaluate the environmental stability of the taggants after a fixation process.

Figure 4 illustrates a security device suitable as an anti-counterfeit measure.

Figure 5 illustrates another security device suitable as a covert taggant having an immediately identifiable image; Figure 6 illustrates a security system in which a signature image can be detected and stored; and

Figure 7 illustrates another embodiment in which a taggant composition serves as an internal control during a plating process.

DETAILED DESCRIPTION OF THE INVENTION

In accordance to one embodiment, it is described herein a composition suitable as a microscopic fluorescent taggant. The composition comprises a multifunctional scaffold including one or more binding sites having an affinity for a substrate; and one or more fluorescent particles bound to the multifunctional scaffold, wherein the fluorescent particles, when exposed to an excitation energy, emit light in a range of wavelength from about 400nm to about 1400πm.

As used herein, "fluorescent taggant", or "taggant", refers to a marker added to a material to allow for various forms of detections or testing. The taggant comprises one or more fluorescent particles bound to a multifunctional scaffold. When irradiated with an excitation energy at a first wavelength, the taggant emits a radiative energy at a longer, second wavelength. Typically, the first wavelength is in the ultraviolet region, e.g., from 100nm and up to 400nm. The taggant emits detectable fluorescence in the visible and IR region, e.g., between about 400nm and about 3000nm. More typically, the taggant emits in the visible and near IR region, e.g., between about 400nm and about 1400nm.

In certain embodiments, the taggants are optically invisible (e.g., they do not emit detectable fluorescence when exposed to light in the visible range between 400nm and 700nm), until they are exposed to an excitation energy, as defined herein.

The fluorescent taggants are not only optically invisible but also forensically invisible due to their small physical sizes and the minute quantity typically required to generate detectable signals. Typically, an individual taggant is less than 1000 microns, less than 100 microns, less than 50 microns, less than 10 microns, less than 500nm, less than 50nm in its overall dimensions. They are therefore suitable candidates for creating covert, unique identification codes to be associated with an article. For example, information indicating the source or authenticity of an article can be encoded in a signature image created by the taggants. Taggants embedded in currency, government- issued documents and packaging material offer an easy solution to ensure authenticity and brand protection.

In one embodiment, as illustrated schematically in Figure 1 , a taggant 10 comprises one or more fluorescent particles 14, which are bound to a multifunctional scaffold 18. The multifunctional scaffold is a microscopic framework having diverse functionalities. The presence of the multifunctional scaffold provides one or more capturing groups 22 to bind to the respective fluorescent particles 14. As a result of the inherent aggregation of the fluorescent particles centered around the scaffold, a reasonable number of the fluorescent particles can be brought into relative close proximity. The local concentrations or density of the fluorescent particles can therefore be increased or otherwise optimized.

The same multifunctional scaffold further comprises an additional functionality that facilitates the adhesion of the scaffold to a substrate. As further illustrated in Figure 1 , the multifunctional scaffold 20 is bound to a substrate 24 via one or more first binding sites 28. This allows for an improved environmental stability of the taggants deposited on the substrate making them resistant to washing or abrasion. Conversely, a taggant can serve as an internal control to indicate a stable and successful binding event between the multifunctional scaffold and the substrate, the detail of which will be discussed below.

Furthermore, the multifunctional scaffold may also contain one or more second binding sites for a functional material. As illustrated schematically in Figure 2, a taggant 40 comprises the fluorescent particles 14 bound to the multifunctional scaffold 18, and a functional material 44 bound to the multifunctional scaffold via a second binding site 48. This architecture is particularly useful in a "bottom-up" approach in nano-device fabrications wherein the multifunctional scaffolds can direct the assembly of the functional material to form a functional layer, such as a conductive layer. The taggant 40 again serves as the internal control by illuminating the successful binding between the multifunctional scaffold and the substrate, as well as the binding between the multifunctional scaffold and the functional material.

The components of the taggant composition are described in detail below.

1. Fluorescent Particles In various embodiments, the fluorescent particles include, but are not limited to, microscopic fluorophores based on quantum dots or fluorescent beads.

"Fluorophore" refers to an entity, such as an organic dye or an inorganic quantum dot, that absorbs energy at one wavelength of radiation and subsequently reemits energy at another, different wavelength.

The energy absorbed is also referred to as "excitation energy".

An excitation energy can be any electromagnetic energy sufficient to excite the fluorophore from a steady state to a higher electronic state, also referred to as an "excited state". The fluorophore returns from the excited state to its steady state by photon emission, i.e., fluorescence. An excitation energy is typically higher in energy and shorter in wavelength than the fluorescent light emitted.

Examples of various types of excitation energy include light, X-rays and electron beams.

In certain embodiments, the fluorescent particles emit light in a range of wavelength between 400nm and 1400nm, when exposed to an excitation energy.

In certain embodiments, visible light is not sufficient in energy to excite the fluorescent particles from a non-emitting steady state to an excited state. The fluorescent particles therefore do not fluoresce when exposed to the normal lighting condition. "Normal lighting condition" refers to a visible range of the light spectrum that can be perceived by the human eye. The visible range is from about 400nm to 700nm.

These fluorescent particles typically require an excitation energy higher than the energy of visible light. For example, they may absorb ultraviolet light and fluoresce in the visible and near IR regions (between about 400nm and about 1400nm). Advantageously, these fluorescence particles are optically undetectable under the normal lighting condition, but become fluorescent when exposed to ultraviolet light.

In other embodiments, the fluorescent particles can be excited by white light in the visible range and emit light in a range of wavelength between 400nm and 1400nm.

The fluorescent particles are preferably of microscopic sizes. "Microscopic" refers to a dimensional measure of micron and sub-micron scales, including nanometer scales. Typically, a microscopic size is less than 1000 microns, less than 100 microns, less than 50 microns, less than 10 microns, less than 500nm, less than 50nm in overall dimensions.

In certain embodiments, the fluorescent particles include, but are not limited to, quantum dots and fluorescent beads.

(a) Quantum Dots Quantum dots are inorganic fluorophores based on semiconductor or metallic materials.

Most commonly, quantum dots are colloidal crystals or nanocrystals of a semiconductor material. They are typically structured as three-dimensional groupings or clusters of atoms (ranging from a few to as many as 10,000) in which the electron motion is confined by potential barriers in all three dimensions. This so called "quantum confinement" phenomenon has significant ramifications in the absorptive and emissive behaviors of the nanocrystals. For example, these semiconductor-based nanocrystals exhibit size-dependent tunable photoluminescence with narrow emission bandwidths (FWHM ~ 30 to 45 nm) that span the visible spectrum. They are further characterized with broad absorption bands, which allow for the simultaneous excitation of several particle sizes (colors) at a common wavelength.

Due to their sizes, quantum dots differ significantly from the bulk material in their optoelectrical properties. When the physical size of a semiconductor quantum dot is in the order of its exciton Bohr radius, the exciton is confined. As a result, the spacings of the electronic energy levels in the quantum dot are "discrete" whereas those of the bulk semiconductor are "continuous". The energy separation between the conductive band and valence band {i.e., bandgap) can be altered by changing the boundaries of the bandgap or by changing the geometry of the surface of the quantum dot. Accordingly, the size of the bandgap can be controlled by adjusting the size of the quantum dot. Because the emission frequency of a quantum dot is dependent on the size of the bandgap, it is therefore possible to fine-tune the radiative output wavelength (i.e., fluorescence) of a quantum dot with extreme precision. Similar to a semiconductor quantum dot, a metal quantum dot is a cluster of metallic atoms in nanometer scale. The confinement of the free electrons to a dimension comparable to the Fermi wavelength of an electron (~0.7nm) in a metallic nanocluster results in discrete, quantum-confined electronic transitions. Gold nanoclusters having fewer than 40 atoms exhibit even stronger fluorescence than semiconductor quantum dots. The absorption and fluorescence of gold nanoclusters are also size-tunable. See, e.g., Dickson, R.M. et al., (2004) Physical Rev. Lett. 93:7, 077402.

Because quantum confinement is a phenomenon associated with the size of the quantum dots as well as the nature of the material forming the nanocrystals, typically, the quantum dots are no more than 50nm in diameter. In certain embodiments, they are no more than 20nm in diameter. In other embodiments, they are no more than 5πm in diameter. In general, metal quantum dots are smaller dots than semiconductor quantum dots for the quantum confinement to take effect. For example, gold nanoclusters of fewer than 40 atoms (smaller than 1.4nm) emit fluorescence detectable in the visible and near IR regions. Semiconductor quantum dots such as CdSe, on the other hand, have size-tunable emissions spanning in the visible range with diameters typically larger than 2nm.

Even small variations in the cluster size can cause significant shifts in their spectroscopic properties. Known techniques such as photose lection can be used to select the nanoparticles of certain size or size range depending on the desired spectroscopic properties, such as excitation and/or emission wavelength. "Photoselection" stems from polarized light spectroscopy where randomly oriented fluorophores with a random distribution of excitation dipole vectors are exposed to a polarized light. Only fluorophores with the correct dipole vector orientation relative to the polarized light are excited (selected) out of the pool of randomly oriented fluorophores. In the context of the present application, the term refers to the selection of quantum dots of a certain size or size range (therefore having specific excitation and/or emission wavelength) out of a pool of various sizes of quantum dots having various excitation and/or emission wavelengths.

Although it is not always necessary, the quantum dots of certain embodiments are preferably monodispersed, e.g., the diameter of the dot varying approximately less than 10% between quantum dots in the preparation. In certain embodiments, the quantum dots have a quantum yield of more than 10%. More typically, the quantum yield is more than 20%. More typically, the quantum yield is more than 40%. "Quantum yield" is a direct measure of the light-emitting efficiency of a fluorophore. It can be quantified as the ratio between the photon emitted and the photons absorbed to produce the excited state from which the emission originates. The quantum dots are typically stable fluorophores, the quantum yields of which remain substantially unchanged when bound to a multifunctional scaffold.

In certain embodiments, the quantum dots have a fluorescence lifetime of more than 5ns, more than 20ns, more than 40ns. "Fluorescence lifetime" refers to the average time a fluorophore remains in the excited state, typically ranging from femtoseconds to nanoseconds. Quantum dots are generally marked with much longer fluorescence lifetimes and are therefore more photostable than the typical organic dyes.

Examples of the semiconductor quantum dots include, but are not limited to: a semiconductor material selected from a Group I IB-VIA compound, a Group MB-VA compound, a Group HIA-VIA compound, a Group IHA-VB compound, a Group IVA-VIA compound, a Group IB-IIIA-VIA compound, a Group IIB-IVA-VIA compound, and a Group HB-IVA-VA compound. A "Group IIB-VIA compound" refers to a compound formed by an element selected from Group HB and an element selected from Group VIA of the Periodic Table. The designation of the Group numbers in the Periodic Table is according to the CAS (Chemical Abstract Service) convention and is known to one skilled in the art. For example, Group MB refers to the group of transitional elements (designated by B) having two electrons in the valence shell, whereas Group VIA refers to the group of main group elements (designated by A) having six electrons in the valence shell. Examples of Group IIB-VIA compounds include ZnS₁ ZnSe, ZnTe, CdS, CdSe, CdTe, HgS₁ HgSe and HgTe.

According to the same naming convention as described above, additional examples of the semiconductor quantum dots include, but are not limited to: AIN, AIP, AIAs, AISb, GaN, GaP, GaAs, GaSb, GaSe, InN, InP, InAs, InSb, TIN, TIP, TIAs, TISb₁ PbS, PbSe, PbTe, and mixtures thereof.

Typically, semiconductor quantum dots assume a core/shell structure, in which the above semiconductor material forms a core that is passivated with a different semiconductor material. Passivating the surface of the core quantum dot can result in an increase in the quantum yield of the fluorescence emission, depending on the nature of the passivation coating. Typical shell materials include ZnS and CdS. These quantum dots of the core/shell structure can also be represented by, for example, CdSe/ZnS, CdTe/ZnS and CdTe/ZnS.

The semiconductor quantum dots can be prepared by known methods in the art. For example, they can be synthesized according to the methods described in, e.g., Klimov, V.I., Semiconductor and Metal Nanocrystals: Synthesis and Electronic and Optical Properties (Optical Engineering) (CRC, November 7, 2003), Li, L. H. et ai, (2004) Nano Lett. 4:1 , 2261-2264, and Battaglia, D. et ai, (2002) Nano Lett. 2:9, 1027-1030.

In addition, quantum dots can be fabricated through biological means. In particular, biological materials such as viruses and proteins can function as templates to create quantum dots. In certain embodiments, the dimensions of the quantum dots thus created correlate to the dimensions of the biological templates. In other embodiments, the biological templates can be engineered to exhibit selective affinity for particular types of semiconductor material. More detailed description of biofabrication of quantum dots can be found in, e.g., Mao, et at. (2003) PNAS (supra), and U.S. Published Application 2005/0130258.

Commercial sources including Evident Technologies (Troy, NY) and Invitrogen (Carlsbad, CA) also provide a wide range of quantum dot material systems.

Examples of the metallic quantum dots include, but are not limited to: a noble metal selected from gold (Au), silver (Ag) and copper (Cu). Fluorescent metal clusters can be prepared according to the methods as described in, for example, Dickson, R.M. etai, (2004) Physical Rev. Lett. 93:7, 077402. Gold nanodots of discrete sizes have been synthesized by reducing a gold salt in the presence of poly(amidoamine) dendrimers. Dickson, R.M. (Supra). Similarly, individual silver nanodots (Ag₂-Aga) have been prepared through photo-activated reduction, also in the presence of poly(amidoamine) dendrimers. Dickson, R.M. etai (2002) J. Am. Chem. Soc. 124, 13982-13983. More recently, the formation of silver nanocluster using DNA as templates was reported. Dickson, R.M. et ai (2004) J. Am. Chem. Soc. 126:16, 5207-5211. (b) Fluorescent Beads

Fluorescent beads, or "fluorescent microspheres" have been widely developed for diagnostic applications in biological systems, including flow cytometry. Fluorescent beads are typically beads made of chemically inert polymer materials labeled or loaded with fluorophores, such as organic dyes or quantum dots

Polystyrene and copolymers based on polystyrene (e.g., polystyrene-divinylbenzene) are commonly used to prepare the polymer beads with high uniformity. Other suitable polymeric materials include polymethylmethacrylate (PMMA), polyvinyltoluene (PVT), styrene/butadiene (S/B) copolymer and styrene/vinyltoluene (SΛ/T) copolymer.

The fluorophores can be loaded or embedded into the internal cavities of the beads. Alternatively, the fluorophores can be tethered to the beads. These techniques are known to one skilled in the art. Fluorescent beads are also commercially available from vendors including Bangs Laboratories, Inc. (Fishers, IN), ProSciTech Corporation (Australia), and Fisher Scientific International Inc. (Hampton, NH).

2. Multifunctional scaffold Colloidal semiconductor quantum dots have been used as fluorescent taggants in security devices, as described in, e.g., U.S. Patent No. 6,692,031 and U.S. Published Patent Application No. 2004/0233465, however, they experience drawbacks such as poor adhesion to the substrates and unsatisfactory emission intensities. In contrast, the fluorescent particles are bound to a multifunctional scaffold, which further comprises one or more binding sites for a substrate of interest. The taggants thus configured provide more intense fluorescent emission and superior environmental stability.

As used herein, a "multifunctional scaffold" refers to a stable framework having diverse functional elements, including various binding sites for binding to one or more fluorescent particles, a substrate material, and/or a functional material, as defined herein. Structurally, the framework can be a discrete molecular structure or multimolecular structure of a biological or synthetic origin. The multifunctional scaffold can be chosen to have dimensions large enough to separate the fluorescent particles from the substrate to prevent quenching interactions. In particular embodiments, the multifunctional scaffold can be an individual molecule, also referred herein as a "multifunctional molecular scaffold". For example, a multifunctional molecular scaffold can be a biomolecule. As used herein, the term "biomolecule" refers to a carbon-based organic molecule of a biological origin or an organic molecule that mimics or resembles biological activities, properties and interactions. Typically, a biomolecule comprises a plurality of subunits (building blocks) joined together in a sequence via chemical bonds. Each subunit comprises at least two reactive groups such as hydroxyl, carboxylic and amino groups, which enable the bond formations that interconnect the subunits. Examples of the subunits include, but not limited to: amino acids (both natural and synthetic) and nucleotides. A biomolecule can be naturally occurring or synthetic. Examples of the suitable biomolecules include peptides, proteins, nucleic acids, polynucleotides, organic polymers and other simple or complex carbon-containing molecules, and combinations thereof.

In certain embodiments, the biomolecules are further characterized by their ability to recognize and bind to an inorganic material with specificity and/or selectivity. In particular, biomolecules comprising subunits of amino acids are found to exhibit sequence-specific binding behavior toward inorganic materials. Examples of amino acid-based biomolecules include, but are not limited to peptides, antibodies, block copolypeptides or amphiphilic lipopeptides.

As used herein, "peptide" refers to a sequence of two or more amino acids joined by peptide (amide) bonds, including proteins. The arnino- acid building blocks (subunits) include naturally-occurring α-amino acids and/or unnatural amino acids, such as β-amino acids and homoamino acids. Moreover, an unnatural amino acid can be a chemically modified form of a natural amino acid. In particular, an amino acid coupled to a labile protecting group can be incorporated into a peptide sequence. A 'labile protecting group" refers to a protecting group that deactivates a functionality and can be readily removed to restore the functionality. The removal of the protecting group is typically triggered by an external event, such as light irradiation, heat, enzymatic or chemical manipulation. A functionality, such as the binding activity of an amino acid, can therefore be manipulated through controlling the external event. More detailed description of using labile protecting groups to manipulate a peptide's activity can be found in U.S. Provisional Application entitled "Fabrication of Inorganic material Using Templates With Labile Linkage", filed on March 9, 2006, in the name of Cambrios Technologies Corporation, which application is incorporated herein by reference in its entirety. As used herein, "block copolypeptide" refers to polypeptides having at least two covalently linked domains ("blocks"), one block having amino acid residues that differ in composition from the composition of amino acid residues of another block. "Amphiphilic lipopeptide" refers to a hydrophilic peptide head group conjugated to a hydrophobic group, such as a fatty acid or steroid. As used herein, "polynucleotide" refers to an oligomer of about 3-

50 nucleotide units interconnected by a phosphate backbone, whereas the term "nucleic acid" typically refers to more than 50 nucleotide units. A polynucleotide of 2-10 nucleotide units is also referred to as "oligonucleotide". The nucleotide subunits include all major heterocyclic bases naturally found in nucleic acids (uracil, cytosine, thymine, adenine and guanine) as well as naturally occurring and synthetic modifications and analogs of these bases such as hypoxanthine, 2-aminoadenine, 2-thiouracil and 2-thiothymine. The nucleotide subunits further include deoxyribose, ribose and modified glycosides.

Typically, the multifunctional molecular scaffold is of a nano-scale architecture having any one or more of the following physical forms: linear, curved, branched, folded, looped, hinged, circular, resilient, elastic and/or flexible.

In other embodiments, the multifunctional scaffold can be a complex multimolecular structure, for example, of a biological origin. Multimolecular, pre-organized biomolecular architectures are chemically and spatially confined environments suitable for the attachments and/or construction of nanoclusters. They are typically rich in functionalities that can affect various binding events. Examples of such biological multimolecular structures include but are not limited to biological cells, viruses, bacteria, phages, self-assembled peptide structures, and proteins. These multimolecular structures range from a few nanometers to a few microns in their dimensions. Their monodispersity and ability to self-assemble make them suitable for highly ordered smectic- ordering structure. See, e.g., Hartgerink, J.D. et a/., (2002) PNAS 99, 5133- 5138. Additionally, certain biological multimolecular structures (e.g., a M13 bacteriophage) can be genetically engineered and the binding sequences having the most desirable binding characteristics (e.g., material specificity) be selected. See, e.g., Belcher, A. er a/., (2004) Science, 303, 213-217; Belcher, A. et al., (2002) Science 296, 892-895; Belcher, A. et ai, (2000) Nature 405 (6787) 665-668.

In yet another embodiment, the multimolecular structure can be based on synthetic polymers, also referred to as "polymeric scaffold". Typically, the polymeric scaffolds include polymer beads comprising surface functionalities such as -COOH, -SH and -NH₂ and/or a binding partner of a recognition pair. Like biomolecule-based multifunctional scaffolds, polymeric scaffold contains at least one binding site that binds to a substrate of interest; typically, the binding site is a surface functionality.

Typically, polystyrene or polystyrene cross-linked by divinylbenzene can be used to form beads of high uniformity. Other suitable polymeric materials include polymethylmethacrylate (PMMA), polyvinyltoluene (PVT), styrene/butadiene (S/B) copolymer, styrene/vinyltoluene (S/VT) copolymer. The sizes of the bead are readily controllable through controlling the polymerization processes. Such techniques are known to one skilled in the art, as noted herein. Suitable polymer beads can be purchased from Bangs Laboratories, Inc. (Fishers, IN).

In further embodiments, the multifunctional scaffold can be a combination of the molecular and multimolecular structures described above. A non-limiting example includes a peptide molecule displayed on a phage. In certain embodiments, the multifunctional scaffold comprises multiple functional elements for binding to an assortment of target materials. These functional elements include one or more capturing groups for binding to one or more fluorescent particles, one or more first binding sites having an affinity for a substrate of interest and an optional second binding site having an affinity for a functional material.

"Binding site", used interchangeably herein with "binding sequence", refers to the minimal structural elements within a multifunctional scaffold that are associated with or contribute to its binding activities. As used herein, the terms "bind" and "couple" and their respective nominal forms are used interchangeably to generally refer to one entity being attracted to another to form a stable complex. The underlying force of the attraction, also referred herein as "affinity" or "binding affinity", can be any stabilizing interaction between the two entities, including adsorption and adhesion. Typically, the interaction is non-covalent in nature; however, covalent bonding is also possible. A covalent bond is formed between two atoms sharing at least a pair of electrons. A non-covalent bond can be based on van de Waals force, electrostatic interaction, hydrogen bonding, dipole-dipole interaction or a combination thereof. In one embodiment, a binding site comprises a chemically reactive functional group of the biomolecule, such as thiol (-SH)₁ hydroxy (-OH), amino (-NH₂) and carboxylic acid (-COOH). For example, the thiol group of a cysteine effectively binds to a gold quantum dot.

In another embodiment, a binding site is a defined sequence of subunits of the biomolecule and more than one functional group may be responsible for the affinity. Additionally, conformation, secondary structure of the sequence and localized charge distribution can also contribute to the underlying force of the affinity.

The magnitude of the binding affinity can be quantitatively represented by an association constant of the binding equilibrium. Known methods in the art, such as Langmuir model for adsorption of analytes on a surface, can be used to measure the association constant- Typically, the association constant can be greater than 1x10⁵ M^"1, greater than 1x1θ7 M^"1, greater than 1x10⁹ M^"1 or greater than 1x10¹¹ M^"1.

The binding activities of the multifunctional scaffold include but are not limited to: their ability to specifically recognize and bind to a material or to display a favorable affinity toward one material over another. The term "specifically" is a term of art that would be readily understood by the skilled artisan to mean, when referring to the binding capacity of a biomolecule, a binding reaction that is determinative of the presence of the substrate in a heterogeneous population of other substrates, whereas the other substrates are not bound in a statistically significant manner under the same conditions. Specificity can be determined using appropriate positive and negative controls and by routinely optimizing conditions. The phrase further applies to a binding reaction that is determinative of the presence of the functional material in a heterogeneous population of other inorganic materials.

For a multifunctional scaffold, a binding event can occur by either "conjugation" or "nucleation". The terms "conjugation" and "conjugate" refer in general to a process in which a multifunctional scaffold binds to a pre-made target material, with or without a cross-linker. A "cross-linker" is a chemical moiety that bridges two functional groups by forming a stable bond with each functional group. A suitable cross-linker can be selected based on such factors as the reactivity of the functional groups to be bridged and the desired length of the linker, the tactics of which is readily recognized by a skilled person.

The terms "nucleation" and "nucleate" refer to a process in which a precursor is converted to a target material in the presence of a multifunctional scaffold, the in situ generated target material binds to and grows on the multifunctional scaffold. The "in situ generated target material" contrasts from the "pre-made target material" in that the former requires an initial conversion from the precursor in the presence of the multifunctional scaffold, whereas the latter has been prepared independently of the multifunctional scaffold. In one embodiment, the target material is a nanoparticle, as defined herein. For example, peptides of certain sequences selectively nucleate metal nanoparticles through reduction of a metal salt in a solution. Likewise, certain peptides selectively nucleate semiconductor nanoparticles. See, e.g., Flynn, Mao, et al., (2003) J. Mater. Sci. (supra); Mao, et al., (2003) PNAS (supra).

In a further embodiment, the initially nucleated nanoparticle can act as a seed material that catalyzes the growth of another target material. The term "seed material" therefore refers to a first target material that causes the growth of a second target material thereon. The first and second target material may be the same or different. For example, when a seed material is exposed to a precursor of a second target material in a solution phase, the seed material catalyzes the conversion of the precursor into the second target material. Typically, the second inorganic material can form a "shell" to the "core" represented by the seed material. More typically, the second target material forms a continuous layer over a seed material layer. This process is also referred to as "mineralization". More particularly, when the second target material is a metal, the process forming a metal layer over a seed layer is also referred to as "metallization" or "plating". Examples of the seed material include Ni, Cu, Pd, Co, Pt, Ru, Ag,

Co alloys or Ni alloys nanoparticles. The second inorganic material that can be subsequently plated include metals, metal alloys and metal oxides, for instance, Cu, Au, Ag, Ni, Pd, Co, Pt₁ Ru, W, Cr, Mo, Ag, Co alloys (e.g., CoPt), Ni alloys, Fe alloys (e.g., FePt) or TiO₂, Co₃O₄, Cu₂O, HfO₂, ZnO, vanadium oxides, indium oxide, aluminum oxide, indium tin oxide, nickel oxide, copper oxide, tin oxide, tantalum oxide, niobium oxide, vanadium oxide or zirconium oxide. More details of using seed layers to direct functional layer formations are described in co-pending applications U.S. Provisional Application 60/680,491, entitled "Biologically Directed Seed Layers and Thin Films", filed May 13, 2005, and U.S. Provisional Application 60/752,019, entitled "Seed Layers, Cap Layers, and Thin Film and Methods of Making Thereof, filed December 21 , 2005, both in the name of Cambrios Technologies Corporation, which references are incorporated herein in their entireties.

As used herein, a "capturing group" refers specifically to a binding site that binds to a fluorescent particle. A capturing group includes, but is not limited to: a functional group, a defined binding sequence and a binding partner that is a member of a recognition pair, as defined herein.

A functional group directly binds to a fluorescent particle or a surface functionalized fluorescent particle by forming a covalent attachment. A defined binding sequence either conjugates to a fluorescent particle or nucleates a fluorescent particle from a solution containing a precursor of the fluorescent particle. The term "binding partner", as used herein, refers to a member of a specific recognition pair, each member of the specific recognition pair being a binding partner of the other member. They are also referred to as "ligand" and "anti-ligand", respectively. As described herein, a capturing group can recognize and bind a corresponding binding partner on a fluorescent particle. The interactions between the binding partners are typically non- covalent in nature. They can nonetheless be sufficiently stable to withstand the conditions of the typical use of the fluorescent particles. An example of a recognition pair is streptavidin and biotin, which forms one of the most stable interactions in biology. Fluorescent particles can be surface functionalized with streptavidin, whereas a multifunctional scaffold such as a peptide can be biotinylated, by known methods in the art.

In certain embodiments, the functional elements of the multifunctional scaffold are also a part or a segment of the structural framework of the multifunctional scaffold. For example, a peptide scaffold may comprise a defined "binding sequence" that binds to a target material such as a fluorescent particle or a substrate. The defined "binding sequence" therefore contributes both to the structural and functional elements of the multifunctional scaffold.

In other embodiments, the functional elements are independent of the structural elements of the multifunctional scaffold. Typically, these involve chemically reactive functionalities that do not contribute to the physical form of the scaffold, or are tethered to the scaffold.

In certain embodiments, the multifunctional scaffolds can also be optionally selected, designed or engineered to provide suitable spacing between the functional elements, or to control the number of the functional elements.

Suitable multifunctional scaffolds are therefore selected based on such criteria as specific binding characteristics toward a fluorescent particle, a substrate, as well as toward a functional material, collectively referred as a "target material" herein.

The term "target material" refers to a material that binds to the multifunctional scaffold, including the fluorescent particles, the substrate and the functional material. The target material can be classified into non-biological material and biological material. Examples of the non-biological target material include, but are not limited to inorganic materials such as metals, metal oxides, metal alloys, semiconductive materials, minerals, ceramic, glass, salts, and combinations thereof. Metals may include Ag, Au, Sn, Zn, Ru, Pt, Pd, Cu, Co, Ni, Fe, Ba, Sr, Ti₁ Bi, Ta, Zr, Mn, Pb, La, Li, Na₁ K, Rb₁ Cs, Fr, Be₁ Mg₁ Ca, Nb, Tl, Hg₁ Rh, Sc₁ Y, Cr , Mo, W, or their alloys and oxides, including brass and steel. Additional inorganic materials may also include, e.g., high dielectric constant materials (insulators) such as barium strontium titanate, barium zirconate titanate, lead zirconate titanate, lead lanthanum titanate, strontium titanate, barium titanate, barium magnesium fluoride, bismuth titanate, strontium bismuth tantalite, and strontium bismuth tantalite niobate, or variations, thereof, known to those of ordinary skill in the art. Biological target material, as further discussed herein, can be animal matters (including humans), plant matters or parts thereof.

As used herein, a "substrate" is a solid or semi-solid surface to which the multifunctional scaffold binds through either covalent or non-covalent interactions. A substrate is typically a non-biological material, including an inorganic material, as defined herein. A substrate can also be organic, such as a polymer. In one embodiment, a substrate is a micro-fabricated material.

Examples of the suitable substrate materials include, but are not limited to: semiconductor materials (e.g., silicon, germanium, etc.), Langmuir films, glass (including functionalized glass), paper, ceramic, carbon, polymer materials, including polycarbonates, polyamides (e.g., Nylon®), polyimides (e.g., Kaptoπ®), polystyrene, PTFE (e.g., Teflon®), and polyesters (e.g., Mylar®), dielectric materials, mica, quartz, gallium arsenide, metals, metal alloys, metal oxides, fabric, and combinations thereof. The surface may be large or small and not necessarily uniform but should act as a contacting surface (not necessarily in monolayer). The substrate may be porous, planar or nonplanar.

In some embodiments, the substrate can also be a biological material, such as a surface of a plant or animal matter. A "plant matter" can include both a living plant and a plant product. For example, wood surface is a suitable substrate whether it is part of a tree or a piece of furniture. An "animal matter" refers to an animal (including a human) or an animal product such as leather, hide and features. For example, the skin of an animal is a suitable substrate. Moreover, the phrase "animal matter" extends to man-made articles incorporating the animal products as noted herein. For example, shoes or jackets made of leather are contemplated within the definition of the "substrate".

The substrate may comprise functional groups such as amino, carboxyl, thiol or hydroxyl on its surface. Advantageously, the functional groups on the substrate allow the multifunctional scaffold to be covalently bound to the substrate, directly or through a cross-linker. This covalently binding can be the sole affinity between the substrate and the multifunctional scaffold, or in addition to the sequence-specific binding affinity of the multifunctional scaffold.

In other embodiments, the multifunctional scaffold can form covalent bonds with the substrate in the absence of any functional groups on the substrate. Typically, a functional group of the multifunctional scaffold, also referred as an "adhesive group", can form a covalent bond with the surface of the substrate. A number of functional groups can act as adhesive groups, including catechol derivatives, which forms strong bonds with metal surfaces such as aluminum or steel as well as other inorganic surfaces such as CaCCh or silicate. See, e.g., Fan, X. et al., (2005) J. Am. Chem. Soc, 127, 15843- 15847. They also bind to organic surfaces such as wood. For example, a peptide multifunctional scaffold comprising the amino acid Tyrosine can be enzymatically oxidized to L-3,4-dihydroxyphenylana!ine (L-DOPA), a catechol derivative. L-DOPA has been demonstrated to form strong covalent bonds with a substrate, as shown in Scheme I. See, e.g., Dalsin, J. L. et al., (2005) Materials Today, 8(9), 38-46.

SCHEME I

tyrosine-containing peptide

As noted herein, in certain embodiments, the multifunctional scaffold exhibits sequence-specific affinity to a given target material. Table 1 shows examples of peptides (SEQ ID NO: 1-24) exhibiting specific affinity for a variety of materials. Table 1

SEQ ID Peptide Sequence Material Type of Binding

1 CNNPMHQNC ZnS nucleation, affinity¹¹²-³'⁴

2 LRRSSEAHNSIV ZnS nucleation, affinity¹'^{3 4}

3 QNPIHTH PbS conjugation¹

4 CTYSRLHLC CdS nucleation, affinity¹

5 SLTPLTTSHLRS CdS nucleation, affinity¹

6 WDPYSHLLQHPQ Streptavidin conjugation⁵

7 HNKHLPSTQPLA FePt nucleation, affinity⁶'⁷

8 CNAGDHANC CoPt nucleation, affinity⁶

9 SVSVGMKPSPRP L10 FePt: nucleation, affinity⁷

10 VISNHRESSRPL L10 FePt: nucleation, affinity⁷

11 KSLSRHDHIHHH LIO FePt: nucleation, affinity⁷

12 VSGSSPDS Au nucleation, affinity⁸

13 AEEEED Ag, Co₃O₄ nucleation, affinity⁹

14 THRTSTLDYFVI PPyCI affinity¹⁰

15 KTHEIHSPLLHK CoPt affinity

16 EPGHDAVP Co²⁺ nucleation, affinity¹¹

¹ Flynn, CE. et al., "Synthesis and organization of naπoscale H-VJ semiconductor materials using evolved peptide specificity and viral capsid assembly," (2003) J. Mater. ScL, 13, 2414- 2421.

² Lee, S-W et al., Ordering of Quantum Dots Using Genetically Engineered Viruses," (2002) Science 296, 892-895.

³ Mao, CB. et al., "Viral Assembly of Oriented Quantum Dot Nanowires," (2003) PNAS, vol. 100, no. 12, 6946-6951.

⁴ US2005/0164515

⁵ Lee, S-W et al., "Viral-based alignment of inorganic, organic and biological πanosized materials" (2003) Advanced Material (Weinheim, Germany) 15(9), 689-692.

⁶ Mao, CB. et al., "Virus-Based Toolkit for the Directed Synthesis of Magnetic and Semiconducting Nanowires," (2004) Science, 303, 213-217.

⁷ Reiss, B.D. et al., "Biological route to metal alloy ferromagnetic nanostructures" (2004) Nano Letters 4(6), 1127-1132.

⁸ Huang, Y. et al., "Programmable assembly of nanoarchitectures using genetically engineered viruses" (2005) Nano Letters 5(7), 1429-1434.

⁹ U.S. Patent Application No. 11/254,540.

10 US2004/0127640 SEQ ID Peptide Sequence Material Type of Binding

17 HTHTNNDSPNQA GaAs affinity¹²-¹³

18 DVHHHGRHGAEHA

Dl CdS nucleation, affinity¹⁴

19 KHKHWHW ZnS₁ Au, CdS affinity¹⁵

20 RMRMKMK Au affinity¹⁵

21 PHPHTHT ZnS affinity¹⁵

22 CSYHRMATC Ge dislocations affinity¹⁶

23 CTSPHTRAC Ge dislocations affinity¹⁶

24 LKAHLPPSRLPS Au affinity⁹

The term "functional material" refers to a target material that binds to a multifunctional scaffold and can be henceforth directed to assemble into a functional structure. Such a functional structure includes, for example, a functional layer in semiconductor fabrications such as an integrated circuit layer. Examples of the functional material include but are not limited to: metal, metal oxide, a semiconductive material, an insulating material and a magnetic material. Advantageously, in one embodiment, the multifunctional scaffolds have a tendency to self-assemble and enable an orderly construction of the target material, which makes it possible for a "bottom-up" approach in fabricating nano-sized components. See, e.g., U.S. Patent Application Publication Nos. 2005/0170336, 2003/0073014.

In one embodiment, the target material is one or more nanoparticles. It should be noted that "nanoparticle" encompasses any

Lee, S-W. θt al., "Cobalt ion mediated self-assembly of genetically engineered bacteriophage for biomimic Co-Pt hybrid material" Biomacromolecules (2006) 7(1 ), 14-17.

¹² Whaley, S.R. et al., "Selection of peptides with semiconductor binding specificity for directed nanocrystal assembly" (2000) Nature, 405(6787), 665-668.

13

US2003/0148380

14 US2006/0003387

¹ Peelle, B. R. et al., "Design criteria for engineering inorganic material-specific peptides" (2005) Langmuir 21 (15), 6929-6933.

¹⁶ U.S. Patent Provisional Application 60/620,386. inorganic particles of less than 100nm in diameter. More typically, the nanoparticles are less than 50nm in diameter, less than 25nm in diameter or less than 10nm in diameter. The quantum dots therefore belong to a special subset of the nanoparticies that emit size-dependent fluorescence due to "quantum confinement".

The nanoparticles can include pre-made nanoparticles, such as colloidal gold, which can be directly conjugated to a multifunctional scaffold. Alternatively, the nanoparticles can be nucleated on a multifunctional scaffold out of a solution phase. Typically, the solution phase contains a precursor material. For example, metallic nanoparticles can be nucleated onto a multifunctional scaffold, such as a peptide, by reducing a precursor metal salt to the metal. In certain embodiments, reducing agents such as NaBHU and dimethylamine borane can be used. The metallic nanoparticles may also be nucleated without an added reducing agent when the peptide itself contains a reducing component. For example, a peptide may comprise a cysteine residue in which a free thiol group contributes to the reduction of a metal salt and subsequent nucleation of the resultant metal on the peptide.

Examples of the inorganic nanoparticles include particles of metals, metal oxides, semiconductive materials, magnetic materials, and dielectric materials. Examples of suitable inorganic particles are summarized in Table 2. TABLE 2

Elemental conductors Au, Ag, Pd, Mo, Cu, Fe, Co, Pt, Ru, Ni, Zn, Sn, Cr₁ W

Group IIB-VIA materials ZnS, CdS, CdSe, CdTe

Group IHA-VA materials GaAs, GaN, InP, BN

Magnetic materials Fe₂θ₃, Fe₃O₄, CoPt, FePt, SmCo₃, Co

Other semi-conductive ITO, PbS, AI₂O₃, SiNx, TaN, ZnO, CuO, CoWP, and conductive materials CoWB₁ CoP, NiWP, NiWB, NiP, carbon nanotubes, Si, Ge, Ce₁ TiO₂, Co₃O₄, HfO₂, indium oxide, niobium oxides, vanadium oxides Dielectric material SiO₂, BaTiO₃, LiNbO₃

Other inorganic material CaCO₃, Ca₃(PO₃)₂

As noted herein, the multifunctional scaffolds are characterized as having diverse functionalities. These functionalities can bind to an assortment of target materials, often with selectivity and/or specificity. Advantageously, the multifunctional scaffolds can be designed and engineered to comprise the desired binding functionalities following an initial identification of the binding sequences specific to a given target material.

In one embodiment, biomolecules having desired binding behaviors can be selected by combinatorial library screening. In particular, exact binding sequences can be identified using tools and protocols developed in the field of molecular biology, such as phage display libraries.

More specifically, biological structures (e.g., a bacteriophage) that are genetically engineered can be used to express or display one or more random biomolecules, such as a peptide. For example, the biomolecule can be a random peptide of a specified length expressed as a portion of the virus' exterior coat.

The advantage of using an expression system to obtain biomolecules is that large numbers of different biomolecules (e.g., libraries) can be provided (i.e., displayed on the phage) and screened for material- recognition, which enables rapid identification of sequences that have specific and/or selective affinity for one or more materials.

More specifically, a filamentous virus (i.e., bacteriophage) may be engineered to produce large amounts of one or more types of biomolecules, such as peptides. Commercially available libraries that contain random assortments of biomolecules with diversified attributes (e.g., length, innate structure, species) may also be used. For example, bacteriophage libraries (also referred to herein, as phage libraries) have been developed that include peptides of specific lengths (e.g., 12 amino acid linear, 7 amino acid linear, or 7 amino acid constrained where cysteines are at the first and ninth position on the peptide to create a loop by the disulfide linkage between the two cysteines) on the minor coat protein (pill) of the M13 coliphage. In one embodiment, a Ph.D.- 12™ Phage Display Peptide Library Kit (New England Biolabs, Beverly, Mass.) can be used. The kit contains a library with approximately 10⁹ discrete linear peptide inserts fused to the pill coat protein of the M13 coliphage.

The phage libraries can be screened against one or more materials, a process known as biopanning. Initially in the biopanning process, phages with randomized peptides that have specific binding affinity for a given material can be collected after cycles of incubation with the material and washing to remove those phages displaying peptides that are non-binding or non-specifically binding. The peptides on the phages that exhibit specific binding can be collected and introduced to bacteria, such as Escherichia coli (E. coli) ER2837 bacteria (New England Biolabs, Ipswich, Mass.) that has been cultured at least about overnight. The techniques used are those well known to one of ordinary skill in the art of molecular biology and includes plating the phage or allowing a various concentrations of phage solutions to infect a known amount of bacteria. When using the infection technique, bacteria with lacZ gene may be used and plated in the presence and absence of isopropylthio-β- D-galactoside (IPTG) and 5-bromo-4-chloro-3-hydroxyindolyl-β-D-galactose (X- gal) for visual determination of bacterial growth on "titer plates." The phage concentration may then be determined by the following:

Concentration of phage from titer plate (pfu/μL) x (1 μl_/1 E⁶L) x (1 mole/6.023x10²³ molecules), wherein, pfu = plaque forming unit.

Several biopanning rounds are generally used to determine material-specific biomolecules and their material-specific binding sites. For each biopanning round, the phage concentration is used to determine the amount (as volume) used in the next, round of biopanning against the material. A fresh piece of material is then used for the next screening, where the phage amount is at least about 10⁹ pfu. Multiple rounds of biopanning are to follow, generally at least about five rounds to determine the consensus sequence involved in binding the material.

Some or all of the above steps can be automated for rapid analysis (high-throughput screening) to identify specific biomolecules that can bind or recognize a selected material with specificity and/or selectivity. These techniques are further described in detail in the following U.S. patent publications: (1) US 2003/0068900 entitled "Biological Control of Nanoparticle Nucleation, Shape, and Crystal Phase"; (2) US 2003/0073104 entitled "Nanoscale Ordering of Hybrid Materials Using Genetically Engineered Mesoscale Virus"; (3) US 2003/0113714 entitled "Biological Control of

Nanoparticles"; and (4) US 2003/0148380 entitled "Molecular Recognition of Materials"; and (5) US 2004/0127640 entitled "Composition, Method and US of Bi-Functional Biomaterials", all of which, including the sequence listings described, are incorporated herein by reference in their entireties. Biomolecules (e.g., peptides) that successfully bind to a specific material can thus be recovered and amplified. The identity of the biomolecule can be ascertained by known techniques including isolation of the phage, sequencing its DNA and translating the DNA sequence to peptide sequence. The peptide thus identified can also be synthesized independently of the virus, as is known in the art, e.g., by solid phase synthesis, with the same function and affinity as seen while displayed on the virus.

Typically, a phage-display library is based on a combinatorial library of random peptides containing between 7-12 amino acids. A peptide exhibiting specific binding to a material can be unambiguously identified by its sequence according to the process described above. Moreover, the part of a peptide sequence that in fact contributes to the binding, i.e., the binding sequence, can be determined by identifying a consensus sequence based on multiple rounds of biopanning. Additionally, screening libraries of shorter peptides against a substrate can assist with pinpointing the exact binding sequences. Furthermore, given the small size of the peptides in a phage library, computer analysis can also be used to accurately predict or confirm the identity of a binding sequence.

In another embodiment, the structural knowledge of the desired binding sequences enables a rational design of a multifunctional scaffold, particular with respect to multifunctional molecular scaffold based on peptides and oligonucleotides. Well-known techniques such as site-directed mutagenesis can be used to rationally introduce modifications to one of more areas of the multifunctional molecular scaffold in order to produce variants in other species. The mutation that leads to a desirable change (e.g., better specificity) in the binding characteristics can be used as a guide to work in other sequences.

Through peptide (or polynucleotide) engineering, many different varieties of binding sequences can be placed at different locations on a multifunctional scaffold. Suitable multifunctional scaffolds can therefore be designed and manufactured to combine a number of desired binding characteristics. For example, a multifunctional scaffold may comprise one or more peptide sequences, e.g., SEQ ID No:24, to nucleate one or more Au nanoparticles. Moreover the same multifunctional scaffold may further comprise another peptide sequence, e.g., SEQ ID No: 14, to bind to a conductive polymer substrate such as oxidized polypyrrol doped with chlorine (PPyCI). Sequences may also be selected to create nanoparticles of different sizes or materials to allow for a more complex fluorescence emission.

More detailed information on genetically engineering peptides to create binding sequences are described in: e.g., Belcher, A. et al., Virus-Based Toolkit for the Directed Synthesis of Magnetic and Semiconducting Nanowires," (2004) Science, 303, 213-217; Belcher, A. et al., "Ordering of Quantum Dots Using Genetically Engineered Viruses," (2002) Science 296, 892-895; Belcher, A. et a/., "Selection of Peptides with Semiconductor Binding Specificity for Directed Nanocrystal Assembly," (2000) Nature 405 (6787) 665-668. Furthermore, Reiss et at., "Biological Routes to Metal Alloy Ferromagnetic

Nanostructures" Nanoletters, 2004, Vol. 4, No. 6, 1127-1132 describes peptides for binding to metals, including mediating nanoparticle synthesis. Flynn, et ai, "Synthesis and Organization of Nanoscale U-Vl semiconductor materials using evolved peptide specificity and viral capsid assembly," J. Mater. Sci., 2003, 13, 2414-2421 , describes peptides for binding to and nucleation of semiconductor nanoparticles. Mao, Flynn et al., "Viral Assembly of Oriented Quantum Dot Nanowires," PNAS, June 10, 2003, vol. 100, no. 12, 6946-6951 further describes peptides for binding to and nucleation of semiconductor nanoparticles. All of the above references are hereby incorporated by reference in their entireties.

3. Fluorescent particles bound to a multifunctional scaffold

In certain embodiments, pre-made fluorescent particles can be bound to the multifunctional scaffold through, for example, the formation of a covalent bond, sequence-specific binding affinity, or through the formation of a recognition pair. In certain embodiments, the fluorescent particles can be directly bound to the multifunctional scaffold. For example, a capturing group such as a cysteine can directly bind to the surface of a gold nanocluster, forming a stable Au-S covalent bond.

More typically, the fluorescent particles are surface functionalized for stable and/or selective binding to the multifunctional scaffold. These functionalities are stable when attached to the surface of the fluorescent particle, and are reactive to certain functional groups present on the multifunctional scaffold to form covalent bonds. For example, quantum dots can be functionalized to comprise a chemically reactive functionality that forms a covalent attachment with a functional group on the multifunctional scaffold. Similarly, fluorescent beads can comprise reactive functionalities, such as -COOH or -NH₂ on the surface of the beads.

For instance, a multifunctional scaffold is naturally rich in functional groups such as thiol, hydroxy, carboxyl and amino group. Typical chemically reactive functionalities on the surface of the fluorescent particles therefore include, but are not limited to: maleido (reacts with a thiol group), sulfo-N-hydroxysuccinimide (reacts with a primary amine), carboxyl group (reactive with an amine) and amino group (reacts with a carboxyl group). Optionally, a spacer group, such as low molecular weight polyethylene glycol (PEG) or a functionalized latex sphere, can be incorporated between the chemically reactive functionality and the surface of the fluorescent particle. Typically, the chemically reactive functionalities described above form stable bonds with a functional group on the multifunctional scaffold under mild conditions. In other embodiments, a cross-linker can be used to link the chemically reactive functionality with the functional group on the multifunctional scaffold. It is within the knowledge of one skilled in the art to select a suitable cross-linker based on the respective functional groups to be connected. As noted herein, the fluorescent particles can also be functionalized to comprise a first binding partner (ligand) that specifically recognize and bind to a second binding partner (anti-ligand) positioned on the multifunctional scaffold. Surface functionalized fluorescent particles can be prepared by known methods in the art. See, Klimov, V.I., (supra). They are also available from the commercial sources identified above, see, e.g., Qdot^® Streptavidin and Biotin Conjugates from Invitrogen (Carlsbad, CA).

In another embodiment, the fluorescent particles can be functionalized to comprise a ligand such as a biotin. Such a functionalized fluorescent particle can be bound to, for example, a streptavidin coated polymeric scaffold. In a further embodiment, fluorescent particles can be bound directly to a multimolecular scaffold based on a biological structure. As used herein, "biological structure" refers to an organic scaffold of a biological origin that contains multiple binding sites. More specifically, the multiple binding sites are situated on the outer walls of the biological structure in segments of oligomeric sequences, e.g., peptides. The sequences responsible for the binding are preferably linked to the genes within the biological structure. Examples of suitable biological structures include, but are not limited to cells, phage, viruses, yeasts, self-assembled peptide structures, and proteins.

The outer walls of these biological structures contain multiple binding sites that render them ideal candidates as multifunctional scaffolds. For example, some cell walls contain intrinsic amines or can be derivatized to contain thiol moieties. As mentioned above, these functional groups can be coupled to a fluorescent particle (e.g., surface-fuπctionalized fluorescent particles) in a variety of known methods. In another example, the cell walls may be labeled with a biotin, which specifically recognizes and binds to a fluorescent particles functionalized with streptavidin. In yet another example, peptides can be expressed on the surface of the outer walls of these biological structures. The biological structure (e.g., a virus) can be engineered to express specific and defined sequences that exhibit desired binding characteristics. U.S. Patent Application Publication Nos. 2005/0221083, 2006/0003387 describe in further detail of the use of biological structures such as virus and yeast as organic multifunctional scaffolds, which applications are incorporated herein by references in their entirety.

It is particularly desirable to manufacture specially designed and discrete metallic or semiconductor nanoclusters in which the emission spectra will depend on the size and shape of the nanoclusters. See, e.g., Alivisatos, P. (2004) Nature Biotechnology 22:1 , 47-52. Genetic engineering provides a powerful tool to display the phage binding sequences that bind to the desirable target material.

Biological structures such as phage have been shown to nucleate size-constrained crystalline semiconductor materials, and to control the crystallographic phase of nucleated nanoparticles. See, e.g., Mao, et a/. (2003) PNAS (supra), and Flynn, et al., (2003) J. Mater. Sci. (supra). Moreover, the aspect ratio of the nanoparticles can be controlled and, therefore, so can the electrical, magnetic, and optical properties. Accordingly, as an alternative to the direct conjugation of pre-made quantum dots to a multifunctional scaffold, fluorescent quantum dots can be prepared and bound to a multifunctional scaffold based on a nucleation process.

In certain embodiments, both semiconductor and metallic quantum dots can be formed through a nucleation process in the presence of a multifunctional scaffold. More specifically, a metallic (or semiconductor) precursor material in a solution can be nucleated onto the multifunctional scaffold to form a nanocluster of a certain size for the quantum effect to take effect.

In particular, a multifunctional scaffold can be selected or designed to have metal-specific binding sequences positioned in such a way that the in situ generated elemental metal will be brought into close proximity of each other to form a metallic nanocluster. For example, metallic clusters can be formed by reducing a metal salt in the presence of a multifunctional scaffold. In one example, silver bromide salt can be slowly reduced in the presence of an oligonucleotide and form silver nanocluster on the oligonucleotide. See, e.g., Petty, J.T. etal., (2004) J. Am. Chem.Soc. 126:16, 5207-52013. See, also, Bertini, I. et al., (2000) Eur. J. Biochem. 267, 1008-1018, describing Cu nanoclusters formed in the presence of a peptide, i.e., Cu7 metallothionein. For semiconductor quantum dots, a precursor solution can comprise the salts of each components of the quantum dot composition. For example, CdS quantum dots can be formed in the presence of peptide SEQ ID. 5 from a solution of CdCb and Na₂S. Detailed description of such nucleation processes are described in, e.g., Mao, C.B., et al., (2004) Science, 303, 213-217, Mao, et al. (2003) PNAS (supra), and Flynn et al., (2003) J. Mater. ScL (supra), which references are incorporated herein by reference in their entireties.

According to this method, the formation of the nanoclusters and the binding of the thus-formed nanoclusters to the multifunctional scaffold take place contemporaneously. Advantageously, the nucleation process can be controlled to form nanoclusters of highly oriented crystalline structure. The biological structures bound with fluorescent particles can be photoselected to have narrow or broad excitation and emission profiles, This allows either discrete excitation and emission properties or a large flux of emitted photons over a broad range of wavelengths. In some embodiments, a range of nanocluster sizes is created along the biological structure, reflecting various excitation and emission spectra discrete to each size. In this case, different fractions of these nanoclusters could be selectively excited and their emission collected. In other embodiments, only discrete sizes of nanoclusters are formed along the same biological structure. This means they will exhibit narrower bands of excitation and emission spectra but with a larger flux of emitted photons of the same wavelength. In other embodiments, the quantum dots are initially encapsulated in a matrix prior to binding to the multifunctional scaffold. A matrix refers to a stabilizing structure that encapsulates a nanocluster of metal atoms or semiconductor atoms. In one embodiment, the matrix is necessary to impart a desired physical property such as hydrophilicity. Examples of the encapsulating matrix include, but are not limited to, a dendrimer, a star polymer and a micelle.

A dendrimer refers to regularly branched, highly monodispersed polymers with a well-defined molecular architecture consisting of a core, regularly branching repeat units, and terminal groups. Polyamidoamine (PAMAM) dendrimers are synthesized from an ethylenediamine core with branching units containing tertiary amine and amide functionality. Full generation (G1 , G2, etc.) PAMAM dendrimers are terminated with primary amine groups, while half generation (G1.5, G2.5, etc.) PAMAM dendrimers have terminal carboxylate groups. The terminating amine or carboxylate groups can be further functionalized to modulate the physical and chemical properties of the dendrimers. It has been described that silver salts (e.g., AgNOs) can be reduced to elemental silver and forms silver nanoclusters entrapped in a G4 or G2 PAMAM. See, Dickson, R. M., et al., (2002) J. Am. Chem. Soc. (Supra). Similarly, gold salt (HAICI₄) can be reduced in the presence of PAMAM to produce gold nanoclusters of discrete sizes including Au₅, Au₈, Aui3, Au₂3 and Au₃i- See, Dickson, R.M. et al., (2004) Physical Rev. Lett. (Supra).

Dendrimers encapsulating fluorescent nanoparticles can therefore be bound, through one of its surface functionalities, to a capturing group on the multifunctional scaffold. !f desired, a cross-linker can be used. For example, an amino group of the PAMAM can be coupled to an amino capturing group via a cross-linker having dicarboxylic acid, e.g., 1 ,4-dicarboxylic acid butane. Alternatively, a hydroxy group of the PAMAM can be coupled to a thiol- capturing group via N-(maleimidophenyl)isocyanate. Advantageously, quantum dots of discrete sizes encapsulated in the dendrimer can be separated according to their sizes by centrifugation, filtration or other means prior to being bound to the multifunctional scaffold. It is therefore more likely to form taggants with improved monodispersity.

Alternatively, one or more dendrimers can be initially bound to a multifunctional scaffold followed by forming and encapsulating a nanocluster of the metal atoms.

A star polymer has a similar branching structure to a dendrimer, and can be in fact built upon a dendrimer core by adding branches of polyethylene glycol. It has been shown that CdS quantum dots can be formed and entrapped in a star polymer out of a solution of Cd²⁺ and S^2". See, e.g., Smith, A. P. et al., (2002) NIST Technipuhs, polymer division, 854.

Other encapsulating matrix such as micelles can be used to enhance the water solubility of quantum dots. See, e.g., Dubertret et al., (2002) Science 298, 1759-1762. As used herein, "micelle" refers to a unit of structure composed of an aggregate or oriented arrangement of amphiphilic molecules. An amphiphilic molecule typically comprises a polar (or hydrophilic) end and a hydrophobic portion (e.g., a hydrocarbon chain). Micelles can be formed in an aqueous solution when the amphiphilic molecules aggregate such that their polar ends are in contact with water and their hydrophobic portions are in the interior of the aggregate. The interior of the micelle is therefore hospitable to encapsulate the normally hydrophobic quantum dots. Typical micelle-forming amphiphilic molecules include but are not limited to: surfactants and amphiphilic diblock copolymers (e.g., poly(lactide-co-ethylene glycol)).

Pre-made quantum dots can be initially encapsulated into the micelles followed by conjugating the micelles to the multifunctional scaffold. It is understood that the micelles are stable structures, the structural integrity of which is maintained during the conjugation process.

4. Use of the taggants

As discussed herein, the taggant composition can bind to more than one fluorescent particle, which thus leads to a more intense fluorescence signal. Moreover, the taggant composition provides better environmental stability than colloidal quantum dots owing to one or more binding sites on the multifunctional scaffold having an affinity for the substrate. These attributes make the taggant composition suitable candidates as sensitive and versatile fluorescent markers. Because the fluorescent particles bound to the multifunctional scaffold can be controlled and modulated to exhibit specific spectroscopic properties, the taggant composition can be used to provide a means to store information on a surface of a substrate, thereby distinguishing a valid article or identity from invalid ones. Advantageously, the multifunctional scaffold has an affinity for the surface, which ensures the environmental stability of the fluorescent particles adhered thereto. The stability can be further enhanced by a fixation process. It is well-known to one skilled in the art that the use of cross-linking agents, e.g. formaldehyde and glutaraldehyde, or dehydrating/denaturing agents, e.g. acetone and ethanol, can improve the adhesion of proteins to substrates. This is a standard process to make cells more stable on microscope slides (See, e.g., Leong, A., Fixation, Woods and Ellis, Laboratory Histopathology: A Complete Reference, 1994 Edinburgh; Churchill Livingstone). Typically, a fixation agent such as acetone and ethanol can be applied, e.g., by spraying, onto a film of biomolecule-based taggants deposited onto a substrate. The improved environmental stability of these taggants can be assessed by measuring the retention of the optical response of the taggants after multiple washing or mechanical abrasion steps. A tape test, which is a standard method for measuring adhesion or resistance to abrasion of a coating on a substrate, can be employed. Typically, an adhesive tape, such as 3M Scotch® 600 tape, can be used to produce the abrasion. The removal of the taggants can be readily evaluated by the loss of the optical response after each application of the adhesive tape. Conversely, the retention of the optical response is correlated to the fraction of the taggants that are firmly bound to the substrate and therefore resistant to the abrasion.

Figure 3 shows the results of a series of tape tests conducted on taggant layers deposited on a glass substrate. The taggant layers were sprayed with different fixation agents, including acetone, ethanol and isopropyl alcohol (IPA). In the control test, no fixation agent was used. As shown in Figure 3, the taggant layers sprayed with acetone and ethanol present better environmental stability than the control test or the IPA-sprayed test. In all four tests, the optical response reached an asymptotic limit, following an initial drop in the optical response after a first application of the adhesive tape.

In one embodiment, the taggants can be added to explosives, plastics or other substrate to indicate their sources, identity or authenticity.

In other embodiment, the taggants can be formulated with a carrier material, such as ink, paint, an adhesive, a binder, etc. The formulation can be sprayed, printed, pressed, or otherwise affixed to the substrate of interest. For example, the taggants can be mixed with an ultraviolet-curable ink composition for anti-counterfeit applications.

In one embodiment, it is described herein a security device comprising: a substrate; and an invisible image indicia comprising a plurality of taggants positioned on the substrate, each taggant comprising one or more nanoparticles bound to a multifunctional scaffold, the nanoparticles emitting light in a range of wavelength from about 400nm to about 1400nm when exposed to an excitation energy, the multifunctional scaffold further including one or more first binding sites having an affinity for the substrate, wherein the invisible image indicia is substantially invisible under an illumination of light within a visible range but fluoresces in a detectable way at visible or IR wavelengths when exposed to the excitation energy. In another embodiment, the invisible image indicia displays a signature image of a fluorescence pattern when exposed to an excitation energy. As used herein, the term "signature image" refers to a unique image of a distribution of fluorescent dots. The distribution can be random or controlled. The fluorescent dots can vary in their physical sizes and spectroscopic properties (e.g., color). The signature image draws its uniqueness from the limitless possible arrangements of the taggants in a given area, as will be discussed further in detail below.

As used herein, the term "invisible image indicia" refers to a marker that is invisible under an illumination of light within a visible range and only readable upon exposure to an excitation energy. For example, an "invisible" image indicia can reveals a unique signature image under an ultraviolet light source. The invisible image indicia can therefore be read by a reading device and tracked to, for example, the source or authenticity of an article to which the indicia is affixed. An embodiment is illustrated in Figure 4, in which a security device 48 comprises invisible image indicia 50 present on the substrate 24, the invisible image indicia being only visible under illumination with ultraviolet light. The invisible image indicia 50 comprise randomly distributed taggants 10. Depending on the specific physical distribution of the taggants and the color emitted from each taggant, a fluorescence pattern 54 can be revealed under the ultraviolet light. In Figure 4, the fluorescence pattern can be random until it is stored as a readable image file in a database and can be optionally associated with a unique identification number. The invisible image indicia can therefore be used as an anti-counterfeiting measure for brand protection or authenticating government issued papers or documents. Because the typical size of the taggants is on the order of nanometer-scale or micron-scale, a surface can hold as many as 2.5 million taggants per square microns {i.e., at 10nm dimension). Assuming the taggant can be prepared in 20 or more distinct sizes either by precision control of growth or by physical separation methods, approximately 40,000,000,000 bits per square centimeters can be yielded. The virtually limitless diversity and randomness of the fluorescence patterns generated is therefore impossible for potential counterfeiters to reproduce.

Optionally, the identification number can be encrypted by known methods in the art. Moreover, it can also be associated with a descriptive message indicating the source of the article.

In another embodiment, the fluorescence pattern generated is not random but an immediately identifiable image, such as a sign, a word, a symbol or a code. As illustrated in Figure 5, a security device 58 comprises the taggants 10 deposited on the substrate 24 according to a selected pattern 60 (e.g., the letters I and D). The pattern is invisible until being excited by an ultraviolet light. The security device is particularly useful in the context of defense. For example, the security device can be incorporated into the fabric of armor. The reading device can be equipped in goggles or surveillance system of tanks and airplanes to instantly distinguish friends from foes.

In a further embodiment, it is described herein a security system comprising a security device including an invisible image indicia, the invisible image indicia having a plurality of taggants positioned on a substrate, each taggant comprising one or more fluorescent particles bound to a multifunctional scaffold, wherein the invisible image indicia is substantially invisible under an illumination of light within a visible range but displays a signature image of a fluorescence pattern when exposed to an excitation energy; a database storing the signature image in a readable format; and a reading device for reading the invisible image indicia. An example of the security system is illustrated in Figure 6, in which a security device 58 is affixed to an article 62. The security device 58 includes an invisible image indicia (not shown). Upon exposure to an excitation energy, the security device 58 displays a signature image 64 of a fluorescence pattern. The signature image 64 can be read by a reading device 66. As would be understood by one of ordinary skill in the art, the reading device can be an optical detector, as well as a thermographic detector. The signature image 64 can thereafter be stored in a readable format in a database 67.

Due to its uniqueness, a signature image can be associated with an article to provide an authentication tool. In a related use, it is described herein a method of authenticating an article, comprising: marking an authentic article with an invisible image indicia, the invisible image indicia having a plurality of taggants bound to a surface of the authentic article, each taggant comprising one or more fluorescent particles bound to a multifunctional scaffold, wherein the invisible image indicia is substantially invisible under an illumination of light within a visible range but displays a signature image of a fluorescence pattern when exposed to an excitation energy; irradiating the invisible image indicia to display a signature image of a fluorescence pattern; storing the signature image in a readable format in a database; irradiating the article to be authenticated; detecting the presence of an image of a fluorescent pattern; and matching the image to the signature image of the authentic article in the database.

In a further embodiment, it is described herein a method for the control or monitoring of specific binding events. In this embodiment, the taggants serve as an internal label to indicate successful binding between the multifunctional scaffold and the substrate. As illustrated in Figure 7, a plurality of the taggants 40 are deposited on the substrate 24 in a pre-selected pattern 60 (e.g., a cross), the taggants being bound to the substrate 24 via the first binding site 28. Upon exposure to an excitation energy, such as ultraviolet light, any defects (non-binding of the substrate) 68 can be visualized (e.g., substrate 24a). For substrates that are free of defects, as can be visualized on a substrate 24b, an optional step of plating can be carried out. More specifically, the multifunctional scaffold 18 of each of the taggant 40 in the layer 60 acts as a template to bind to a target material 44. In one embodiment, the target material 44 is a seed material on to which the layer 70 can be plated. Interestingly, if the target material 44 is also a fluorescent nanoparticle, such as the fluorescent nanoparticle 14, the taggants 40 can serve as an internal control for the plating process. As the plating progresses, the fluorescence signal is expected to diminish as the fluorescent particles are either encapsulated by non-fluorescing material, or loses fluorescence as their sizes grow beyond the dimensional restraints for the "quantum confinement" to take effect. Such diminishing signal can also be monitored as a means to monitor successful plating.

It is noted that in all of the above embodiments, while the multifunctional scaffolds may be present during deposition on the substrate, they may be later removed leaving behind only the nanoparticles on the substrate. Typically, the multifunctional scaffolds can be removed from the material they are bound to by thermal annealing or sintering. U.S. Patent Application No. 10,976,179 and Mao et al. (2004) Science, 300, 213-217 describe in detail the techniques of burning biomolecules scaffolds off; both are incorporated herein by reference in their entireties.

Finally, it is clear that numerous variations and modifications may be made to method and apparatus described and illustrated herein, all falling within the scope of the invention as defined in the attached claims.

All of the above U.S. patents, U.S. patent application publications, U.S. patent applications, foreign patents, foreign patent applications and non- patent publications referred to in this specification and/or listed in the Application Data Sheet, are incorporated herein by reference, in their entirety.

Claims

1. A composition comprising: a multifunctional scaffold including one or more first binding sites having a binding affinity for a surface of a substrate; and one or more fluorescent particles bound to the multifunctional scaffold, wherein the fluorescent particles emit light in a range of wavelength from about 400nm to about 1400nm when exposed to an excitation energy.

2. The composition of claim 1 wherein the fluorescent particles are metallic quantum dots that display size-dependent fluorescence emissions upon irradiation with light.

3. The composition of claim 2 wherein the metallic quantum dots are nanoclusters of gold, silver, or copper atoms.

4. The composition of claim 3 wherein each nanocluster has no more than 40 metal atoms.

5. The composition of claim 3 wherein each nanocluster is no more than 4nm in diameter.

6. The composition of claim 2 wherein the metallic quantum dots are encapsulated in a matrix, the matrix being bound to the multifunctional scaffold.

7. The composition of claim 6 wherein the matrix is a dendrimer, a star polymer or a micelle.

8. The composition of claim 1 wherein the fluorescent particles are semiconductor quantum dots that display size-dependent fluorescence emissions upon irradiation with light.

9. The composition of claim 8 wherein the semiconductor quantum dots are CdSe, CdS₁ CdTe, CdSe/ZnS, CdTe/ZnS or CdTe/ZnS.

10. The composition of claim 1 wherein the fluorescent particles are polymer-based fluorescent beads.

11. The composition of claim 1 wherein the multifunctional scaffold is a biomolecule, a protein, a self-assembled peptide structure, a cell, a phage, a yeast, a virus or a polymer bead.

12. The composition of claim 11 wherein the multifunctional scaffold includes one or more capturing groups that bind the respective one or more fluorescent particles.

13. The composition of claim 12 wherein the capturing group is a functional group selected from thiol, hydroxy and amino group.

14. The composition of claim 12 wherein the capturing group is a peptide sequence having an affinity for the fluorescent particle.

15. The composition of claim 14 wherein the peptide sequence is selected from the group consisting of SEQ ID NO: 1-24.

16. The composition of claim 14 wherein the peptide sequence nucleates the fluorescent particle out of a solution by converting a precursor material to the fluorescent particle.

17. The composition of claim 11 wherein the capturing group is a first binding partner that binds to a fluorescent particle surface functionalized with a second binding partner, the first binding partner and the second binding partner forming a recognition pair.

18. The composition of claim 17 wherein the first binding partner is a biotin and the second binding partner is streptavidin.

19. The composition of claim 11 wherein the first binding site includes one or more peptide sequences that non-covalently bind to the surface of the substrate.

20. The composition of claim 19 wherein the multifunctional scaffold further comprises an adhesive group that covalently binds to the substrate.

21. The composition of claim 20 wherein the multifunctional scaffold comprises an amino acid, the amino acid being converted to the adhesive group.

22. The composition of claim 21 wherein the amino acid is tyrosine and the adhesive group is ι_-3,4-dihydroxyphenylanaline.

23. The composition of claim 19 wherein the multifunctional scaffold further comprises one or more second binding sites having an affinity for a functional material.

24. The composition of claim 23 wherein the functional material is a conductive material, a dielectric material, a semiconductor material or a magnetic material.

25. A security device comprising a substrate; and an invisible image indicia comprising a plurality of taggants positioned on the substrate, each taggant comprising one or more fluorescent particles bound to a multifunctional scaffold, the fluorescent particles emitting light in a range of wavelength from about 400nm to about 1400nm when exposed to an excitation energy, the multifunctional scaffold further including one or more binding sites bound to the substrate, wherein the invisible image indicia is substantially invisible when exposed to visible light but emits detectable fluorescence when exposed to the excitation energy.

26. The security device of claim 25 wherein the invisible image indicia displays a signature image of a fluorescent pattern when exposed to the excitation energy.

27. The security device of claim 25 wherein the plurality of the taggants display fluorescence emissions depending on the size of the fluorescent particles.

28. The security device of claim 25 wherein the plurality of the taggants have a random assortment of sizes and shapes.

29. The security device of claim 25 wherein the fluorescent particles are metallic quantum dots, semiconductor quantum dots or fluorescent beads.

30. The security device of claim 25 wherein the multifunctional scaffold is a biomolecule, a protein, a self-assembled peptide structure, a cell, a phage, a yeast, a virus or a polymer bead.

31. The security device of claim 25 wherein the signature image is an immediately identifiable image.

32. The security device of claim 25 wherein the signature image is identifiable by matching the signature image with a previously stored image.

33 A method of forming a security device, comprising: depositing a plurality of taggants on a substrate to form an invisible image indicia, each taggant comprising one or more fluorescent particles bound to a multifunctional scaffold, the multifunctional scaffold including one or more binding sites bound to the substrate, wherein the fluorescent particles emit light in a range of wavelength from about 400nm to about 1400nm when exposed to an excitation energy; irradiating the invisible image indicia to display a signature image of a fluorescence pattern; and storing the signature image in a readable format in a database.

34. The method of claim 33 wherein the depositing step comprises randomly distributing the taggants on the substrate, the taggant having fluorescent particles of a random assortment of sizes and shapes.

35. The method of claim 34 wherein the signature image encodes a pattern of distribution and properties of the fluorescent particles.

36. The method of claim 35 further comprising assigning an identification code to the signature image.

37. The method of claim 36 further comprising encrypting the identification code.

38. The method of claim 37 further comprising associating a descriptive message with the identification code.

39. The method of claim 33 wherein the fluorescent particles are metallic quantum dots, semiconductor quantum dots or fluorescent beads.

40. The method of claim 33 wherein the substrate is a semiconductor, metal, glass, ceramic, polymer, fabric, paper or composite material.

41. The method of claim 33 further comprising applying a fixation agent to the taggants deposited on the substrate.

42. The method of claim 41 wherein the applying step comprises spraying acetone or ethanol to the taggants deposited on the substrate.

43. The method of claim 33 further comprising removing the multifunctional scaffolds.

44. A security system comprising: a security device comprising an invisible image indicia, the invisible image indicia having a plurality of taggants positioned on a substrate, each taggant comprising one or more fluorescent particles bound to a multifunctional scaffold, the multifunctional scaffold including one or more binding sites bound to the substrate, wherein the invisible image indicia is substantially invisible when exposed to light within a visible range but displays a signature image of a fluorescence pattern when exposed to an excitation energy; a database storing the signature image in a readable format; and a reading device for reading the invisible image indicia.

45. The security system of claim 44 wherein the excitation energy is ultraviolet light.

46. The security system of claim 44 wherein the Invisible image indicia encodes a pattern of distribution and properties of the fluorescent particles.

47. A method of authenticating an article, comprising: marking an authentic article with an invisible image indicia, the invisible image indicia having a plurality of taggants bound to a surface of the authentic article, each taggant comprising one or more fluorescent particles bound to a multifunctional scaffold, the multifunctional scaffold further including one or more binding sites bound to the surface, wherein the invisible image indicia emits light in a range of wavelength from about 400nm to about 1400nm and displays a signature image of a fluorescence pattern when exposed to an excitation energy; irradiating the invisible image indicia to display a signature image of a fluorescence pattern; storing the signature image in a readable format in a database; irradiating the article to be authenticated; detecting the presence of an image of a fluorescent pattern; and matching the image to the signature image of the authentic article in the database.

48. The method of claim 47 wherein the signature image is unique.

49. The method of claim 48 wherein the signature image is associated with an identification code.

50. A method comprising: encapsulating a nanocluster of metal atoms in a matrix; and conjugating the matrix to a multifunctional scaffold, the multifunctional scaffold having a binding affinity for a substrate.

51. The method of claim 50 wherein the encapsulating step comprises converting a precursor of the metal to the metal atoms in the presence of the matrix.

52. The method of claim 51 wherein the converting step comprises reducing a metal salt to the metal.

53. The method of claim 50 further comprising a step of controlling a size of the nanocluster encapsulated by adjusting the relative amounts of the matrix and the precursor of the metal.

54. The method of claim 50 wherein the metal is gold, silver or copper.

55. The method of claim 50 wherein the conjugating step comprises covalently linking the matrix to the multifunctional scaffold via a cross-linker.

56. The method of claim 50 wherein the matrix is a dend rimer, a star polymer or a micelle.

57. The method of claim 50 wherein the multifunctional scaffold is a protein, a self-assembled peptide structure, biological cell, phage, virus, bacteria, peptide, oligonucleotide, antibody, block copolypeptide or amphiphilic lipopeptide.

58. The method of claim 50 wherein the nanocluster comprises no more than 40 metal atoms.

59. The method of claim 50 wherein the nanocluster is no more than 4nm in diameter.

60. The method of claim 50 wherein the nanocluster displays a size-dependent fluorescence emission.

61. A method comprising: depositing a layer of taggants on a substrate according to a preselected pattern, each taggant comprising one or more fluorescent particles bound to a multifunctional scaffold, the fluorescent particles emitting light in a range of wavelength from about 400nm to about 1400nm when exposed to an excitation energy, the multifunctional scaffold further including one or more first binding sites bound to the substrate and one or more second binding sites having an affinity for a functional material; irradiating the substrate to display a fluorescent pattern, wherein the taggants act as an interna! control to indicate any non-binding of the multifunctional scaffold to the substrate; and binding a functional material to the multifunctional scaffold via the second binding site thereof to form a functional material layer in the preselected pattern.

62. The method of claim 61 wherein the taggants comprises metallic quantum dots, semiconductor quantum dots or fluorescent beads.

63. The method of claim 61 wherein the multifunctional scaffold is a biomolecule, a protein, a self-assembled peptide structure, a cell, a phage, a yeast, a virus or a polymer bead.

64. The method of claim 63 wherein the second binding site of the multifunctional scaffold binds to a pre-made functional material.

65. The method of claim 63 wherein the second binding site of the multifunctional scaffold nucleates the functional material from a solution comprising a precursor of the functional material using a seed layer.

66. The method of claim 65 wherein the seed layer is formed of a plurality of nanoparticles.

67. The method of claim 66 wherein the nanoparticles are fluorescent particles.

68. The method of claim 61 wherein the functional material is a conductive material, a dielectric material, a semiconductor material or a magnetic material.

69. The method of claim 68 further comprising irradiating the seed layer to indicate any non-binding to the functional material.