Nothing Special   »   [go: up one dir, main page]

WO1994002830A1 - Analyse de cellules - Google Patents

Analyse de cellules Download PDF

Info

Publication number
WO1994002830A1
WO1994002830A1 PCT/GB1993/001561 GB9301561W WO9402830A1 WO 1994002830 A1 WO1994002830 A1 WO 1994002830A1 GB 9301561 W GB9301561 W GB 9301561W WO 9402830 A1 WO9402830 A1 WO 9402830A1
Authority
WO
WIPO (PCT)
Prior art keywords
cell
antibody
labelled
process according
cells
Prior art date
Application number
PCT/GB1993/001561
Other languages
English (en)
Inventor
Melvyn Greaves
Catherine Mary Price
Susan Colman
Original Assignee
The Institute Of Cancer Research
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by The Institute Of Cancer Research filed Critical The Institute Of Cancer Research
Publication of WO1994002830A1 publication Critical patent/WO1994002830A1/fr

Links

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
    • G01N33/56966—Animal cells
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6813—Hybridisation assays
    • C12Q1/6841—In situ hybridisation
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
    • G01N33/56966—Animal cells
    • G01N33/56972—White blood cells

Definitions

  • the present invention relates to the genetic screening of cells, eg cells of cancer (including leukemic) or pre-natal (fetal) lineages.
  • the correlation of cell type and chromosomal abnormality can provide additional information relating to lineage involvement and diagnosis in leukaemia and the developmental level of the clonal leukemic cells which would be the target for any therapy.
  • Kibbelaar et al (Blood, 29(7); 1823-1828, (1992)) have found that a cell can be stained in solution with an antibody probe attached to a fluorescent label and that a cell thus stained can be further stained with a DNA probe attached to a different fluorescent label, allowing simultaneous identification of a phenotype and a genotype.
  • Kibbelaar has a number of disadvantages.
  • the procedures involved in contacting cells with antibodies and conducting in situ hybridization with DNA probes are both tasks which require technical expertise. Such expertise is not always available in pathology laboratories who perform only routine analysis of cells of patients, but rely on other, fewer, centres for more complicated analysis of such samples.
  • the technique of Kibbelaar must be performed on fresh cell samples because the cells are contacted with the labelled antibody in solution.
  • a further disadvantage of the need to perform rapid analysis of cells is that this does not allow for historical analysis of the cells at a later date. For example, it may be desirable to determine the results of a first phenotyping experiment before deciding which antibodies should be used on a second phenotyping experiment. It may also be the case that as new antibodies become available and the understanding of diseases increases, information can be obtained from samples taken from patients who have long since died.
  • Kibbelaar et al show that cells can be analyzed by combined immunotyping and genotyping, the method disclosed has limited practical application as a routine clinical technique.
  • the present invention thus provides a process for the analysis of a cell which comprises (i) contacting a cell with an antibody, which antibody is labelled, either before or after being brought into contact with the cell, with a colorimetric label; (ii) contacting the cell with a DNA probe, which DNA probe is labelled, either before or after being brought into contact with the cell, with a different colorimetric label, characterised in that the cell is fixed to a solid support prior to labelling.
  • the cell sample can be stored for a considerable length of time prior to conducting the reaction with the antibody and DNA probes. This would allow the cells to be obtained in any pathology laboratory and subsequently analyzed in a specialist analytical laboratory elsewhere where there is the requisite level of
  • SUBSTITUTE SHEET expertise at cell labelling techniques This would allow the cells to be analyzed one or two weeks, or a month, or even a year or more after they have been removed from a patient. It would also allow a first sample of cells to be analysed and a second sample, taken at the same time, to be analysed with different antibody and/or DNA markers which are selected in the light of the result of the first analysis.
  • a further advantage of the present invention is that by fixing the cell on a solid support, the cell membrane is made permeable, which allows the contents of the cell cytoplasm to be available for labelling. This will increase the signal obtained from cells positive for the marker being analyzed, which will help distinguish such positive cells from mere background signals.
  • colorimetric refers to two types of label. These are histochemical markers and fluorescent markers. Histochemical stains are permanent whereas, as described above, fluorescent stains fade after relatively short periods of time.
  • alkaline phosphatase-Fast Red is also capable of fluorescing but does not, unlike conventional fluorescent stains, fade.
  • the present invention thus further provides a process for the phenotyping and genotyping of a cell which comprises (i) contacting the cell with an antibody, which antibody is labelled, either before or after being brought into contact with the cell, with a colorimetric label; (ii) contacting the cell with a DNA probe, which DNA probe is labelled, either before or after being brought into contact with the cell, with a different colorimetric label, characterised in that the colorimetric label used to label the antibody is a histochemical stain capable of fluorescing.
  • the histochemical stain is alkaline phosphatase-Fast
  • SUBSTITUTESHEET Red Other histochemical stains include fuschin.
  • the cells can be stored after step (i) above for any desired length of time prior to staining with a DNA probe.
  • the phenotyping and genotyping of cells is conducted in a process which comprises (i) fixing a cell to a solid support; (ii) contacting the cell with an antibody, which antibody is labelled, either before or after being brought into contact with the cell, with a histochemical stain which stain is capable of fluorescing; (iii) contacting the cell with a DNA probe, which DNA probe is labelled, either before or after being brought into contact with the cell, with a colorimetric label which fluoresces at a different wavelength from the histochemical stain.
  • the present invention also provides a method for the phenotyping and genotyping of a cell fixed to a solid support which comprises (i) contacting the cell with an antibody, which antibody is labelled, either before or after being brought into contact with the cell, with a histochemical stain which stain is capable of fluorescing; (ii) contacting the cell with a DNA probe, which DNA probe is labelled, either before or after being
  • SUBSTITUTESHEET brought into contact with the cell, with a colorimetric label which fluoresces at a different wavelength from the histochemical stain.
  • the steps of contacting the antibody to the cell sample, and labelling the antibody are known in the art per se.
  • an unlabelled antibody produced in a first species eg mouse
  • a second antibody against the antibody of the first species is used to detect the first antibody, and this second antibody is either labelled, or capable of being labelled by a colorimetric stain. Variations on this are known in the art and can be used in the present invention.
  • a DNA probe may be labelled with a first marker, suitably biotin.
  • the first marker can be detected with a further probe, eg. fluoresceinated avidin. Variations of this are again known in the art (eg amplification of the avidin by biotinylated anti-avidin) and may be used in the present invention.
  • DNA probe means a probe capable of detecting genomic DNA in a cell.
  • the probe will usually comprise DNA.
  • the probe may also comprise modified bases or even comprise RNA.
  • the probe will usually comprise at least one base which is suitably labelled, eg with biotin.
  • the precise nature of the DNA probe may be determined by those of skill in the art, taking into account the nature of the target DNA and reagents available for the probe target.
  • the cell is contacted with the antibody prior to being contacted with the DNA.
  • the contacted cell is usually exposed to radiation of a wavelength
  • SUBSTITUTESHEET that will cause both the labelled antibody and the labelled probe to fluoresce.
  • the result of the fluorescence may be recorded in any manner desired, eg by photography or by digitisation and recordal in a computer.
  • the latter method may be advantageous when the two wavelengths at which the probe and antibody stains fluoresce appear similar to the naked eye, since the digitised record can be manipulated and represented by the recordal system in other ways, eg with "false colours".
  • the invention may be used to examine cells, eg human cells or other mammalian cells, in situations where information on cell lineage and genotype are required simultaneously. This is useful in the study of cancers, especially leukemias and preleukemic conditions, eg polycythemia vera. Such diseases are often manifested by a clonal lineage of cells which have an abnormal chromosome content (eg trisomies, deletions or translocations) and aberrant expression of cell markers. Information concerning the nature of such markers will assist the physician in the specific diagnosis of the disease, to monitor the progress of the disease and to develop improved therapies for the treatment of the disease, eg by the use of toxic antibodies directed against markers expressed by the leukemic cells, or by monitoring cell numbers after treatment with different drugs.
  • diseases are often manifested by a clonal lineage of cells which have an abnormal chromosome content (eg trisomies, deletions or translocations) and aberrant expression of cell markers. Information concerning the nature of such markers will assist the physician in the specific
  • the invention can also find utility in examining fetal cells for prenatal diagnosis.
  • chorionic villus sampling which can be conducted in the first trimester of pregnancy, is in theory preferable to amniocentesis as a method of prenatal diagnosis, which is conducted in the middle of the second trimester, since late termination of pregnancy (in the event of the detection of a genetic abnormality) is generally undesirable for both medical and social reasons.
  • CVS is a difficult procedure and may result in samples which contain a mixture of fetal and maternal cells. Such samples will be
  • SUBSTITUTESHEET difficult to analyze by conventional methods.
  • a fetal antibody marker and conduct an examination for genomic abnormalities, eg trisomy 21 or genetic defects inherited in a normal mendelian fashion. Examples of such defects include muscular dystrophy and cystic fibrosis.
  • Fetal nucleated erythrocytes have also been isolated from maternal blood during pregnancy using monoclonal antibodies and flow sorting 16 " 21 and have been detected as early as 11 weeks of gestational age 16,19,20 .
  • Analogous techniques may be used to obtain fetal cells from maternal blood using a Magnetic-Activated Cell Sorter (MACS) , using a suitable MALS column (eg available from Milteny Biotec GmbH) .
  • MALS Magnetic-Activated Cell Sorter
  • Trisomy 8 is a non-random chromosome abnormality in PV occurring in approximately 16% of karyotypically abnormal cases and in common with other chromosomal changes (eg. del(20q) and +9) it may occur alone or as a secondary event.
  • SUBSTITUTESHEET Patient 1 This 67 year old man presented in October 1991 with a scrotal hydrocele. He was noted to have a raised haemoglobin and further investigations, including the production of erythropoietin independent erythroid colony growth, confirmed the diagnosis of PV. Cytogenetic analysis showed a normal male karyotype and the presence of two hyperdiplid clones both with trisomy 8 in addition to other chromosomal abnormalities: 46,XY/47,XY,+8,+9, -12,t (6;12) 48,XY, +8,+9, 12,t (6;12) ,del (12q) . Peripheral blood samples were obtained from this patient at this time. Shortly after presentation he required emergency surgery for an ischaemic lower limb and sustained a fatal cerebro- vascular event post-operatively.
  • Peripheral blood samples from both patients and a normal female control were obtained by venesection and placed in preservative free heparin.
  • Whole peripheral blood was cultured from 2 to 48 hours and harvested for routine cytogenetic analysis. The remainder of the sample was centrifuged over Lymphoprep (Nycomed, Oslo, Norway) to obtain a mononuclear cell (MNC) fraction.
  • the MNC were used in hemopoietic colony assays,
  • MNC were cultured at a concentration of 2 x 10 5 /mL in Iscove's medium containing 40% fetal calf serum, 2% bovine serum albumin (BSA) , 10 "4 M 2-mercaptoethanol, 1% methylcellulose, 5 units of erythropoietin and 30 units GM-CSF. Cultures were incubated at 37oC in a humidified 5% C0 2 atmosphere for 14 days. Individual erythroid (BFU-E) and granulocyte/macrophage (CFU-GM) colonies were removed using a micropipiette, suspended in phosphate buffered saline (PBS) and cytocentrifuged onto glass microscope slides. The slides were either fixed in methanol and Giemsa stained to confirm morphological cell type or air dried, wrapped in foil and stored at -20oc for later staining and FISH.
  • BFU-E fetal calf serum
  • BSA bovine serum albumin
  • the slides were washed in a TBS for 20 minutes and 20 ⁇ L of the second layer, an unlabelled rabbit anti-mouse antibody (DAKO Ltd, High Wycombe, UK) , was added and the slides incubated for 45 minutes in the humid chamber.
  • the slides were washed in TBS as above. 20 ⁇ L of anti-rabbit APAAP
  • T-cells CD3 T28; a gift from
  • CD13 MY7; Coulter Electronics Ltd, Luton, UK
  • monocytes CD14 MY4 : a gift from Dr J Griffen
  • megakaryocytes CD41 J15; a gift from Dr A McMichael
  • Hematopoietic progenitors CD34 (QBEND10; Quantum Biosystems, Cambridge, UK)
  • Erythrocytes Anti-glycophorin A (AGA) R10; a gift from Dr P Edwards) .
  • Hybridization buffer 50% formamide/10% dextran sulphate in 2X SSC pH 7.0
  • biotin labelled probe [chromosome 8 specific alpha-satellite probe D8Z1
  • the cells were mounted in fluorescence antifade (Citifluor, Nottingham University, UK) or antifade containing the counterstains propidium iodide (3ng/ ⁇ L) or DAPI (diamidino-2-phenyl-indole dihydrochloride, 200ng/mL) (both Sigma, Poole, UK) .
  • Double labelled fluorescent cells were analysed using either a Zeiss photomicroscope III equipped with epifluorescence and a dual band-pass filter (Omega, Brattleboro, VT) or a Bio-Rad MRC-600 laser scanning confocal microscope equipped with akrypton/argon ion laser (488/568nm line excitation with dual channel, 522nm and 585nm, emission filters) . Cells viewed on the Zeiss photomicroscope were photographed on either a Zeiss photomicroscope III equipped with epifluorescence and a dual band-pass filter (Omega, Brattleboro, VT) or a Bio-Rad MRC-600 laser scanning confocal microscope equipped with akrypton/argon ion laser (488/568nm line excitation with dual channel, 522nm and 585nm, emission filters) . Cells viewed on the Zeiss photomicroscope were photographed on
  • the false negative rate for the probe was assessed by scoring 200 cells APAAP control slides (immunoglobulin sub-class specific) from the normal control (Table 1) . In common with other centromeric repeat probes the false negative rate (1 signal per cell) for the chromosome 8 probe is higher (2-5%) than the false positive rate (3 signals per cell) at less than 1% in the normal control. Scorable cells with no signal contributed to less than 0.5% of the total cells observed. Cells with three signals in APAAP control slides from patients 1 and 2 were observed in 21% and 71% respectively (Table 1) . In addition, patient 1 had a small percentage 3%) of cells with 4 signals.
  • Colony analysis Colonies from patient 1 and 2 showed the presence of the trisomy in both myeloid (1/2 and 13/13) and erythroid (3/3 and 3/3) hematopoetic progenitors (Table 2) .
  • the number of cells analysed per colony and the percentage of cells showing 1, 2 or 3 chromosome 8 specific signals are given in Table 2.
  • the hybridization efficiency in colony cytospin cells is reduced so that the. number of informative cells is lower in comparison with normal MNC cytospins.
  • the false negative rate for centromeric probes in this type of preparation is higher at 5-17% whereas the false positive rate remains similar to that observed in normal MNC cytospins (2-2.5%) .
  • This problem however, is offset by the clonal nature of the preparations whereby only a few cells in the colony need be scored to confirm the genotype.
  • si tu and immunophenotype analysis In si tu and immunophenotype analysis. In si tu results with the chromosome 8 probe on the immunophenotyped cells are given in Table 4. Erythroid (RIO) positive cells only could be scored for patient 2 and of these the majority (90%) had three signals confirming the involvement of the erytroid lineage in the neoplastic clone. An R10+ cell photographed before and after FISH illustrated the dual fluorescence signal obtained in antibody positive cells. B- and T-lymphocytes from patient 1 showed a normal two signal distribution of chromosome 8 whereas monocytes and granulocytes showed two separate populations; 63%- 76% with two signals and 21%-35% with three signals. A small proportion of CDllc+ cells (1%) had 4 signals.
  • RIO Erythroid
  • Example 1 shows it is possible to analyse both phenotype and genotype simultaneously in the same interphase cell. Two major advantages of this method are fist that it allows a direct cell
  • SUBSTITUTESHEET by cell analysis of lineage and chromosomal markers and negates the need for sequential photography followed by relocation of specific cells, and second that the number of analysable cells is high due to the absence of harsh pretreatment steps which damage cytoplasmic and cell surface features. This technique has been used successfully to show discordant lineage involvement of the neoplastic clone in the myeloproliferative disease polycythemia vera.
  • trisomy 8 In both patients studied the clonal marker (trisomy 8) was present in early myeloid (CFU-GM) and erythroid (BFU-E) cells. In patient 1 trisomy 8 was present in a subset of mature myeloid cells (CDllc ⁇ , CD13+, CD14+) but was absent in all B- and T- lymphocytes scored.
  • G-6-PD glucose-6-phosphate dehydrogenase
  • PGK phosphoglycerate kinase
  • HPRT hypoxanthine phospribosyl transferase
  • SUBSTITUTE SHEET for clonal selection in PV is not the lymphoid/myeloid stern cell but a myeloid lineage committed cell. Alteratively, the longevity of T-cells and the slow rate of new T-cell production in adults could obscure a clonal contribution to the lineage. Rare cases of ly phoblastic transformation of PV 10,11 as in T-cell blastic crisis of CML 12,14 indirectly implicate the lymphoid/myeloid stem cell.
  • Blood samples from 6 pregnant women were obtained and the blood enriched for fetal nucleated erythrocytes. Three blood samples from non-pregnant females were used as controls.
  • AGA An i glycopho in A
  • Jacobs P and Greaves MF Leuk. Res. 8:737, 1984.

Landscapes

  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Immunology (AREA)
  • Hematology (AREA)
  • Molecular Biology (AREA)
  • Cell Biology (AREA)
  • Urology & Nephrology (AREA)
  • Biomedical Technology (AREA)
  • Zoology (AREA)
  • Organic Chemistry (AREA)
  • Biotechnology (AREA)
  • Physics & Mathematics (AREA)
  • Microbiology (AREA)
  • Biochemistry (AREA)
  • Analytical Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Wood Science & Technology (AREA)
  • Tropical Medicine & Parasitology (AREA)
  • Virology (AREA)
  • Food Science & Technology (AREA)
  • Medicinal Chemistry (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Genetics & Genomics (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • General Engineering & Computer Science (AREA)
  • Biophysics (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

Un procédé qui permet de définir le phénotype et le génotype d'une cellule, comprend: i) la fixation d'une cellule sur un support solide, ii) la mise en contact de cette cellule avec un anticorps qui est marqué, avant ou après cette mise en contact, avec un colorant histochimique apte à la fluorescence, 3) la mise en contact de cette cellule avec une sonde à ADN qui est marquée, avant ou après cette mise en contact, avec un marqueur colorimétrique qui devient fluorescent à une longueur d'onde différente de celle concernant le colorant histochimique.
PCT/GB1993/001561 1992-07-23 1993-07-23 Analyse de cellules WO1994002830A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
GB9215710.6 1992-07-23
GB929215710A GB9215710D0 (en) 1992-07-23 1992-07-23 Analysis of cells

Publications (1)

Publication Number Publication Date
WO1994002830A1 true WO1994002830A1 (fr) 1994-02-03

Family

ID=10719211

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/GB1993/001561 WO1994002830A1 (fr) 1992-07-23 1993-07-23 Analyse de cellules

Country Status (2)

Country Link
GB (1) GB9215710D0 (fr)
WO (1) WO1994002830A1 (fr)

Cited By (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1998002577A1 (fr) * 1996-07-15 1998-01-22 The Children's Medical Center Corporation Proceder pour detecter conjointement des genes introduits et leurs produits
FR2751989A1 (fr) * 1996-08-01 1998-02-06 Soc D Rech Et De Dev En Activa Methode d'identification de cellules eucaryotes par fixation desdites cellules sur un support et support pour mettre en oeuvre ladite methode
WO1998052018A1 (fr) * 1997-05-14 1998-11-19 Applied Imaging Corporation Identification d'objets par formation d'images multiples
US6136540A (en) * 1994-10-03 2000-10-24 Ikonisys Inc. Automated fluorescence in situ hybridization detection of genetic abnormalities
US6259807B1 (en) 1997-05-14 2001-07-10 Applied Imaging Corp. Identification of objects of interest using multiple illumination schemes and finding overlap of features in corresponding multiple images
US6956695B2 (en) 2001-03-19 2005-10-18 Ikonisys, Inc. System and method for increasing the contrast of an image produced by an epifluorescence microscope
EP1991708A2 (fr) * 2006-03-13 2008-11-19 Ikonisys, Inc. Procédé permettant de combiner immunocoloration et hybridation en fluorescence in situ (fish) au moyen de fluorophores pouvant se lier de manière covalente
WO2010022332A1 (fr) 2008-08-22 2010-02-25 Ventana Medical Systems, Inc. Procédé pour la détection chromogénique d’au moins deux molécules cibles dans un unique échantillon
US11413296B2 (en) 2005-11-12 2022-08-16 The Regents Of The University Of California Viscous budesonide for the treatment of inflammatory diseases of the gastrointestinal tract

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4888278A (en) * 1985-10-22 1989-12-19 University Of Massachusetts Medical Center In-situ hybridization to detect nucleic acid sequences in morphologically intact cells
WO1992009704A1 (fr) * 1990-11-29 1992-06-11 Diagnostic Hybrids, Inc. Procede de detection et d'identification in situ de sequences d'acide nucleique

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4888278A (en) * 1985-10-22 1989-12-19 University Of Massachusetts Medical Center In-situ hybridization to detect nucleic acid sequences in morphologically intact cells
WO1992009704A1 (fr) * 1990-11-29 1992-06-11 Diagnostic Hybrids, Inc. Procede de detection et d'identification in situ de sequences d'acide nucleique

Non-Patent Citations (5)

* Cited by examiner, † Cited by third party
Title
C.M. VAN DER LOOS ET AL.: "Combined Immunohistochemistry and DNA in situ Hybridization in One Tissue Section", THE JOURNAL OF HISTOCHEMISTRY AND CYTOCHEMISTRY, vol. 37, no. 6, 1989, NEW YORK, NY, US, pages 938, XP000404472 *
D. PINKEL, T. STRAUME, AND J.W. GRAY: "Cytogenetic analysis using quantitative, high-sensitivity, fluorescence hybridization", PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF USA, vol. 83, no. 9, May 1986 (1986-05-01), WASHINGTON US, pages 2934 - 2938, XP002914775, DOI: doi:10.1073/pnas.83.9.2934 *
R.E. KIBBELAAR ET AL.: "Combined Immunophenotyping and DNA In Situ Hybridization to Study Lineage Involvement in Patients with Myelodysplastic Syndromes", BLOOD, vol. 79, no. 7, 1 April 1992 (1992-04-01), NEW YORK, NY, US, pages 1823 - 1828, XP000400795 *
R.E. KIBBELAAR ET AL.: "Detection of trisomy 8 in hematological disorders by in situ hybridization", CYTOGENETICS AND CELL GENETICS, vol. 56, no. 3-4, 1991, BASEL, CH, pages 132 - 136, XP000400788 *
TSIEH SUN ET AL.: "Comparison of Phenotyping and Genotyping of Lymphoid Neoplasms", JOURNAL OF CLINICAL LABORATORY ANALYSIS, vol. 3, no. 3, 1989, NEW YORK, NY, US, pages 156 - 162, XP000400787 *

Cited By (16)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6221607B1 (en) 1993-10-07 2001-04-24 Ikonisys Inc. Automated fluorescence in situ hybridization detection of genetic abnormalities
US6136540A (en) * 1994-10-03 2000-10-24 Ikonisys Inc. Automated fluorescence in situ hybridization detection of genetic abnormalities
WO1998002577A1 (fr) * 1996-07-15 1998-01-22 The Children's Medical Center Corporation Proceder pour detecter conjointement des genes introduits et leurs produits
FR2751989A1 (fr) * 1996-08-01 1998-02-06 Soc D Rech Et De Dev En Activa Methode d'identification de cellules eucaryotes par fixation desdites cellules sur un support et support pour mettre en oeuvre ladite methode
WO1998005965A1 (fr) * 1996-08-01 1998-02-12 Societe De Recherche Et De Developpement En Activation Et Communication Cellulaire Methode d'identification de cellules eucaryotes par fixation desdites cellules sur un support et support pour mettre en oeuvre ladite methode
US6633662B2 (en) 1997-05-14 2003-10-14 Applied Imaging Corporation Identification of objects of interest using multiple illumination schemes and finding overlap of features in corresponding multiple images
US6169816B1 (en) 1997-05-14 2001-01-02 Applied Imaging, Inc. Identification of objects of interest using multiple illumination schemes and finding overlap of features in corresponding multiple images
US6259807B1 (en) 1997-05-14 2001-07-10 Applied Imaging Corp. Identification of objects of interest using multiple illumination schemes and finding overlap of features in corresponding multiple images
WO1998052018A1 (fr) * 1997-05-14 1998-11-19 Applied Imaging Corporation Identification d'objets par formation d'images multiples
US6956695B2 (en) 2001-03-19 2005-10-18 Ikonisys, Inc. System and method for increasing the contrast of an image produced by an epifluorescence microscope
US7330309B2 (en) 2001-03-19 2008-02-12 Ikonisys, Inc. System and method for increasing the contrast of an image produced by an epifluorescence microscope
US11413296B2 (en) 2005-11-12 2022-08-16 The Regents Of The University Of California Viscous budesonide for the treatment of inflammatory diseases of the gastrointestinal tract
EP1991708A2 (fr) * 2006-03-13 2008-11-19 Ikonisys, Inc. Procédé permettant de combiner immunocoloration et hybridation en fluorescence in situ (fish) au moyen de fluorophores pouvant se lier de manière covalente
EP1991708A4 (fr) * 2006-03-13 2009-12-02 Ikonisys Inc Procede permettant de combiner immunocoloration et hybridation en fluorescence in situ (fish) au moyen de fluorophores pouvant se lier de manière covalente
WO2010022332A1 (fr) 2008-08-22 2010-02-25 Ventana Medical Systems, Inc. Procédé pour la détection chromogénique d’au moins deux molécules cibles dans un unique échantillon
US8481270B2 (en) 2008-08-22 2013-07-09 Ventana Medical Systems, Inc. Method for chromogenic detection of two or more target molecules in a single sample

Also Published As

Publication number Publication date
GB9215710D0 (en) 1992-09-09

Similar Documents

Publication Publication Date Title
Sohda et al. The proportion of fetal nucleated red blood cells in maternal blood: Stimation by FACS analysis
Bianchi et al. Erythroid‐specific antibodies enhance detection of fetal nucleated erythrocytes in maternal blood
US5714325A (en) Prenatal diagnosis by isolation of fetal granulocytes from maternal blood
EP0500727B1 (fr) Procede non envahissant d'isolation et de detection d'adn f tal
GÄNSHIRT‐AHLERT et al. Detection of fetal trisomies 21 and 18 from maternal blood using triple gradient and magnetic cell sorting
US5641628A (en) Non-invasive method for isolation and detection of fetal DNA
Al-Mufti et al. Increased fetal erythroblasts in women who subsequently develop pre-eclampsia
Bianchi et al. Detection of fetal cells with 47, XY,+ 21 karyotype in maternal peripheral blood
Hammadeh et al. Comparison of sperm preparation methods: effect on chromatin and morphology recovery rates and their consequences on the clinical outcome after in vitro fertilization embryo transfer
Al-Mufti et al. Fetal cells in maternal blood of pregnancies with severe fetal growth restriction
Choolani et al. Simultaneous fetal cell identification and diagnosis by epsilon-globin chain immunophenotyping and chromosomal fluorescence in situ hybridization
Little et al. Frequency of fetal cells in sorted subpopulations of nucleated erythroid and CD34+ hematopoietic progenitor cells from maternal peripheral blood
WO2004087863A2 (fr) Diagnostic genetique prenatal non invasif utilisant des cellules transcervicales
Price et al. Simultaneous genotypic and immunophenotypic analysis of interphase cells using dual-color fluorescence: a demonstration of lineage involvement in polycythemia vera
Bischoff et al. Prenatal diagnosis with use of fetal cells isolated from maternal blood: five-color fluorescent in situ hybridization analysis on flow-sorted cells for chromosomes X, Y, 13, 18, and 21
Reading et al. Nucleated erythrocytes in maternal blood: quantity and quality of fetal cells in enriched populations
de Graaf et al. Enrichment, identification and analysis of fetal cells from maternal blood: evaluation of a prenatal diagnosis system
WO1994002830A1 (fr) Analyse de cellules
Holzgreve et al. Fetal cells in cervical mucus and maternal blood
Christensen et al. Studies on the isolation and identification of fetal nucleated red blood cells in the circulation of pregnant women before and after chorion villus sampling
AU667723B2 (en) Process for the prenatal diagnosis of genetic anomalies
Von Koskull et al. Fetal erythroblasts from maternal blood identified with 2, 3‐bisphosphoglycerate (BPG) and in situ hybridization (ISH) using Y‐specific probes
Zheng et al. Fetal cell identifiers: Results of microscope slide–based immunocytochemical studies as a function of gestational age and abnormality
Coata et al. Prenatal diagnosis of genetic abnormalities using fetal CD34+ stem cells in maternal circulation and evidence they do not affect diagnosis in later pregnancies
WO2000075647A1 (fr) Systeme et procede de depistage diagnostic prenatal

Legal Events

Date Code Title Description
AK Designated states

Kind code of ref document: A1

Designated state(s): JP US

AL Designated countries for regional patents

Kind code of ref document: A1

Designated state(s): AT BE CH DE DK ES FR GB GR IE IT LU MC NL PT SE

121 Ep: the epo has been informed by wipo that ep was designated in this application
122 Ep: pct application non-entry in european phase