WO2011087544A1

WO2011087544A1 - Efficacy diagnosis of multiple sterilization modalities or processes

Info

Publication number: WO2011087544A1
Application number: PCT/US2010/054078
Authority: WO
Inventors: Gregg A. Mosley
Original assignee: Mosley Gregg A
Priority date: 2009-10-26
Filing date: 2010-10-26
Publication date: 2011-07-21

Abstract

Assemblies, kits, and methods for diagnosing microbial inactivation efficacies are disclosed. Assemblies and kits can include an enclosure and one or more chemical or biological indicators, such as at least two indicators, associated with the enclosure. The one or more chemical or biological indicators can be configured to diagnose the microbial inactivation efficacies of at least two sterilization modalities or at least two sterilization processes of a sterilization modality. The assemblies and kits can be used to challenge the at least two modalities or processes, and provide a means to verify the microbial inactivation efficacy of each modality or process. In some examples, the assemblies and kits can include a plurality of substrates that can reduce rapid penetration of inactivation conditions to the chemical or biological indicators. The substrates, when positioned within the enclosure, can form at least one cavity for placement of a chemical or biological indicator.

Description

EFFICACY DIAGNOSIS OF MULTIPLE STERILIZATION MODALITIES OR PROCESSES

CLAIM OF PRIORITY

5 Benefit of priority is hereby claimed to U.S. Provisional Patent

Application Serial No. 61/255,029, entitled "TEST PACK AND METHOD OF USE," (Attorney Docket No. 3138.001PRV), filed on October 26, 2009, which is hereby incorporated by reference in its entirety.

10 TECHNICAL FIELD

This patent document pertains generally to diagnosis of a microbial inactivation efficacy related to a sterilization modality or process. More particularly, but not by way of limitation, this patent document pertains to assemblies, kits, and methods including or using one or more chemical or

15 biological indicators or inoculated carriers for diagnosing microbial (bacteria, fungi, virus, etc.) inactivation efficacies of at least two sterilization modalities or at least two sterilization processes of a sterilization modality.

BACKGROUND

20 In recent years, there has been an increasing need for improved

techniques of sterilizing, disinfecting, sanitizing or otherwise inactivating articles, surfaces, or areas that may have been intentionally or accidentally exposed to biological or chemical contaminants harmful to humans or animals. For example, such articles, surfaces, or areas may have been inadvertently

25 tainted with biological or chemical contaminants as a result of a laboratory or

industrial accident. Alternatively, such articles, surfaces, or areas may have been intentionally contaminated with harmful substances during the commission of a criminal or terrorist act.

One way of guarding against contamination is to inspect, sterilize, and

30 test each and every article, surface, or area. However, this approach is generally regarded as unworkable due to the time and expense required if multiple articles, surfaces, or areas are present. Another approach to assuring successful sterilizing, disinfecting, sanitizing or otherwise inactivating microorganisms in or on articles, surfaces, or areas is to validate sterilization using a single sterilization, disinfection, sanitization, or biological inactivation modality or process and use a surrogate that is designed to represent a worst case challenge as a replacement for testing the normal microbes. For example, bulk quantities of articles, surfaces, or areas can be sterilized by chlorine dioxide gas. Similarly, electron beam ("e- beam") irradiation has been used to inactivate anthrax spores on or within pieces of mail. However, these approaches also have drawbacks in that the effectiveness of a single sterilization modality or process can be limited. Articles, surfaces, or areas can be contaminated with one or more of a variety of biological or chemical agents for which the single sterilization modality or process may be incapable of inactivating. For example, although e- beam technology can be effective in killing anthrax spores, it is incapable of destroying other biological contaminants such as botulism toxin or ricin.

There are several conventional methods to test the efficacy or effectiveness of a single sterilization modality or process on a sample article, surface, or area or load of articles, surfaces, or areas. An example of a method includes inoculating a sample article, surface, or area with a known quantity of a specific indicator organism (the "inoculate"), subjecting the inoculated article, surface, or area to a specific sterilization process, recovering the sample inoculate, and culturing the inoculate in a specific growth medium to determine whether any organisms survived.

OVERVIEW

The present inventor has recognized, among other things, a need for assemblies, kits, and methods configured to diagnose microbial inactivation efficacies of at least two sterilization modalities or at least two sterilization processes of a sterilization modality influencing or configured to influence a load of articles, surfaces, or areas. In this way, the inactivation effectiveness of multiple simultaneous modalities or consecutive sterilization modalities or processes impressed on a load of articles, surfaces, or areas can be efficiently and accurately tested. The assemblies, kits, and methods can be used to efficiently, accurately, and cost effectively test varying sterilization modalities and processes in industries related to bioterrorism, health care, medical devices, pharmaceuticals, paper currency, mail, food packaging and preparation, and other industries, and further determine an inactivation efficacy of each modality or process. The present inventor has further recognized a need for self-contained assemblies, kits, and methods that do not require destruction of a sample load article, surface, or area during a microbial inactivation efficacy test.

The present invention is directed to assemblies, kits, and methods for diagnosing microbial inactivation efficacies. Assemblies and kits can include an enclosure and one or more chemical or biological indicators, such as at least two chemical or biological indicators, associated with the enclosure. The one or more chemical or biological indicators can be configured to diagnose the microbial inactivation efficacies of at least two sterilization modalities or at least two sterilization processes of a sterilization modality. The assemblies and kits can be used to challenge the at least two modalities or processes, and provide a means to verify the microbial inactivation efficacy of each modality or process. In some examples, the assemblies and kits can include a plurality of substrates that can reduce rapid penetration of inactivation conditions to the chemical or biological indicators. The substrates, when positioned within the enclosure, can form at least one cavity for placement of a chemical or biological indicator. In some examples, the assemblies and kits can include one or more chemical indicators to provide visual assurance of exposure to a sterilization modality or process.

To better illustrate the assemblies, kits, and methods disclosed herein, a non-limiting list of examples is provided here:

In Example 1, an assembly for diagnosing a microbial inactivation efficacy comprises an enclosure and at least two chemical or biological indicators positioned on or within the enclosure. The at least two chemical or biological indicators are configured to diagnose the microbial inactivation efficacy of at least two sterilization modalities, such as two different sterilization modalities, or at least two sterilization processes of a sterilization modality, such as two sterilization processes of a single sterilization modality. In Example 2, the assembly of Example 1 is optionally configured such that the enclosure includes a first enclosure and a second enclosure. At least one of the first and second enclosures includes at least two chemical or biological indicators associated therewith. Optionally, the other of the first and second enclosures includes only one chemical or biological indicator associated therewith.

In Example 3, the assembly of any one or any combination of Examples 1 or 2 is optionally configured such that the at least two chemical or biological indicators include a chemical indicator positioned on an outside surface of the enclosure.

In Example 4, the assembly of any one or any combination of Examples 1-3 is optionally configured such that the at least two chemical or biological indicators include a first and a second chemical indicator positioned within the enclosure. The first chemical indicator is configured to monitor a first sterilization modality and the second chemical indicator is configured to monitor a second, different sterilization modality. Alternatively, the first chemical indicator can be configured to monitor a first sterilization process of a specified modality and the second chemical indicator can be configured to monitor a second, different sterilization process of the specified modality.

In Example 5, the assembly of any one or any combination of Examples

1-4 is optionally configured such that the at least two chemical or biological indicators include at least one biological indicator placed within the enclosure.

In Example 6, the assembly of Example 5 is optionally configured such that the at least one biological indicator positioned within the enclosure includes a first and a second biological indicator. The first biological indicator is configured to monitor a first sterilization modality and the second biological indicator is configured to monitor a second, different sterilization modality. Alternatively, the first biological indicator can be configured to monitor a first sterilization process of a specified modality and the second biological indicator can be configured to monitor a second, different sterilization process of the specified modality. In Example 7, the assembly of Example 5 is optionally configured such that the at least one biological indicator positioned within the enclosure includes a first biological indicator configured to monitor a heat sterilization modality, a second biological indicator configured to monitor a chemical sterilization modality, and a third biological indicator configured to monitor a radiation sterilization modality.

In Example 8, the assembly of Example 7 is optionally configured such that the first biological indicator includes Geobacillus stearothermophilus, the second biological indicator includes Bacillus atrophaeus, and the third biological indicator includes Bacillus pulmilus.

In Example 9, the assembly of any one or any combination of Examples 1-8 optionally comprises a plurality of substrates having a size and a shape configured for positioning within the enclosure.

In Example 10, the assembly of Example 9 is optionally configured such that the plurality of substrates, when positioned within the enclosure, form at least one cavity.

In Example 11, the assembly of Example 10 is optionally configured such that at least one chemical or biological indicator is positioned within the at least one cavity.

In Example 12, the assembly of any one or any combination of Examples

9-11 is optionally configured such that the plurality of substrates, when positioned within the enclosure, form at least two cavities.

In Example 13, the assembly of any one or any combination of Examples

1-12 is optionally configured such that each of the at least two chemical or biological indicators is configured to be independently tested for microbial inactivation efficacy.

In Example 14, the assembly of any one or any combination of Examples

1-13 optionally comprises a carrier for the at least two chemical or biological indicators. The carrier is formed of a material selected from the group consisting of: cellulose, cotton, wood, sand, polyester, ceramic, plastic, glass, fiberglass, and metal. In Example 15, the assembly of Example 14 is optionally configured such that a combination of the enclosure, the at least two chemical or biological indicators, and the carrier is configured to mimic or exceed the resistance of a challenge to the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality.

In Example 16, the assembly of any one or any combination of Examples 1-15 is optionally configured such that the at least two chemical or biological indicators are configured to provide varying levels of resistance to the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality.

In Example 17, the assembly of any one or any combination of Examples 1-16 is optionally configured such that the at least two chemical or biological indicators include a chemical indicator configured to provide a visual indication of exposure to a sterilization modality or a sterilization process.

In Example 18, a kit comprises the assembly according to any one or any combination of Examples 1-17; and a set of instructions for using the assembly to diagnose the microbial inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality.

In Example 19, a method of diagnosing a microbial inactivation efficacy comprises subjecting an enclosure and one or more associated chemical or biological indicators to at least two sterilization modalities or at least two sterilization processes of a sterilization modality, and diagnosing the microbial inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality using the one or more chemical or biological indicators during a single challenge.

In Example 20, the method of Example 19 is optionally configured such that subjecting the enclosure to the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality includes making use of a first enclosure and a second enclosure. At least one of the first and second enclosures includes at least two chemical or biological indicators associated therewith. In Example 21, the method of any one or any combination of Examples 19 or 20 is optionally configured such that diagnosing the microbial inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality includes monitoring a hierarchical microbial inactivation efficacy of the at least two modalities or processes.

In Example 22, the method of Example 21 is optionally configured such that monitoring the hierarchical microbial inactivation efficacy includes comparing the microbial inactivation efficacy of a heat sterilization modality, the microbial inactivation efficacy of a chemical sterilization modality, and the microbial inactivation efficacy of a radiation sterilization modality.

In Example 23, the method of any one or any combination of Examples 21 or 22 is optionally configured such that monitoring the hierarchical microbial inactivation efficacy includes evaluating at least one biological indicator in a culture medium and detecting any surviving spores.

In Example 24, the method of any one or any combination of Examples

19-23 is optionally configured such that diagnosing the microbial inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality includes determining whether a sterilization modality or a sterilization process is ineffective.

In Example 25, the method of any one or any combination of Examples

19-24 is optionally configured such that diagnosing the microbial inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality includes diagnosing an efficacy of each sterilization modality or process simultaneously.

In Example 26, the method of any one or any combination of Examples

19-24 is optionally configured such that diagnosing the microbial inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality includes diagnosing an efficacy of each sterilization modality or process consecutively.

In Example 27, the method of any one or any combination of Examples

19-26 is optionally configured such that diagnosing the microbial inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality includes monitoring degradation of a sterilization process.

In Example 28, the method of any one or any combination of Examples 19-27 optionally comprises changing a process variable parameter of a sterilization process using the microbial inactivation efficacy diagnosis of one or more of the at least two sterilization modalities or processes.

In Example 29, the method of Example 28 optionally comprises subjecting a second enclosure and one or more associated chemical or biological indicators to a sterilization process for which the process variable parameter was changed.

In Example 30, the method of any one or any combination of Examples 19-29 optionally comprises diagnosing a viral inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality.

In Example 31, the assembly, kit, or method of any one or any combination of Examples 1-30 is optionally configured such that all elements or options recited are available to use or select from.

These and other examples, advantages, and features of the present assemblies, kits, and methods will be set forth in part in following Detailed Description. This Overview is intended to provide non-limiting examples of the present subject matter— it is not intended to provide an exclusive or exhaustive explanation. The Detailed Description is included to provide further information about the present assemblies, kits, and methods.

BRIEF DESCRIPTION OF THE DRAWINGS

In the drawings, like numerals can be used to describe similar components throughout the several views. Like numerals having different letter suffixes can be used to represent different instances of similar components. The drawings illustrate generally, by way of example, but not by way of limitation, various embodiments discussed in the present document.

FIG. 1 illustrates a component view of an assembly for

diagnosing microbial inactivation efficacies of at least two sterilization modalities or at least two sterilization processes of a sterilization modality, as constructed in accordance with at least one embodiment.

FIG. 2 illustrates varying biological indicators for use in

FIG. 3 illustrates varying chemical indicators for use in

FIG. 4 illustrates a partially assembled view of an assembly for diagnosing microbial inactivation efficacies of at least two sterilization modalities or at least two sterilization processes of a sterilization modality, as constructed in accordance with at least one embodiment.

FIG. 5 illustrates another partially assembled view of an

assembly for diagnosing microbial inactivation efficacies of at least two sterilization modalities or at least two sterilization processes of a sterilization modality, as constructed in accordance with at least one embodiment.

FIG. 6 illustrates a perspective view of an enclosure for use in diagnosing microbial inactivation efficacies of at least two sterilization modalities or at least two sterilization processes of a sterilization modality, as constructed in accordance with at least one embodiment.

FIG. 7 illustrates a cross-sectional view, such as along line 7-7,

of the enclosure shown in FIG. 6, as constructed in accordance with at least one embodiment. FIG. 8 illustrates an assembled view of an assembly for diagnosing microbial inactivation efficacies of at least two sterilization modalities or at least two sterilization processes of a sterilization modality, as constructed in accordance with at least one embodiment.

FIG. 9 illustrates a perspective view of a sterilization system that can be used with an assembly for diagnosing microbial inactivation efficacies of at least two sterilization modalities or at least two sterilization processes of a sterilization modality.

DETAILED DESCRIPTION

Validation and verification of a sterilization modality's or process's efficacy can be important aspects of knowledge for bioterrorism, health care articles, medical devices, paper currency, mail, or any treatment process for sterilization, disinfection, sanitization, or biological inactivation of food products. Unfortunately, conventional means to test the efficacy of a sterilization modality or process often result in article, surface, or area destruction due to requiring the inoculation of a sample article, surface, or area with a known quantity of a specific indicator organism, subjecting the inoculated article, surface, or area to a specified modality or process known to be associated with the specific indicator organism, recovering the sample inoculate, and culturing the inoculate in an appropriate growth medium to determine whether any indicator organisms survive. Additionally, conventional test means are limited to use with a single sterilization modality or process.

Advantageously, the present invention provides assemblies, kits, and methods configured to diagnose the microbial inactivation efficacy of at least two sterilization modalities, such as two different sterilization modalities, or at least two sterilization processes of a sterilization modality, such as two sterilization processes of a single sterilization modality, inflicted on a single load of articles, surfaces, or areas ("challenge") without necessarily requiring destruction of sample load articles, surfaces, or areas. In this way, the present assemblies, kits, and methods can provide a time and cost savings over conventional test means, as well as preserve a load article's, surface's, or area's integrity.

FIG. 1 illustrates a component view of an assembly 100 for diagnosing microbial inactivation efficacies of at least two sterilization modalities or at least two sterilization processes of a sterilization modality, as constructed in accordance with at least one embodiment. Components of the assembly 100 can be tailored to mimic or exceed the resistance of a challenge to the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality. The assembly 100 can be used to challenge the modalities or processes, thereby providing a means to validate the microbial inactivation efficacy of each. The assembly 100 can include one or more chemical or biological indicators, such as at least two chemical or biological indicators, on or within an enclosure 102 (e.g., a pressed paper box having dimensions of about 3 inches x 5 inches x 1 inch). In various examples, the assembly 100 can include a multi-organism set of biological indicators 104 and a multi-set of chemical indicators 106, which alone or together can verify a minimum level of delivered microbial inactivation from the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality.

In the example shown, the assembly 100 includes four biological indicator 104 organisms, each contained in or on a carrier 108 and optionally enclosed within a barrier package 110. The specific biological indicator 104 organisms and carriers 108 can be chosen for their appropriateness for a specific set of sterilization modalities operating simultaneously or sterilization modalities or processes operating consecutively. One of the biological indicator 104 organisms can be used in or on two different carrier 108 materials allowing differential diagnostic measurements and all can be placed within a stack of paper substrates 112 and positioned inside the enclosure 102. Some of the substrates 112 can include cut-out regions 114 that are used to form cavities 116 for placement of the chemical or biological indicators 104. The substrates 112 can be configured to increase the measured resistance of each biological organism, for example, to the given at least two sterilization modalities or the at least two sterilization processes of a sterilization modality. The assembly 100 can also includes at least two chemical indicators 106 inside the enclosure 102, an instruction label 118, and a tamper seal 150 configured to seal the enclosure 102. The tamper seal 150 can include a chemical indicator 106, such as a heat sterilization modality indicator.

FIG. 2 illustrates a plurality of biological indicators 104 for use in an assembly configured to diagnose microbial inactivation efficacies of at least two sterilization modalities or at least two sterilization processes of a sterilization modality, as constructed in accordance with at least one embodiment. As disclosed above, each of the monitored sterilization modalities or processes can call for a unique biological indicator 104 organism to confirm microbial inactivation efficacy. Multiple simultaneous modalities or consecutive sterilization modalities or sterilization processes can uniquely effect biological organisms.

The resistance of a biological indicator 104 to a particular sterilization modality or process can be given as a D-value, which is defined as the exposure time required under a defined set of conditions to cause a 1 -logarithm or 90% reduction in the population of a particular organism. Each biological indicator 104 can have a listed D-value for that modality or process to which it provides the greatest resistance. For example, the organism Geobacillus

stearothermophilus can include an appreciable resistance to a process using moist heat ("steam") exposure as the sterilization modality, the organism Bacillus atrophaeus can include an appreciable resistance to a process using dry heat exposure as the sterilization modality, and the organism Bacillus pumilis can include an appreciable resistance to a process using ionizing radiation exposure as the sterilization modality. A process using ultraviolet ("UV") radiation exposure as the sterilization modality can also be used in the methods disclosed herein with appropriate biological indicator organisms to test for its effectiveness included in the assembly 100. The varying biological indicators 104 can provide different levels of resistance to a process using radiation exposure, a process using dry heat exposure, or a process using moist heat exposure, for example. Each of the biological indicators 104 can include a carrier 108 comprised of a material selected from the group consisting of, but not limited to: cellulose, cotton, wood, sand, polyester, ceramic, plastic, glass, fiberglass, and metal, and optionally shaped in the form of strips inoculated with a known population of an organism. In some examples, such as is shown in FIG. 1, a glass carrier can be placed in a cavity formed from a plurality of substrates including a cut-out region 114 (FIG. 1) to prevent damage during a challenge. The carriers 108 can be used to carry or contain the organisms of the biological indicators 104. The biological indicators 104, via the carriers 108, can include a known quantity of a biological indicator organism. For example, a concentration of Bacillus atrophaeus for use in an ethylene oxide gas sterilization modality can be 10⁸ (microorganisms per carrier) and stored on a cellulose carrier 108, and a concentration of Geobacillus stearothermophilus for use in a steam sterilization modality can be 10⁷. For food processing, a biological indicator 104 can include Clostridium. Other microorganisms that can be used in the biological indicators 104 include, but are not limited to, Bacillus circulans, Bacillus cereus, Bacillus pumilus, Bacillus thuringiensis, Bacillus amyloliquefaciens, or a biological enzyme. Table 1 lists the biological indicators 104 shown in FIG. 2. Table 1: Example biological indicator organism list.

Following exposure, each biological indicator 104 can be removed from its associated carrier 108 and placed in an appropriate microbiological culture medium, which allows outgrowth and detection of any surviving spores. FIG. 3 illustrates a plurality of chemical indicators 106 for use in an assembly configured to diagnose microbial inactivation efficacies of at least two sterilization modalities or at least two sterilization processes of a sterilization modality, as constructed in accordance with at least one embodiment. The chemical indicators 106 can each monitor different sterilization modalities, such as dry heat or steam, or processes, such as dry heat at a first temperature and dry heat at a second, elevated temperature, as well as provide a means for visually indicating that the assembly 100 (FIG. 1) has been exposed to a sterilization modality or sterilization process. Table 2 lists the chemical indicators 106 shown in FIG. 3. Other chemical indicators can include, but are not limited to: radiation indicators, ethylene oxide gas indicators and other specific chemical gas indicators.

Table 2: Example chemical indicator list.

FIGS. 4 and 5 illustrate partially assembled views of an assembly 100 for diagnosing microbial inactivation efficacies of at least two sterilization modalities or at least two sterilization processes of a sterilization modality, as constructed in accordance with at least one embodiment. As shown, one or more chemical 106 indicators can be placed within an enclosure 102 near an outer enclosure surface 402, while one or more biological indicators 104 can be placed within the enclosure 102 and spaced from the outer enclosure surfaces 402. Due to their relative outer enclosure 102 placement, the chemical indicators 106 can diagnose microbial inactivation efficacies of the at least two sterilization modalities or processes earlier than the more-centralized biological indicators 104. In some examples, one or more biological indicators 104 can be placed near an outer enclosure surface 402 or placed near the center, thereby more closely mimicking or exceeding the resistance of a certain challenge to the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality.

Domestic and international regulatory guidelines permit the use of assemblies 100 that demonstrably have resistance to sterilization modalities or sterilization processes equal to or greater than the one or more articles, surfaces, or areas to be subjected to the at least two sterilization modalities or processes. The enclosure 102 of the assembly 100 can be fabricated from a suitable single or multiple layer material that is chosen to offer the appropriate level of resistance to the at least two sterilization modalities or sterilization processes. In various examples, the enclosure 102 includes a box-like configuration.

Similarly, a plurality of substrates 112 positionable inside the enclosure 102 can be chosen to increase the measured resistance of the included biological 104 or chemical 106 indicators to the given modalities or processes. Accordingly, the assembly 100 can be used alone to mimic or exceed the resistance of a challenge to the at least two sterilization modalities or at least two sterilization processes of a sterilization modality, rather than requiring that the assembly 100 be processed with an actual load of articles, surfaces, or areas. In some examples, the assembly 100 is used with a challenge.

A retail kit can also be packaged for consumer purchase. The retail kit can include an assembly 100 and a set of instructions for using the assembly to diagnose a microbial inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality. The instructions can include one or more of: subjecting the assembly 100 to at least two sterilization modalities or processes; allowing the assembly 100 to cool; monitoring a status of a chemical indicator 306 (FIG. 3) on a tamper seal surrounding a portion of the enclosure 402; and testing or sending for testing of the chemical and biological indicators enclosed with the assembly 100. Time is saved by not having to inoculate sample load articles, surfaces, or areas before they are packaged and no articles, surfaces, or areas have to be sacrificed as part of the microbial inactivation efficacy testing process. FIGS. 6 and 7 illustrate perspective views of an enclosure 102 and cavities 116 formed by a stack of substrates 112, some of which include cut-out regions 114 (FIG. 1), positionable with the enclosure 102. The cavities 116 can be selectively positioned within the enclosure 102 to provide the desired resistance insulation to sterilization modalities or sterilization processes for one or more chemical or biological indicators. This selective cavity 116 positioning can be effectuated by tailoring the placement of the substrate 112 cut-out regions. As shown in the example of FIG. 7, ten full substrates 112 can form the base of an enclosed substrate stack, followed by forty- five substrates 112 including a centralized void region, followed by two full substrates 112, followed by five substrates 112 including a non-centralized void region, and finally topped off with ten full substrates 112. The full substrates 112 positioned throughout the substrate stack can help insulate and protect the one or more chemical or biological indicators positioned in the cavities 116, as well as provide the desired resistance insulation to the sterilization modalities or sterilization processes.

FIG. 8 illustrates an assembled view of an assembly 100 for diagnosing microbial inactivation efficacies of at least two sterilization modalities or at least two sterilization processes of a sterilization modality, as constructed in accordance with at least one embodiment. A chemical indicator 306 integrated with a tamper seal can be placed on an outside surface 402 of an assembly enclosure 102. The chemical indicator 306 can provide an initial visual means for indicating that the assembly 100 has been exposed to a sterilization modality or sterilization process.

In use, one or more assemblies 100 can be placed at various locations within a challenge or can be used alone. The load can then be subjected to the chosen sterilization modalities or sterilization processes. After the modalities or processes are complete, the one or more assemblies 100 can be removed for analysis to diagnose microbial inactivation efficacies of the sterilization modalities or processes. In examples using one or more biological indicator organisms, the biological indicators can be removed from the enclosure 102 and incubated in an appropriate culture medium for culturing. The absence of growth of indicator organisms can indicate the successful sterilization by at least one sterilization modality or process. In examples using biological enzyme indicators, the biological indicator can be removed and exposed to an appropriate enzyme substrate to determine whether any enzyme activity remains. The absence of enzymatic activity can indicate the successful sterilization by at least one sterilization modality or process.

FIG. 9 illustrates a perspective view of a sterilization system 900 that can be used with an assembly 100 for diagnosing microbial inactivation efficacies of at least two sterilization modalities. A system similar to the sterilization system 900 can be configured to deliver at least two sterilization modalities, which can be selected from, among others: steam, dry heat, ionizing radiation, radiation beam technology, electromagnetic field technology, ultraviolet radiation technology, chemical decontamination technology, hydrogen peroxide, ethylene gas, ozone, plasma, or combinations thereof. A system similar to the sterilization system 900 can also be configured to deliver at least two

sterilization processes of a sterilization modality, such as varying parameters for process variables. The assembly 100 can be tailored to measure a minimum level of resistance to the at least two sterilization modalities or processes using a combination of different chemical or biological indicators, such as different biological indicator organisms, each with an identified level of resistance to the different modalities or processes.

The sterilization system 900 can include a test box 902 with a door 904, an article tumbling drum 906, a sterilization in-progress/completed status indicator 908, an input port 910, and an output port 912. The system 900 can further include an irradiator source and applicator 914, a moist heat or steam source and applicator 916, a dry heat source and applicator 918, and a microwave sterilization source and applicator 920. It is to be understood, however, that in some examples, the system 900 can additionally or alternatively employ any other suitable sterilization technology, such as x-ray, broadband light beam, radiation beam, electromagnetic field, UV radiation, or chemical technologies. In the example shown, the system 900 can employ UV ray technology, moist heat technology, dry heat technology, microwave sterilization technology, and/or suitable combinations of these technologies, for effectively sterilizing a load of articles. To this end, a quantity of potentially contaminated articles can be placed and confined in the tumbling drum 906 inside the test box 902, the test box door 904 can be closed, and the articles in the tumbling drum 906 can undergo at least two sterilization modalities using one or more of the above- mentioned technologies.

In some examples, the tumbling drum 906 can be configured to handle various masses and volumes of articles during each sterilization process.

Further, the tumbling drum 906 can be configured for rotationally oscillating about a hub 922, as depicted by directional arrows 924. The speed and direction of rotation of the tumbling drum 906 can be pre-set or pre-programmed to assure that all portions of the load articles are exposed to the applied irradiation, dry heat, moist heat, and/or microwave technologies. For example, the speed may be pre-set to a single speed, or pre-programmed to a number of varying speeds. Similarly, the direction of rotation may be pre-set to a single rotation direction, or pre-programmed to change direction a predetermined number of times.

Moreover, the surfaces of the tumbling drum 906, and the internal surfaces of the test box 902 including the door 904, can be made reflective to amplify the light ray disinfection energy applied to the articles during one or more sterilization modalities.

The test box 902 and door 904 can be suitably shielded and sealed to prevent leakage of irradiation, dry heat, or moisture during the at least two sterilization modalities. The test box 902 can be further configured to prevent potentially harmful biological and/or chemical substances from escaping until the substances are either destroyed or otherwise rendered inactive by the sterilization modalities. To this end, the air pressure inside the test box 902 can be made to be below atmospheric pressure. Specifically, the input port 910 can be configured to allow ambient air to pass therethrough, and to enter the test box 902 via one or more orifices. It is noted that a filter can be employed to filter the ambient air before it enters and after it leaves the test box 902. The output port 912 can be configured to draw the ambient air from the input port 910, through the inside of the test box 902, and back outside the test box, using, for example, an air blower. As a result, even if there were any unwanted air leaks in the system 900, the air would simply be drawn into the test box 902 and be subsequently expelled through the output port 912. As shown in FIG. 9, the test box door 904 can include a transparent section 920 to allow a human operator to observe the articles in the tumbling drum 906.

EXPERIMENTAL EXAMPLES

In order that the present assemblies, kits, and methods can be more fully understood, the following examples are given by way of illustration. These examples can verify the efficacy of the assemblies, kits, and methods, in particular included biological and/or chemical indicators, to measure microbial inactivation from various types of sterilization modalities or sterilization processes and distinguish effects resulting from the different modalities or processes. When combined, the biological indicators and the chemical indicators can be used to assess deficient and successful performances for varying sterilization modalities or sterilization processes. Experimental Example 1 - Gamma Sterilization Modality

A first experimental example subjected two assemblies 100 to a process using gamma ionizing radiation exposure as the modality. A first assembly was subjected to an 8.3 kGy gamma sterilization process and a second assembly was subjected to a 26 kGy gamma sterilization process. After the processes were complete, the assemblies were removed from a sterilization system and sent to a laboratory for analysis and to record diagnosed microbial inactivation efficacies of the gamma sterilization processes.

Experimental Example 2 - Steam Sterilization Modality A second experimental example subjected three assemblies 100 to a process using pre- vacuum moist heat ("steam") exposure as the modality. A first assembly was subjected to 5 minutes of a steam sterilization process at 121 °C, a second assembly was subjected to 10 minutes of a steam sterilization process at 121 °C, and a third assembly was subject to 30 minutes of a steam sterilization process at 121 °C. After the processes were complete, the assemblies were removed from a sterilization system, allowed to cool, and sent to a laboratory for analysis and to record diagnosed microbial inactivation efficacies of the steam sterilization processes.

Experimental Example 3 - Dry Heat Sterilization Modality A third experimental example subjected four assemblies 100 to a process using dry heat exposure as the modality. A first assembly was subjected to 15 minutes of a dry heat sterilization process at 160°C, a second assembly was subjected to 30 minutes of a dry heat sterilization process at 160°C, a third assembly was subject to 60 minutes of a dry heat sterilization process at 160°C, and a fourth assembly was subject to 120 minutes of a dry heat sterilization process at 160°C. After the processes were complete, the assemblies were removed from a sterilization system, allowed to cool, and sent to a laboratory for analysis and to record diagnosed microbial inactivation efficacies of the dry heat sterilization processes. Experimental Example 4 - Multiple Sterilization Modalities

A fourth experimental example subjected a single assembly 100 and a 3- pound article mail load ("challenge") to multiple sterilization modalities operated in a single process including dry heat, microwave activated moist heat, and UV irradiation ("Ra") assisted by rotation of an inner cage and operation below atmospheric pressure for 90 minutes with an approximate 60 minute temperature exposure time. After the process was complete, the assembly was removed from a sterilization system, allowed to cool, and sent to a laboratory for analysis and to record diagnosed microbial inactivation efficacies of the multiple sterilization modalities.

In each of the four experimental examples, the biological indicators 104 included in the assemblies 100 were removed from their associated carriers 108 and placed in an appropriate microbiological culture medium, which allowed outgrowth and detection of any surviving spores. Biological indicator strips of Bacillus atrophaeus ("DH") and Bacillus pumilis (Ra) were cultured in Tryptic Soy Broth ("TSB"), incubated at 30-35°C for 7 days and observed incrementally for indications of growth. Positive cultures were compared to the positive control to assure growth was due to the indicator organism. Dry Amp ("DA") ampoules were opened at the neck and the contents cultured in TSB, incubated at 30-35 °C for 7 days and observed incrementally for indications of growth.

Positive cultures were compared to the positive control to assure growth was due to the indicator organism. Biological indicator strips of Geobacillus

stearothermophilus were cultured in TSB, incubated at 55-60°C for 7 days and observed incrementally for indications of growth. Positive cultures were compared to the positive control to assure growth was due to the indicator organism.

In each of the four experimental examples, the chemical indicators 106 included in the assemblies 100 were also tested. A chemical indicator configured for moist heat sterilization detection on the outside of the assembly ("process") 100 was observed and its color was recorded prior to opening an assembly enclosure 102 for culturing the biological indicators 104. A chemical indicator configured for moist heat sterilization and/or dry heat sterilization ("MH/DH") detection and positioned inside the assembly enclosure 102 was removed, observed and its color recorded prior to culturing the biological indicators 104. A chemical indicator configured for dry heat sterilization detection and positioned inside the assembly enclosure was also removed, observed and its color recorded prior to culturing the biological indicators 104.

In the result tables 3-6, biological indicator results were recorded as 0/n when all test samples were found to be negative, where "n" is the total number of tested samples. If one sample is positive, then the results were listed as l+/n.

Chemical indicators were recorded as "NC" for no change or a C→ indicating a change, and the color was recorded with "Lt" for light, "Drk" for dark, "Par" for partial, "OvExp" for over exposed, "Grn" for green, "Gry" for grey, "Blk" for black, and "Brn" for brown. Table 3: Efficacy Diagnosis Results of a Process Using Gamma Exposure as the Sterilization Modality.

Table 4: Efficacy Diagnosis Results of a Process Using Steam Exposure as the Sterilization Modality.

Table 5: Efficacy Diagnosis Results of a Process Using Dry Heat Exposure as the Sterilization Modality.

Table 6: Efficacy Diagnosis Results of a Process Using Multiple Sterilization Modalities.

These experimental examples demonstrate that the use of one or more 5 assemblies 100 can provide a microbial inactivation efficacy diagnosis for single and multiple sterilization modalities or sterilization processes. This efficacy diagnosis includes the ability to measure or detect: delivered lethality with different types of sterilization modalities or sterilization processes, different levels of delivered lethality based on one or more biological organisms used,

10 inadequate lethality for certain sterilization modalities or sterilization processes, distinguishing characteristics between different types of sterilization modalities or sterilization processes applied to the assembly in a challenge, and which sterilization modalities or sterilization processes are not functioning correctly or are ineffective.

15 Closing Notes:

Assemblies, kits, and methods which, when used, can measure a minimum level of resistance to at least two sterilization modalities or at least two sterilization processes of a sterilization modality. The assemblies, kits, and methods can be used to challenge the sterilization modalities or processes, thereby providing a means to efficiently and cost effectively verify the efficacy of each. The assemblies can include an enclosure and one or more chemical or biological indicators strategically positioned on or within the enclosure. In this way, the assembly can be simply placed into a challenge with having to inject or otherwise destroy a sample article, surface, or area. A combination of indicators can be selected based on an identified resistance to the different sterilization modalities or processes to be used.

The above Detailed Description includes references to the accompanying drawings, which form a part of the Detailed Description. The drawings show, by way of illustration, specific embodiments in which the present assemblies, kits, and methods can be practiced. These embodiments are also referred to herein as "examples."

The above description is intended to be illustrative, and not restrictive. For example, the above-described examples (or one or more features thereof) can be used in combination with each other. Other embodiments can be used, such as by one of ordinary skill in the art upon reviewing the above description. Also, in the above Detailed Description, various features can be grouped together to streamline the disclosure. This should not be interpreted as intending that an unclaimed disclosed feature is essential to any claim. Rather, inventive subject matter can lie in less than all features of a particular disclosed embodiment. Thus, the following claims are hereby incorporated into the Detailed Description, with each claim standing on its own as a separate embodiment. The scope of the invention should be determined with reference to the appended claims, along with the full scope of equivalents to which such claims are entitled.

In this document, the terms "a" or "an" are used, as is common in patent documents, to include one or more than one, independent of any other instances or usages of "at least one" or "one or more." In this document, the term "or" is used to refer to a nonexclusive or, such that "A or B" includes "A but not B," "B but not A," and "A and B," unless otherwise indicated. In this document, the phrase "sterilization modality" refers to a type of sterilization, disinfection, sanitization or otherwise inactivation of microorganisms using, but not limited to, gamma radiation, moist heat, dry heat, electron beam, microwave, ozone, plasma, ethylene oxide, hydrogen peroxide, gaseous peracetic acid or chlorine dioxide. In this document, the phrase "sterilization process" refers to a sterilization modality having specified values ("parameters") for process variables. In this document, the phrase "process variables" refers to a condition within a sterilization process whose changes alter microbiocidal effectiveness and includes all factors affecting the ultimate lethality of the process. In this document, the phrases "microbial inactivation" or "microbial lethality" relate to the inactivation of microorganisms. In this document, the term "diagnose" is used to refer to the identification of cause and effect relationships. In this document "biological indicators" are species which can be used by observers to determine how various conditions in an environment have changed over time; such biological indicators are typically presented as a test system containing viable microorganisms providing a defined resistance to a specified sterilization process. Biological indicators can be available in a variety of presentations such as spore strips and threads, self-contained vials, inoculated articles, etc.

In the appended claims, the terms "including" and "in which" are used as the plain- English equivalents of the respective terms "comprising" and

"wherein." Also, in the following claims, the terms "including" and

"comprising" are open-ended, that is, an assembly, assembly, device, article, kit, or process that includes elements in addition to those listed after such a term in a claim are still deemed to fall within the scope of that claim. Moreover, in the following claims, the terms "first," "second," and "third," etc. are used merely as labels, and are not intended to impose numerical requirements on their objects.

The Abstract is provided to comply with 37 C.F.R. § 1.72(b), to allow the reader to quickly ascertain the nature of the technical disclosure. It is submitted with the understanding that it will not be used to interpret or limit the scope or meaning of the claims.

Claims

WHAT IS CLAIMED IS:

1. An assembly for diagnosing a microbial inactivation efficacy, comprising: an enclosure; and

at least two chemical or biological indicators positioned on or within the enclosure,

wherein the at least two chemical or biological indicators are configured to diagnose the microbial inactivation efficacy of at least two sterilization modalities or at least two sterilization processes of a sterilization modality.

2. The assembly of claim 1, wherein the enclosure includes a first enclosure and a second enclosure, the first enclosure including at least two chemical or biological indicators and the second enclosure including at least one chemical or biological indicator.

3. The assembly of any one of claims 1 or 2, wherein the at least two chemical or biological indicators include a chemical indicator positioned on an outside surface of the enclosure.

4. The assembly of any one of claims 1-3, wherein the at least two chemical or biological indicators include a first and a second chemical indicator positioned within the enclosure, the first chemical indicator configured to monitor a first sterilization modality and the second chemical indicator configured to monitor a second, different sterilization modality.

5. The assembly of any one of claims 1-4, wherein the at least two chemical or biological indicators include at least one biological indicator placed within the enclosure.

6. The assembly of claim 5, wherein the at least one biological indicator positioned within the enclosure includes a first and a second biological indicator, the first biological indicator configured to monitor a first sterilization modality and the second biological indicator configured to monitor a second, different sterilization modality.

7. The assembly of claim 5, wherein the at least one biological indicator positioned within the enclosure includes a first biological indicator configured to monitor a heat sterilization modality, a second biological indicator configured to monitor a chemical sterilization modality, and a third biological indicator configured to monitor a radiation sterilization modality.

8. The assembly of claim 7, wherein the first biological indicator includes Geobacillus stearothermophilus, the second biological indicator includes Bacillus atrophaeus, and the third biological indicator includes Bacillus pulmilus.

9. The assembly of any one of claims 1-7, comprising a plurality of substrates having a size and a shape configured for positioning within the enclosure.

10. The assembly of claim 9, wherein the plurality of substrates, when positioned within the enclosure, form at least one cavity.

11. The assembly of claim 10, wherein at least one chemical or biological indicator is positioned within the at least one cavity.

12. The assembly of any one of claims 9-11, wherein the plurality of substrates, when positioned within the enclosure, form at least two cavities.

13. The assembly of any one of claims 1-12, wherein each of the at least two chemical or biological indicators is configured to be independently tested for microbial inactivation efficacy.

14. The assembly of any one of claims 1-13, comprising a carrier for the at least two chemical or biological indicators, the carrier formed of a material selected from the group consisting of: cellulose, cotton, wood, sand, polyester, ceramic, plastic, glass, fiberglass, and metal.

15. The assembly of claim 14, wherein a combination of the enclosure, the at least two chemical or biological indicators, and the carrier is configured to mimic or exceed the resistance of a challenge to the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality.

16. The assembly of any one of claims 1-15, wherein the at least two chemical or biological indicators are configured to provide varying levels of resistance to the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality.

17. The assembly of any one of claims 1-16, wherein the at least two chemical or biological indicators include a chemical indicator configured to provide a visual indication of exposure to a sterilization modality or a sterilization process.

18. A kit comprising:

the assembly according to any one of claims 1-17; and

a set of instructions for using the assembly to diagnose the microbial inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality.

19. A method of diagnosing a microbial inactivation efficacy, comprising:

subjecting an enclosure and one or more associated chemical or biological indicators to at least two sterilization modalities or at least two sterilization processes of a sterilization modality; and

diagnosing the microbial inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality using the one or more chemical or biological indicators during a single challenge.

20. The method of claim 19, wherein subjecting the enclosure to the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality includes making use of a first enclosure and a second enclosure, at least one of the first and second enclosures including at least two chemical or biological indicators.

21. The method of any one of claims 19 or 20, wherein diagnosing the microbial inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality includes monitoring a hierarchical microbial inactivation efficacy of the at least two modalities or processes.

22. The method of claim 21, wherein monitoring the hierarchical microbial inactivation efficacy includes comparing the microbial inactivation efficacy of a heat sterilization modality, the microbial inactivation efficacy of a chemical sterilization modality, and the microbial inactivation efficacy of a radiation sterilization modality.

23. The method of any one of claims 21 or 22, wherein monitoring the hierarchical microbial inactivation efficacy includes evaluating at least one biological indicator in a culture medium and detecting any surviving spores.

24. The method of any one of claims 19-23, wherein diagnosing the microbial inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality includes monitoring whether a sterilization modality or a sterilization process is ineffective.

25. The method of any one of claims 19-24, wherein diagnosing the microbial inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality includes diagnosing an efficacy of each sterilization modality or process simultaneously.

26. The method of any one of claims 19-24, wherein diagnosing the microbial inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality includes diagnosing an efficacy of each sterilization modality or process consecutively.

27. The method of any one of claims 19-26, wherein diagnosing the microbial inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality includes monitoring degradation of a sterilization process.

28. The method of any one of claims 19-27, comprising changing a process variable parameter of a sterilization process using the microbial inactivation efficacy diagnosis of one or more of the at least two sterilization modalities or processes.

29. The method of claim 28, comprising subjecting a second enclosure and one or more associated chemical or biological indicators to a sterilization process for which the process variable parameter was changed.

30. The method of any one of claims 19-29, comprising diagnosing a viral inactivation efficacy of the at least two sterilization modalities or the at least two sterilization processes of a sterilization modality.