Rapid read-out biological indicator

• US 5,418,167 A
• Filed: 08/13/1993
• Issued: 05/23/1995
• Est. Priority Date: 11/29/1988
• Status: Expired due to Term

• Current Assignee: 3M Innovative Properties Company (3M Company)
• Original Assignee: 3M Company
• Inventors: Woodson, Lewis P., Matner, Richard R., Foltz, William E.
• Primary Examiner(s): Wityshyn, Michael G.
• Assistant Examiner(s): DEGEN, NANCY JILL
• Application Number:
  US08/106,816
• Time in Patent Office: 648 Days
• Field of Search: 422/55, 422/56, 422/58, 422/60, 422/61, 422/83, 422/86, 422/101, 435/31, 435/38, 435/808, 435/810, 435/288, 435/291, 435/296, 435/18, 435/19, 435/21, 435/23, 435/24
• US Class Current: 435/287.4
• CPC Class Codes: C12Q 1/22 Testing for sterility conditions
• CPC Class Codes: Y10S 435/81 Packaged device or kit

Abstract

A rapid method of determining the efficacy of a sterilization cycle, and an indicator adapted to perform such method, comprising subjecting to the sterilization cycle a source of active enzyme having activity which correlates with the viability of a microorganism commonly used to monitor sterilization, and incubating the enzyme source, following the completion of the sterilization cycle, with an effective amount of a substrate system capable of reacting with any residual active enzyme to produce a detectable enzyme-modified product.

Claims

1. A sterility indicator comprising

a) an outer container having liquid impermeable and substantially non-gas absorptive walls, said outer container having at least one opening therein;

b) a gas-transmissive, bacteria-impermeable means covering said opening;

c) contained within said outer container, a source of active enzyme in a detectable concentration, said active enzyme having sufficient activity following a sterilization cycle which is sublethal to at least one test microorganism commonly used to monitor sterilization, to react with an effective amount of a substrate system for said active enzyme to produce a detectable enzyme-modified product within less than twenty-four hours, yet said active enzyme having activity which is reduced to background following a sterilization cycle which is lethal to said at least one test microorganism; and

d) also contained within said outer container, an effective amount of a substrate system capable of reacting in aqueous solution with said active enzyme to produce a detectable enzyme-modified product.

2. The sterility indicator of claim 1 wherein said substrate system is in aqueous solution and is contained within a sealed, openable, gas and liquid impermeable inner container.

3. The sterility indicator of claim 1 wherein said source of active enzyme is bacteria or fungi in either the spore or vegetative state.

4. The sterility indicator of claim 3 wherein said source of active enzyme is bacteria or fungi commonly used to monitor sterilization, and said sterility indicator further comprises within an inner container, an aqueous nutrient medium capable, with incubation, of promoting growth of said bacteria or fungi, and a detector material capable of undergoing a visible color change in response to growth of said bacteria or fungi.

5. The sterility indicator of claim 1 wherein said enzyme is one which has sufficient activity following a sterilization cycle which is sublethal to at least one test microorganism commonly used to monitor sterilization, to react with said substrate system to produce a detectable enzyme-modified product within less than twelve hours.

6. The sterility indicator of claim 1 wherein said enzyme is one which has sufficient activity following a sterilization cycle which is sublethal to at least one test microorganism commonly used to monitor sterilization, to react with said substrate system to produce a detectable enzyme-modified product within less than eight hours.

7. The sterility indicator of claim 1 wherein said enzyme is one which has sufficient activity following a sterilization cycle which is sublethal to at least one test microorganism commonly used to monitor sterilization, to react with said substrate system to produce a detectable enzyme-modified product within less than three hours.

8. The sterility indicator of claim 1 wherein said enzyme is one which

a) when subjected to a sterilization cycle which would be just sufficient to decrease the population of 1×

10⁶ test microorganisms commonly used to monitor sterilization to zero, as measured by lack of outgrowth of said test microorganisms, has activity equal to background, as measured by reaction with an effective amount of any substrate system capable of reacting with said active enzyme to produce a detectable enzyme-modified product; and

b) when subjected to a sterilization cycle sufficient to decrease the population of 1×

10⁶ of said test microorganisms commonly used to monitor sterilization by at least about 1 log, but less than about 6 logs, has activity greater than said background, as measured by reaction with an effective amount of said substrate system capable of reacting with active enzyme to produce a detectable enzyme-modified product.

9. The sterility indicator of claim 1 wherein said enzyme is selected from the group consisting of beta-D-glucosidase, alpha-D-glucosidase, alkaline phosphatase, acid phosphatase, butyrate esterase, caprylate esterase lipase, myristate lipase, leucine aminopeptidase, valine aminopeptidase, chymotrypsin, phosphohydrolase, alpha-D-galactosidase, beta-D-galactosidase, alpha-L-arabinofuranosidase, beta-D-glucuronidase, N-acetyl-β

-glucosaminidase, beta-D-cellobiosidase, alanine aminopeptidase, proline aminopeptidase, tyrosine aminopeptidase, leucine aminopeptidase, phenylalanine aminopeptidase and a fatty acid esterase derived from spore-forming microorganisms.

10. The sterility indicator of claim 1 further comprising within the outer container a predetermined number of viable test microorganisms.

11. The sterility indicator of claim 10 further comprising within a sealed, openable, gas and liquid impermeable inner container an aqueous nutrient medium capable, with incubation, of promoting growth of said viable test microorganisms contained within the outer container and a detector material capable of undergoing a visible color change in response to growth of said viable test microorganisms contained within the outer container.