Rapid read-out sterility indicator

A unitary sterility indicator comprising:

• a) an outer container having liquid impermeable and substantially gas non-absorptive walls, said container having at least one opening therein;
• b) gas-transmissive, bacteria-impermeable means covering said at least one opening;
• c) contained within said outer container a detectable amount of a microorganism selected from the group consisting of Bacillus stearothermophilus and Bacillus subtilis;
• d) a sealed, openable gas and liquid impermeable inner container containing a composition consisting of
• 1) aqueous nutrient medium capable, with incubation, of promoting growth of said viable microorganisms;
• 2) an enzyme substrate capable of reacting in the presence of an aqueous medium with an active enzyme surviving a sublethal sterilization cycle to produce a detectable enzyme-modified product;
• and
• 3) a detector material capable of undergoing a visible color change in response to growth of said microorganism;
• said inner container being disposed in said outer container and said inner container, when opened, permitting said composition to contact said microorganism;
• and
• e) means contained within said outer container for restricting the a volume in which said composition and said microorganism are contained together, after said inner container is opened and said composition is allowed to contact said microorganisms, to a volume which is less than the volume of said composition contained in said inner container.

A method for testing the efficacy of a sterilization cycle comprising the sequential steps of:

• a) subjecting to said sterilization cycle a unitary sterility indicator comprising:
• 1) an outer container having liquid impermeable and substantially gas non-absorptive walls, said container having at least one opening therein;
• 2) gas-transmissive, bacteria-impermeable means covering said at least one opening;
• 3) contained within said outer container a source of active enzyme which provides a detectable amount of said enzyme, said 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 enzyme to produce a detectable enzyme-modified product within less than twenty-four hours, yet said enzyme having activity which is reduced to "background" following a sterilization cycle which is lethal to said test microorganism;
• 4) a sealed, openable gas and liquid impermeable inner container, containing an aqueous reaction medium capable of permitting said active enzyme to react with a substrate for said enzyme, said inner container being disposed in said outer container and said inner container, when opened permitting said aqueous reaction medium to contact said source of active enzyme;
• 5) an enzyme substrate contained in at least one of said containers, and capable of reacting in the presence of said aqueous reaction medium with an active enzyme surviving said sublethal sterilization cycle to produce a detectable enzyme-modified product;
• and
• 6) means contained within said outer container for restricting the volume in which said enzyme-modified product is contained, after said inner container is opened and said medium is allowed to contact said source of active enzyme, to a volume which is less than the volume of said aqueous reaction medium contained in said inner container;
• b) causing said openable inner container inside said outer container to open, after said sterilization cycle, permitting said aqueous rection medium to contact any residual active enzyme and said enzyme substrate;
• and
• c) incubating said indicator under conditions sufficient to promote reaction of any enzyme remaining active after said sterilization cycle with said enzyme substrate.