Biological Indicator for Steam Sterilization, Self-Contained

» Biological Indicator for Steam Sterilization, Self-Contained, is a Biological Indicator for Steam Sterilization, Paper Carrier individually packaged in a suitable container readily penetrable by steam and designed to hold an appropriate bacteriological culture medium, so as to enable the packaged carrier, after subjection to saturated steam sterilization conditions, to be incubated in the supplied medium in a self-contained system. The supplied medium may contain a suitable indicator as a convenience for determining by a color change whether or not spores have survived. The design of the self-contained system is such that, after exposure to the specified sterilization conditions and inoculation of the medium under closed conditions as stated in the labeling, there is no loss of medium and inoculum during subsequent transport and handling, if done according to the provided instructions. The materials of which the self-contained system are made are such that there is no retention or release of any substance that may cause inhibition of growth of surviving spores under the incubation conditions stated in the labeling.

Packaging and storage— Preserve in the original package under the conditions recommended on the label, and protect from light, from substances that may adversely affect the contained microorganisms, from excessive heat, and from moisture.

Expiration date— The expiration date is determined on the basis of stability studies and is not less than 18 months from the date of manufacture, the date of manufacture being the date on which the first determination of the total viable spore count was made.

Labeling— Label it to state that it is a Biological Indicator for Steam Sterilization, Self-Contained; to indicate the D value of the self-contained system, the method used to determine such D value (i.e., by spore count or fraction negative procedure after graded exposures to the sterilization conditions); the survival time and kill time under the specified conditions stated on the label; its particular total viable spore count, with a statement that such count has been determined after preliminary heat treatment; and its recommended storage conditions. State on the labeling that the supplied bacteriological medium will meet requirements for growth-promoting ability, the strain and ATCC number from which the spores were derived, and the instructions for spore recovery and for safe disposal of the indicator unit. Also indicate in the labeling that the stated resistance characteristics are reproducible only under steam sterilization conditions at the stated temperature and only under the exact conditions under which it was determined, that the user would not necessarily obtain the same result, and that the user should determine the suitability of the biological indicator for the particular use.

Identification— It meets the requirements of the Identification test under Biological Indicator for Steam Sterilization, Paper Carrier.

Resistance performance tests—

D value— Proceed as directed for the relevant procedure for D Value under Biological Indicators—Resistance Performance Tests

. The requirements of the test are met if the determined D value is within 20% of the labeled D value for the selected sterilizing temperature and if the confidence limits of the estimate are within 10% of the determined D value.

Survival time and kill time— Follow the procedure for Survival Time and Kill Time in the section Steam Sterilization, Self-Contained, under Biological Indicators—Resistance Performance Tests

. The requirements of the test are met if all of the specimens subjected to the steam sterilization for the survival time show evidence of growth, while none of the specimens subjected to the steam sterilization for the kill time shows growth. If for either the survival time or the kill time requirement, not more than 1 specimen out of both groups fails the test, whichever is applicable, continue the corresponding test with 4 additional groups, each consisting of 10 specimens, according to the procedure described above. If all of the additional specimens subjected to steam sterilization either meet the survival requirement for the survival time test or meet the kill requirement for the kill time test, whichever is applicable, the requirements of the test are met.

Total viable spore count— Proceed as directed for Total Viable Spore Count under Biological Indicators—Resistance Performance Tests

using the procedure applicable to Biological Indicator for Steam Sterilization, Paper Carrier. The requirements of the test are met if the average number of viable spores per carrier is not less than 0.3 log of the labeled spore count per carrier and does not exceed the log labeled spore count per carrier by 0.48.

Medium suitability—

Sterility— Incubate 10 self-contained biological indicator systems at 55

to 60

, or at the optimal recovery temperature specified by the manufacturer, for 48 hours, making sure that there is no contact between the individual spore strips and the supplied medium. Examine the incubated medium visually (for change in color indicator or turbidity) and microscopically (for absence of microbial growth).

Growth promotion of medium prior to sterilization treatment— Submerge 10 self-contained units in a water bath maintained at 95

to 100

for 15 minutes. Start timing when the temperature of the container contents reach 95

. Cool rapidly in an ice-water bath (0

to 4

). Remove the units from the ice-water bath, submerge each spore strip with the self-contained medium, incubate at 55

to 60

, or at the optimal recovery temperature specified by the manufacturer, and examine visually after 48 hours for growth (for turbidity or change in color), and microscopically (for microbial growth). All the specimens under test show growth. If one or more of the specimens do not show growth, repeat the test with 20 additional units. The additional units all show growth.

Growth promotion of medium after exposure to sterilization conditions— Expose the specified number of units for both the Survival Time and Kill time stated in the labeling, as described in the section Biological Indicator for Steam Sterilization, Self-Contained under Biological Indicators—Resistance Performance Tests

. Incubate the spore strips submerged in the self-contained medium according to the instructions of the manufacturer. At the end of the incubation period confirm the existence of growth in each of the specimens that were exposed for each Survival time and the absence of growth in each of the specimens that were exposed for each Kill time by visual inspection (turbidity or color indicator change) and by separate microscopic examination of each specimen and confirm, where applicable, correspondence of the labeled color to the appearance of growth in the supplied medium.

Ability of medium to support growth after exposure to the sterilization conditions— Take a stated number of units (e.g., 10) after they have been exposed for each Kill time stated in the labeling as directed in the preceding section. Aseptically remove and pool the medium from each unit. Prepare a suspension of the indicator microorganism as directed for Total Viable Spore Counts under Biological Indicator for Steam Sterilization, Paper Carrier. Prepare a dilution of that suspension so as to contain 100 to 1000 viable microorganisms in one mL. Inoculate the pooled medium with enough suspension to contain a total of 100 to 1000 microorganisms in a 10 mL aliquot of not more than the volume from 10 units of the pooled medium. Incubate the inoculated pooled medium as directed for Total Viable Spore Count. Clear evidence of growth is obtained within 7 days.

Disposal— Prior to destruction or discard, sterilize it by steam at 121

for not less than 30 minutes, or by not less than an equivalent method recommended by the manufacturer. This includes test strips employed in any test procedures for the strips themselves.

Auxiliary Information— Please check for your question in the FAQs before contacting USP.

Topic/Question	Contact	Expert Committee
Monograph	Radhakrishna S Tirumalai, Ph.D. Senior Scientist 1-301-816-8339	(MSA05) Microbiology and Sterility Assurance

USP32–NF27 Page 1678