Spore strips

Aseptic transfers are also avoided by the use of self-contained units where the spore strip and nutrient medium are present in the same device ready for mixing after use. 

The cold spots recognized during the temperature investigation (worst case) are investigated again by placing a spore strip near each thermocouple location which has been proved to be a cold spot. Besides, the two spore strips shall be used as a positive control. 

Spore strips or spore suspensions are used in the validation studies. The number of micro-organisms per strip or per ml of suspension must be as accurately known as the D value. 

After the cycle, aseptically transfer the spore strip to vessels of culture media. If spore suspensions were used, aseptically transfer the inoculated bottles to a laminar airflow workstation and add culture media to the bottles. Use appropriate positive and negative controls. 

Determine the D value of biological indicator spore strips placed within the product. Determine the location of the lowest radiation dose point within the product. Then determine the dosage required for a 10 6 probability of nonsterility for the product. 

Culturing of Test and Control Samples. Both control and treated spore strip samples were cultured at 37 °C in trypticase soy broth for 48 hr to determine kill. Swab samples, taken from sections of the surface of the muddy-moldy pages before and after exposure to ethylene oxide, were placed in 1 mL of 0.001M phosphate buffered saline solution at pH 7.0 for plating on appropriate media. These samples were taken from a 6.45-cm2 section. 

Series II. In this series of experiments both spore strips and muddy-moldy pages were used, some placed in books which were partially opened and set on their bottom edges while they were being treated. Moisture was added to some sections of the completely dried, contaminated pages. The results listed in Table I show that spores of B. subtilis were not killed in 15 hr when located inside closed books. When a book was left partially open to permit ready access of ethylene oxide, all spores were killed. 

Table I. Treatment of Spore Strips at Room Temperature...

Time (hr) Location of Spore Strips Unexposed Strip Exposed Strip... 

Table IV. Treatment and Evaluation of Spore Strips at Elevated Temperature...

Sterilization of spore strips and muddy-moldy pages in books treated at room temperature was accomplished in 15 hr when the books were set on their edges and left partially open. The importance of sufficient moisture is obvious from the results of treating the muddy-moldy page for 4 hr after being dampened. A convenient practical way to ensure the supply of the necessary amount of moisture to each biological site on each page still needs to be developed. 

For other materials (for instance, equipment and supplies for aseptic manufacture) where the mechanisms of inactivation rely on direct contact between the steam and the item being sterilized and therefore where air removal is matter of importance, the choice of substrate for Bis generally lies between using commercially available paper spore strips and the material itself The decision tree in 4 may be helpful. Regulatory pressure is currently toward use of inoculated product, but commercially available spore strips are more convenient. Tailor-made inoculated product requires substantial amounts of microbiological expertise. The decision tree in Fig. 4 may be helpful in selecting which approach is best in particular circumstances. 

During microbiological challenge studies, all Bacillus stearothermophilus spore strips/suspensions must show negative test for the growth of B. stearothermophilus. 

During Microbiological Challenge studies, spore strips/suspension quantification test must indicate that the population of each manufacturer s lot of spore strips/suspensions is within 50% of their labeled population. 

Commercially available spore strips arc usually intended to have 10 viable recoverable spores per strip. They may be loaded onto carriers by indt-... 

Of the three types of carrier, the spore strip is the most commonly used. Once this choice is made there are several other essential decisions. 

The number of spores per strip and the number of strips per load should for validation purposes be related in some way to the bioburden of the product. What type of relationship this should be is less obvious. There are two broad approaches to this the first is to relate the microbiological challenge on each spore strip to the average bioburden on individual products and the required level of sterility assurance (SAL) the second is to relate the total microbiological challenge in the sterilizer load to the total bioburden within the sterilizer. 

If the number of spores per load is to be related to the total bioburden of the load, the average bioburden per item must be multiplied by the number of items in the load to give an estimate of the total bioburden in the load. The number of spore strips to be used in validation may be calculated by dividing this number by the number of spores per biological monitor thus... 

If the number of spores per spore strip and the number of spores per validation load is to be related to the individual product item and a sterility assurance level of 10. h is first necessary to know the average number of contaminants per product item. In the interests of conservatism this number may be rounded up or supplemented by a safety factor. The target is to use as many spore strips as is necessary to provide assurance that this bioburden is being inactivated to an SAL of 10 This can be calculated from... 

The number of spore strips per load can be obtained by dividing this figure by the number of spores per strip. 

Given that the error term is composed of e, i + 4, the MSe calculation may not be appropriate. Only three hip joints were available and were reused, over time, for the testing. At time 0, spore strips (without hip joints) were heat shocked (spores stimulated to grow by exposure to 150°C water) and the average value recovered was found to be 1.0 x 10 . Then, spore strips attached to the hip joints underwent 1,2,3,4, and 5 min exposures to steam heat in a BIER vessel. Table 3.2 provides the spore populations recovered following three replications at each time of exposure. 

The time taken to sterilise materials depends on the load, but typically loads made up of relatively small volumes would be treated adequately within 20 minutes. Loads containing large volumes take longer and suitable times can be determined by the use of thermal probes or spore strips (see page 86). 

An important medium for the growth of B. stearothermophilus is Spore Strip Broth. Strips containing spores of B, stearothermophilus are frequently placed in autoclaves to determine their efficiency. When removed after autoclaving, these strips are placed in the broth and incubated at 55 °G for 7 days. Growth indicates that the autoclaving process was deficient. 

Dry heat is generally used at 180°C for a period of two hours for the sterilisation of glassware, metal instruments and thermostable products. Spore strips of Bacillus stearothermophilus may be placed in the oven as a biological sensor, and removed for culturing at the end of the heating period. Growth indicates that the sterilisation process was inadequate, whilst no growth shows a suitable temperature was maintained for the requisite time. 

Table 6.1 Effect of RCS conditions at 5 kGy dose on spore strips and residual formaldehyde using Celcon M-90 powder...

Celcon M-90 (mg) Dose rate (kGy/h) Spore strip (cfu/mL) Spore killing (log reduction) Residual CH Olpg) Suture sterility... 

Suture Gamma dose (kGy) Residual formaldehyde ( jg) Spore- strip colony- forming count Verified suture sterility Suture properties ... 

Formaldehyde concentrations can be estimated chemically, although for validation purposes a spore strip assay is commonly used. 

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