◆ SpookStack

l Heat Stability of SPS 02.266, SPS 02.57.03, and SPS 02.88.01 The object of this study is to compare the heat stability at 70°C in a water bath! of SPS 02.266 spores, SPS 02.57.03 spores and SPS 02.88.01 spores in water for injection, WFI! The spores from the three preparations will be diluted to about 5 X 106 CFU/ml in WFl. A "0 time" sample will be diluted in WFI to approximately 5 X 102 CFU/ml and plated onto Tryptic Soy Agar plates TSA!. The spore preparations will then be incubated for 1, 6, and 12 hours. After dilutions are made in WFI, onetenth-ml aliquots will be plated out for 1:1 .000 and 1:10,000 dilutions of the heated spore preparations. All plates will be incubated overnight at 37°C. Counts will be made and the heat stability will be determined and expressed as the length of time at 70°C required for a 50% drop in viability. Preparations to be examined: Approximate Initial Concentration 1 = SPS 02.266 spores 1.0 X 109 CFU/ml 2 = SPS 02.57.03 spores 6.0 X 109 CFU/ml 3 = SPS 02.88.01 spores 5.3 X 109 CFU/ml Dilution Schemes in tubes ofWFl to get to 5 X 106 CFU/ml!: 1 - SPS 02.266 spores! - 0.1 mlinto 9.9 ml, mix, then 5.0 ml into 5.0 ml 2 - SPS 02.57.03 spores! - 0.1 ml into 9.9 ml, mix, then 1.0 ml into 11 ml 3 - SPS 02.88.01 spores! - 0.1 ml into 9.9 ml, mix, then 1.0 ml into 9.6 ml At each time point hours,1 hour, 6 hours, 12 hours!, dilute as follows: 0.1 ml into 9.9 ml, mix; from this dilution, put 1.0 ml into 9.0 ml, mix; from this dilution put 1.0 ml into 9.0 ml and mix. For each of the last two dilutions, dispense 0.1 ml onto 5 TSA plates. Spread the plates to dryness with a glass spreader, then incubate overnight. Count the colonies on the plates. 4 .