1. experiment material
(1) Bacillus stearothermophilus paper (a set of test tubes sold by 3M Company is also available). Bacillus stearothermophilus.
ATCC7053 is a biological indicator bacteria commonly used in this method, containing a 5-cell CFU/tablet.
(2) 121 ° C pressure steam sterilization chemical indicator card (common sterilization indicator tape is also available).
(3) Bromocresol purpura water medium was autoclaved at 116 ° C for 20 minutes and then used (such as the test tube sold by 3M Company, this medium may not be used).
(4) O-150 °C residual thermometer (calibration required by the metrology bureau).
2. Method and result
The Bacillus stearothermophilus paper (hereinafter referred to as the bacterial tablet) was placed in a sealed test tube with a sterile forceps. The chemical indicator card and the spot thermometer are placed in an open test tube. Prepare 5-10 parts of each of the above two tubes. Place them at the autoclave vapor port, the bottom exhaust port and the bottom water outlet or at the center of the upper, lower, left and right. If the sterilizer is on the second floor, you need to put 10 places.
The remaining thermometer can be used for verification test after it has been standardized. Before testing, the mercury column of the thermometer should be smashed to below 40 °C. The temperature difference of the thermometer after each monitoring should be between 1 °C, indicating that the temperature distribution inside the sterilizer is uniform.
The sterilized plaque should be cultured at 56-60 ° C for 24-48 hours under sterile aseptic conditions under sterilized bromocresol purine water medium to observe the color change. If the medium turns yellow, the B. stearothermophilus in the tablet is not completely inactivated, and the bacteria can still grow in the medium, and the glucose is decomposed to produce acid to yellow. If the color of the medium does not change, it is still purple, indicating that the spores have been inactivated. At the same time, unsterilized paper was placed in the medium as a positive control, and blank medium without paper was used as a negative control.
The indicator color on the chemical indicator card changes from light yellow to black during autoclaving. As the color changes in depth, and compared with the control color, it can be judged whether the sterilization effect meets the requirements. The chemical indicator card should be stored in a dry place. It will change color when exposed to moisture, which will affect the observation of sterilization effect.
High-pressure steam sterilization must allow steam to enter the sterilizer smoothly, contact with the sterilized items, and discharge the original cold air to achieve the sterilization effect. The no-load heat distribution and full-load thermal penetration verification are to be performed (not more than 2/3 of the total volume at full load). The two verifications were repeated three times for a total of six times.
Five points and six tests showed that the temperature was at 121 ° C, the chemical indicator card turned black; the degree was consistent with the control color; the medium did not change color, indicating that the high pressure steam sterilization effect was qualified.
Autoclaves are powerful instruments, but strong inspections are only an examination of the physical parameters of the instrument. Therefore, the sterilizer that has been subjected to the strong inspection must also verify the sterilization effect. Some units often overlook this important issue. In domestic portable sterilizers, the instrument pointer often reaches the required temperature, but the actual temperature of the sterilized article does not meet the requirements. Lead to sterilized items is not thorough, affecting the results of the experiment. The medium is not sterilized thoroughly, affecting the use of the medium and judging the results. Therefore, the verification of the aseptic effect of the autoclave is a problem that must be taken seriously.
When using a high-pressure steam sterilizer, first take care to remove cold air from the sterilizer while opening the vapor. The vent hole must be closed before all the cold air in the unit has been removed.
If some air remains in the sterilizer, the pressure gauge has reached a certain pressure value, but the temperature inside the device has not reached the corresponding degree. The more air that remains, the greater the difference between the two, the temperature inside the device is insufficient, and the sterilization is not complete. (Many people will have bubbles in the small catheter when they are sterilized in the fermentation medium. It may also be possible to try to discharge more cold air from the sterilizer. It has a miraculous effect).