Sterile operation precautions - Huaqiang Electronic Network

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【operating】

When cultivating, the action should be accurate and agile, but not too fast, to prevent air flow and increase pollution opportunities. Do not touch the disinfected utensils with your hands. If it has been touched, use a flame to sterilize or replace the spare parts. For convenience, the supplies on the work surface should have a reasonable layout. In principle, the right hand should be placed on the right side, the left hand product on the left side, and the alcohol lamp on the left side. Work must be kept in order from beginning to end, and tissues or cells should not be exposed to the air prematurely before they are treated. Similarly, before the culture solution is used, do not open the bottle too early; if it is not used again after use, immediately close the mouth of the bottle to increase the chance of falling. When extracting nutrient solution, PBS, cell suspension and other various liquids, separate pipettes should be used, and should not be mixed to prevent contamination or cell cross-contamination. In the work, you can't talk or cough in the field of operation, so as to prevent the bacteria from bringing bacteria or mycoplasma into the work surface. Wipe the clean bench top and hands with alcohol before handling. Hands or relatively dirty items cannot pass over the open bottle mouth. The bottle mouth is most susceptible to contamination. If the tip of the pipe touches the bottle mouth when adding liquid, the pipette should be discarded.

[Preparation before training]

Make experimental plans and operating procedures before starting the experiment. The calculation of the data must be done in advance. According to the requirements of the experiment, prepare all kinds of required equipment and articles, put them into the operation place (cultivation room, clean bench) after the inventory is correct, and then start disinfection. This can avoid after starting the experiment. Prison items are not fully taken back and forth to increase pollution opportunities.

[Operation field disinfection]

The sterile culture room should be used to mop the ground with 0.2% Xinjieer every day (the mop should be dedicated). Ultraviolet radiation disinfection for 30-50min, ultra-clean workbench countertops should be scrubbed with 75% alcohol before each experiment. Then UV poisoning for 30min. When disinfecting the countertop, do not expose the culture cells and the culture solution to ultraviolet rays at the same time. Do not place too much or overlap on the countertop when disinfecting. Otherwise it will block the radiation and reduce the disinfection effect. Some operating utensils such as pipettes, waste liquid tanks, dirt boxes, test tube racks, etc. are scrubbed with 75% alcohol and placed in the table while UV disinfecting.

In vitro cultured cells lack anti-infective ability, so prevention of contamination is the primary condition for determining the success or failure of culture. Even with a well-equipped laboratory. If the experimenter is careless and the technical operation is not standardized, it will also lead to pollution. thus. For the utmost guarantee of sterility in all operations. Every job must be done in an orderly manner and completely.

[Washing and dressing]

In principle, it is the same as surgery. Usually only observe and do not do the cultivation operation, you can wear the clean clothes of the cell culture chamber for 30 minutes. When working with the clean bench, the entire forearm should be inserted into the box, and the long-sleeved clean overalls should be used, and the hands should be disinfected with 75% alcohol before starting the operation. If the hand touches potentially contaminated items and enters the culture room during the experiment, wash the hands again with disinfectant. To enter the primary culture room, you need to wash your hands thoroughly and wear a mask and a sterile coat.

[flame disinfection]

When culturing in a sterile environment or doing other aseptic work, first ignite an alcohol or gas lamp. All subsequent operations, such as loading a straw cap, opening or closing the mouth of the bottle, etc., are required to be in the vicinity of the flame and subjected to cauterization. However, it should be noted that the metal equipment can not be burned in the flame for too long, in order to prevent annealing, and the burnt metal crucible should be cooled before the tissue can be taken to avoid tissue damage. The straw after the nutrient solution has been absorbed can no longer be burned by the flame, because the nutrient solution remaining in the pipette head can be charred to form a carbon film, and when used, the harmful substances are brought into the nutrient solution. When the cell culture flask is opened or closed, the flame sterilization time is short. Prevent cells from burning out due to excessive temperature. In addition, when the rubber plug passes through the flame, it cannot be used for a long time, so as to avoid burning charred gas and damaging the cultured cells.