Glass pipette preparation and sterilization experiments
Glass pipette preparation and sterilization experiments
Both glass and plastic pipettes can be used for tissue culture. The advantage of plastic pipettes is that they are single-use in a single package and do not need to be cleaned or sterilized. Glass pipettes are very inexpensive, but each time they are used, the cotton stopper must be removed and replaced with new cotton, and they must be cleaned very carefully with a pipette cleaner to avoid clogging the pipette with residual oily substances. Source: Animal Cell Culture: A Guide to Basic Techniques, Fifth Edition
Operation method
Scheme 11.2 Experiment on preparation and sterilization of glass pipettes
Materials and Instruments
Pipette cylinders Hypochlorite Sterile indicator tape Sterilization oven Move I. Materials Sterilization For more product details, please visit Aladdin Scientific website.
1. Pipette cylinders (for collecting used pipettes)
2. Disinfectant: hypochlorite, diluted with detergent and containing a minimum of 300 μg/g chlorine (e.g., sodium hypochlorite for routine use).
3. cleaning agents (e.g. 7 X or Decon)
4. stainless steel basket (to collect cleaned pipettes and dry them)
5. sterility indicator
6. Pipette jars (square aluminum or stainless steel containers with silicone pads at each end; square jars do not roll) (Thermo Electron)
7. sterile indicator tape or plugs (Bennett)
8. Sterilization oven: temperatures of up to 160°C can be reached with a fan, which is preferable to the use of recordable thermometers and bendable probes.
II. Procedure
Collection and cleaning
1. Pour the cleaning agent containing the disinfectant into the cylinder containing the pipette.
2. Remove the pipette tip after use and dispose of the pipette in the cylinder immediately (Figure 11.5).
(a) Do not place the pipette in the laminar flow hood and do not allow it to dry out.
(b) Do not place pipettes for sucking agar or silicone (water repellents or anti-foam agents, etc.) in the same cylinder as normal pipettes. Silicone can be sucked with disposable pipettes, and pipettes after sucking agar should be washed with heated tap water, or disposable pipettes can also be used.
3. Pipettes should be soaked overnight or for at least 2 h. If pipettes are used in large quantities, the filled cylinders should be retrieved from time to time and soaked for 2 h prior to addition of cleaning solution.
4. Remove the cotton plugs by compressed air after soaking.
5. Place the pipette in the pipette cleaner with the tip facing upwards (Figure 11.4, Figure 11.5, see also Figure 4.6).
6. Wash the pipette with tap water by siphoning through the cleaner for at least 4 h. Alternatively, the pipette can be cleaned with an automatic cleaning machine with a pipette connector, but without detergent.
7. turn the valve to deionized water (DW) or reverse osmosis water (ROW) (Fig. 4.6), or wait until the tap water rinse is complete, then fill, empty and repeat the rinse 3 times with DW or ROW (rinse cycle with automatic deionized water in a glassware washer).
8. Place the pipette in a pipette dryer or oven with the tip facing up.
9. Plug the mouth of the tube with cotton (Figure 11.6).
10. Sort the pipettes by size and store in a clean environment.
1. Place the pipettes in the canister containing the pipettes (it is very important to label the ends with the size of the pipettes).
2. Fill each jar with pipettes of the same size.
3. a few jars with 1 ml, 2 ml, 10 ml, 25 ml pipettes (e.g., 4 per model).
4. apply sterile indicator tape between the lid and the jar and write the date on the tape.
5. Dry heat sterilize at 160°C for 1 h. Use as little tape as possible or switch to a temperature-indicating label (Alpha Medical; Bennet), which is small and non-explosive. Check the temperature in the center of the oven to ensure that minimum sterilization conditions are met in this most difficult to reach area. Ensure that there is some clearance between the jars in the oven so that hot air can circulate (Figure 11.7).
6. Remove the pipettes from the oven, cool them and take the jars to the tissue culture laboratory. Pipettes can be left for more than 48 h, but tape along the perimeter of the jar lid.