How to pay attention to cell culture technology

1. Before the experiment, sterile room and bench (laminar flow) of 30 to 60 minutes with uv lamp radiation sterilization, sterile operation with 70% ethanol to wipe up the surface, and open the bench fan operation after 10 minutes, to begin to experiments.

Only one cell line is processed at each operation, and the medium is not Shared, even if the medium is identical, to avoid confusion or intercellular contamination.

After the experiment, the experimental items were taken out of the workbench, and the sterile operation was lifted by 70 % of the bacteria.

The operation interval should allow the sterile operation station to operate for more than 10 minutes, then the next cell line operation.

2. The working area of sterile operation should be kept clean and spacious, necessary items, such as test tube rack, straw absorbing device or suction box can temporarily placed, such as other test items is finished should be removed, to facilitate air circulation.

The experimental supplies were cleaned with 70 % of the water before they were brought into the sterile counter.

The experimental operation should be carried out in the sterile area of the center of the carrying surface, not in the non-sterile area of the edge.

3. Carefully use sterile laboratory articles to avoid contamination.

Do not touch the top of the pipette or container, or operate the test on the open container.

To handle the clips on the container, cap and holding the bottle, tilt Angle of about 45 ° access, as far as possible don't place the cap opening up on the desktop.

4. The staff should pay attention to their own safety. They should wear the lab coat and gloves before carrying out the experiment.

The cell lines from human or viral infection should be carefully handled and the appropriate level of aseptic work station (at least Class II) should be selected.

In the process of operation, aerosol should be avoided, caution *** *, such as DMSO and TPA, and avoid the injury of sharp needle.

5. The following items are tested regularly: 5.1. CO2 pressure of CO2 cylinders 5.2. CO2 concentration, temperature and water disk of CO2 culture tank (water table of water shall be replaced weekly with sterile water).

5.3. Airflow pressure in the sterile operating table, periodically changing the uv lamp tube and the HEPA filter membrane, pre-filter (300 hours/pre-filter, 3000 hours/HEPA).

6. The sink can add disinfectant (Zephrin 1:750) and replace the water of the sink regularly.

7.Seriously in accordance with the procedures for experiments, generally do not lead to pollution, in many cases is used as the reagent or medium pollution and make the experiment failed, after been powder culture medium preparation (serum), in 4 degrees commonly try to not more than a month, such as storage time can be longer - 20 degrees, but also had better not more than 3 to 4 months, may be not too high requirements for immortalized cell lines, but I in the past four years has been the primary cell culture, cell delicate, experience has shown that placing time shoulds not be too long.

8. Many plastic products can also be sterilized with high pressure, such as the cultivation of bottle caps, glue plugs, and suction first class.

Usually cannot be used for high pressure have been made into a one-off effect of the goods, of course, if the money is limited, such as the cultivation of the imported plate, dish, etc., can also repeat use 1 to 2 times, I at that time is a method to use after completely clean, open under ultraviolet light irradiation before use 1-1.5 h, I have done many times, no problem.

Because of the disinfection and disinfection of plastic culture bottle, it is better not to reuse it. If it is necessary to reuse it, it can be used for disinfection of ethylene oxide.

Under light microscopy, epithelial cells usually show "aura" sample configuration, wiredrawing phenomenon between cells (i.e., far away the cell can be linked together through a slender tentacles), the growth of the cells have piece features.

Mesenchymal cells are usually fusiform and have no growth characteristics and are not closely related to each other.

However, the morphological characteristics of the cells can be altered by changes in the growth conditions, such as HeLa cells, which are epithelial cells, but in the condition of acid culture, they become fusiform.

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