serological testing

Summary

Different lots of serum have different abilities to support cell growth, especially clonal cell growth. Testing different batches of serum and then purchasing serum in bulk not only saves money but also reduces variations in experimental conditions, as some batches of serum may contain toxic or growth-inhibiting compounds. This experiment is based on the Cell Lab Guide (previous volume) by Peitang Huang.

Operation method

serological testing

Principle

Different lots of serum have different abilities to support cell growth, especially clonal cell growth. Testing different batches of serum and then purchasing serum in bulk not only saves money but also reduces variations in experimental conditions, as some batches of serum may contain toxic or growth-inhibiting compounds.

Move

1) Obtain 100ml batches of test serum from different manufacturers.

2) Test the serum with 2-3 types of cells commonly used in our laboratory.

3) Add lml of culture medium containing 20% of the above sera to 9 petri dishes (35-mm), and use 9 dishes for each batch of serum.

4) Inoculate 500, 1,000, and 2,000 cells in each 3 of the 9 dishes. If the cell survival rate is high, the number of cells can be lowered. Cells do not have to grow to full size, but should grow to form a sufficient density of individual clones. Cells should be diluted in serum-free culture plants to avoid residual blood淸remaining in the existing culture medium. Cells should be added to a volume of 1 ml to achieve a final serum concentration of 10%. The existing serum is used as an internal standard for this protocol.

5) Prepare 80 ml of culture medium containing 10% of the serum to be tested. Use this culture medium to change the cells twice a week for two weeks. Cloning should be evident during this period.

6) Perform clonal staining to compare the number of clones in different batches of serum. Staining of clones is done by rinsing with 1xPBS, draining the PBS, adding 2 ml of Wright's staining solution (Fisher Scientific), staining for 8 minutes at room temperature, adding 2 ml of water, and rinsing with water for 10 minutes at room temperature to observe the number and size of clones.

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