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Detailed procedure and tips for setting up macrophage cell line cell plates ready for the toxin neutralization assay.
Prepare cell growth medium
Culture media used will depend on cell type used. For example:
|10% FBS||50 mL|
|2 mM glutamine||5 mL|
|100 U penicillin/0.1 mg/mL streptomycin||5 mL|
Use aseptic technique working in a class II safety hood.
Whilst growing, cells should be checked EVERY day to make sure they are healthy and growing as expected. They should be mainly attached to the bottom of the flask, round and plump or elongated in shape and refracting light around their membrane. This indicates they are healthy.
Media should be pinky orange in color. Discard the cells and do not use for the assay if they are detaching in large numbers and/or look shriveled and grainy/dark in color.
100 mL x dilution factor
This will give a value for the volume of media the cells should be in.
If volume required for the correct cell density is less than 100 mL: