I will be treating heparinized whole blood with a small molecule for various timepoints and then extracting RNA for qPCR.
Do people recommend incubations in tubes or plates/dishes? 5% CO2/37C or only 37C? I am set on the RNA extraction part, but fairly inexperienced with the prior steps, so any hints or tips would be well appreciated.
I did several whole blood experiments with tlr stimulants and virus or bacterial infection in whole blood.
After optimization done by the first authors of the linked paper I now always work with corning cell culture plates 96 well, 100 ul whole blood per well with a plastic plate cover incubated at 37C 5% co2
Article Potent Innate Immune Response to Pathogenic Leptospira in Hu...
Dear Stevan,
Usually, when we perform whole blood cultures the incubation is held at 5%CO2/37C. One important thing is the dilution of the blood (at least 1:2 but 1:10 in RPMI is ideal). Eosinophils and neutrophils will last about 6-8 and 16 hours, respectively. Lymphocytes would last up to 6 days. No need for supplementation with FBS (but sometimes antibiotics is desired for longer cultures).
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