I am planning to isolate human monocytes from buffy coat and wondering how well isolated monocytes can grow in cell culture? if I need more than 4.5 million cells per flask, how much blood is needed to isolate a certain amount of monocytes in order to expand them into this cell amount?
Hello,
Isolated monocytes do not survive for long in culture. Nevertheless, you can try using the protocol mentioned in the research article given below. May be it might help.
https://doi.org/10.1189/jlb.0513272
Best.
One quick and easy way to isolate monocytes from PBMCs is to use immunomagnetic negative selection cell separation.
Hello Saikal, We required monocytes for cytokine differentiated dendritic cells for a trial. We found that you could recover about 10% monocytes(105 cells/ml) relative to the number of PMBC(106 cells/ml) from fresh whole blood following density separation. However, numbers dropped off as the blood aged, as did functional ability. Blood older than 24hours after collection resulted in less than 5% monocyte recovery, poor differentiation and low growth. Fresh monocytes are easily isolated by a simple 2-4 hours incubation of isolate PBMC under standard culture conditions - lymphocytes are removed by recovering non-adherent cells by gentle washing. Better than 95% adherent cells CD14 positive. CD14 contribution verified by flow cytometry for both adherent and non-adherent wash off
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