I am working with tumor xenografts and preparing single cell suspension! I am currently using 20 mg/ml of collagenase IV, 100 ug of DNase and CaCl2 in plain RPMI medium. I incubate them for 45 min.
When I label my single cell suspension afterwards, CD 115 conjugate fluorescent dyes does not work (I have used two different clone and different colours). In addition to CD 115, CD8 as well as CD 3 and CD4 do not work.
I have also reduced the collagenase concentrations but still it does not work. So I am looking for the reason of it and also what else I can use to dissociate the tumors?
Nicola @ D you know what liberase can be used? In this paper they only mentioned Liberase, but did not mention what kind (TM or TL)?
Hi,
I work with Liberase TH and that increased the viability of my cell in a 40%. I work with knee fat tissue and the total amount of cell that I am able to get after digestión is very low. This reagent helped me a lot.
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