I performed this assay many times but without any consistent result. I usually isolate mouse spleen CD4+CD25+ cells, seed 100K non-activated cells on 3um-pore size membranes (24well plate, corning) in RPMI + 0.5% HI-FBS, with my chemoattractant in the lower chamber, 6h or overnight migration at 37 degree. The number of migrated cells is extremely varied (few hundreds-few thousands) and the results as well.
I want to try coating the membranes with fibronectin (or collagen) to improve migration. Does anyone know the optimal concentration of fibronectin (or collagen) for Tregs? I've found only few works using 10ug/mL.
Abdelkader Dehbi
this paper can help you
Article Technical Advance: New in vitro method for assaying the migr...
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