I am trying to decide the best method between flow cytometry or qPCR to be used as I compare Glut1 expression in two different T cell lines. Is there an ideal protocol?
I had read articles where they used flow cytometry, however seems that T cells have to be transfected before. Should I used intracellular staining in instead.
Thanks for your advice.
Flow cytometry but using two or three "colors". At least one T cell surface marker plus Ab against Glut1. Try finding anti.Glut1 already conjugated with fluorophore.
Transfection/transduction is done when you want to overexpress Glut-1 in the cells. If you want to look at the endogenous expression levels of Glut-1 in your T cell lines, you could do either q-PCR (if you are interested in mRNA levels) or Flow cytometry/Western Blot (to look at protein levels). Might be worthwhile measuring both the mRNA and protein levels. If going down the flow cytometry route, I think surface staining would be enough (since the receptor is expressed on surface of cells). Like Rafael mentioned, stain the cells for CD4/CD8 and Glut1 (all fluorescently labelled).The papers below might help.
Article The Glucose Transporter Glut1 Is Selectively Essential for C...
https://www.nature.com/articles/srep24129#results
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