Hello, im trying to visualize surface antigens of lymphocyte cell line cells with primary antibody immunoFluorescence. I cant have a proper protocol about this issue. I followed flow cytometry protocol: pelleting cells, incubating with Fluorescent antibody then fixing, but it didnt work. Do you have suggestions ? do you think these antibodies would work for Fµscopy ? or not at all ? Protocols ? THANKS
Good day!
I think that antibodies for flow cytometry are quite suitable for immunofluorescence. In my practice, I also used antibodies for flow cytometry in an immunofluorescence experiment. see this work (reference), can it help you?
https://www2.vet.cornell.edu/sites/default/files/PrepReagents.pdf
Alexandr Chernov Thanks for you answer and valuable protocols, however i believe i already did something similar to that in my setup. I am talking about imaging, using microscopy do you confirm using these protocols for imaging ?
Ps. I use anti-CD3 conjugated Phycoertherin and aCD4 conjugated FITC to observe T Lymphocytes
Thanks again
PE can be readily bleached by light sources used in microscopy. FITC is more stable. Watch our for how fast PE fades in your set-up. You might make it work, but you may also have to be quick with your photos , or use a more stable fluorochrome.
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