I am interested in cell fixation methods not using aldehyde, alcohol, or acetone for preparing cells for immunofluorescence microscopy and flow cytometry.
That's a tough one. What about chemical cross linkers with reactive primary amines, carboxyls, sulfidryls or carbonyls? I've used them to cross link exogenous molecules to cell surface but I belive that in a normal cell they would cross link cell receptors creating a fixation mechanism (similar to PFA).
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I agree with Andre Fernandes - that is tricky. I don't know of anything else that will work for flow (not to say there isn't anything, I just haven't tried it), but for immunofluorescence, you could do cryofixation and avoid chemical fixation entirely.
cryofixation works for EM only without post-fixation by conventional way.
you can inject pre-stained primary into a cell. the amount is critical, i think. i never try it.
A mix of glutaraldehyde and formaldehyde should do the trick. Formaldehyde will cross-link fast but reversible and the glutaraldehyde will enforce the fixation permanently. Should work for both FC and EM. Just wash excess fixative away before adding antibodies.
the question whether is the target to leave out dehydration and aldehyde base cross-linking as method completly? if that is, there is no answer according to my best knowledge. (considering that later one wants to stain by abs).
It depends a lot of what you want to do. In example, cryosections without fixation work in IF (might need very short incubation in 2% aldehyde and then very good wash – air dry slides in RT when come from -80°C). For culture cells use Fisher coverslips and grow your cells on top (for suspension cells use cytospin) . Wash cells well (3x10min PBS) and then depends a lot of your targets and experiment. If, you do not want to use 1st and 2nd Abs, best is to use fluo-target in culture, then wash well coverslips and air-dry. Mount and you get nice IF data. Keep in -20°C.
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