Hi, I'm planning to quantify cells with different receptor expressions in stem cells throughout the 21 cell culture phase. Can I fix cells in 4% formaldehyde on selected days and conduct the flow cytometer in one day?
Thank you
It depends on what you are trying to measure. Some molecules are more durable than others. Preferably, you can ask your supervisor to help you find scientific data - aka, if someone has done this before - so you will be able to follow an established protocol. If this is not available, I would always recommend trying it out first with a few batches, to see if it makes any difference. This means, that you would measure every day and fix an aliquot for measurement at the end of the week. The same goes for keeping the samples cool. Only when you have established that your results are reliable, you may use them in your research.
Hello J.W.Nirmani Yasara I think you mean slide-based cytometry since you are considering to fix cells on slides not flowcytometry that requires a cellular suspension. The answer is yes you can, and it has been done before, see (PMID: 16527301). Also, if you said slides by mistake, then yes also suspensions of fixed cells can be made but not stored at room temperature depending on duration of storage and fixative used you may need to put them in the fridge or -20 (consult : https://bitesizebio.com/22141/fixation-and-flow-cytometry/, and https://www.cellsignal.com/learn-and-support/protocols/protocol-flow). Good luck
I concur with G. Weckman; it definitely depends on which antigen you are looking at.
From my own experience examining hematopoietic stem cells from bone marrow and cord blood, formalin fixation does not affect the ability to stain the cells for either CD45 or CD34 expression, but it completely abolishes all CD38 staining. This is known as fixation sensitivity.
If the antibody binding site on the antigen of interest contains a lysine residue, it is highly likely that formalin fixation will affect the ability of the monoclonal antibody to bind to it's target and stain the cells.
- Badges
- Science topic