Immunstaining cells fixed in 10% formalin- possible? - (Dec/02/2008 )

I have been given slides with cells fixed in 10% formalin (slides were dipped in this fixation).
Now my job is to immunostain the cells. I have tried and tested many Immunofluorescence protocols
and now am wondering if it is even possible to immunstain these cells fixed in 10% formalin (not IHC, but specifically IF, since this is not tissue, but cells). 1% formalin may have been a better fixation method?
But I am not sure, just speculating...

Has anyone used immunofluorescence on formalin fixed cells ?

Thanks for any feedback in advance!

I have stained cell suspension or FCM with 1%FA and worked well. But, first you will have to check if your antigen is not disturbed with fixation.

Thanks for your reply Bungalow Boy. What I am really concerned about is the use of 10% formalin fixed cells compared to 1% formalin fixation?

I have to do immunofluorescence on 10% formalin fixed cells (not tissues) and was wondering
if this % formalin was still stainable and if so with what protocol....
I have tried many....

I have been given slides with cells fixed in 10% formalin (slides were dipped in this fixation).
Now my job is to immunostain the cells. I have tried and tested many Immunofluorescence protocols
and now am wondering if it is even possible to immunstain these cells fixed in 10% formalin (not IHC, but specifically IF, since this is not tissue, but cells). 1% formalin may have been a better fixation method?
But I am not sure, just speculating...

Has anyone used immunofluorescence on formalin fixed cells ?

Thanks for any feedback in advance!

we use 3% formalin. I guess the only thing you can do is give it a go, maybe with an antibody you know works very well. 10% might overfix, but I'm sure you will find out quick smart if it has or not. How long were they fixed for?

I used 1%FA for fixation for FCM but am not sure for Immunoflourescence Microscopy.

Googled for protocol and found this

http://www.sfu.ca/~mbbweb/microscopy/pdf/pfa.pdf

This protocol speaks of 3.7% PFA which should be same as 10% Formalin. So the protocol should be OK.

QUOTE (NKB @ Dec 3 2008, 10:24 AM)

I have been given slides with cells fixed in 10% formalin (slides were dipped in this fixation).
Now my job is to immunostain the cells. I have tried and tested many Immunofluorescence protocols
and now am wondering if it is even possible to immunstain these cells fixed in 10% formalin (not IHC, but specifically IF, since this is not tissue, but cells). 1% formalin may have been a better fixation method?
But I am not sure, just speculating...

Has anyone used immunofluorescence on formalin fixed cells ?

Thanks for any feedback in advance!

I have just started my experiments on immunofluorescence. Gone through many references and protocols (--never seen 10% formalin, correct me if I am wrong), and seleted 3.7% formalin (PFA/PBS; pH 7.2) for 10 min at RT. It worked perfectly to my epithelial cells.

Just sharing my own experience, really cant say what if 10% formalin is there??

If you are particularly worried about background, you can wash several times in 0.1 M glycine in PBS. The glycine should mop up any free aldehydes from the formalin.

there is a common misunderstanding, when it comes to formalin - in fact, "formalin is a saturated solution of formaldehyde, water, and typically another agent, most commonly methanol. In its typical form, formalin is 37% formaldehyde by weight (40% by volume).." So, it is the same solution, but concentration values in % differ. 10% formalin - 3.7% fomaldehyde
In some protocols the word "formalin" may be even misused instead or formaldehide without substitution of the corresponding concentration values...
So, may be your slides are ok? did you ask the person, who gave you the slides, about the mode of fixation?
Honestly, I have never done immunohistochemistry, just guessing - did you try the heat retrieval of antigens on your slides?