Ethanol disrupts hydrophobic and hydrogen bonding thus exposing the internal hydrophobic groups of proteins and altering their tertiary structure. Ethanol interacts more strongly with hydrophobic areas than for example methanol (so methanol would be better choice to use). Ethanol will cause distortion of nuclear and cytoplasmic detail, it will remove lipids while dehydrating the cells and also precipitate proteins on the cellular architecture.

On the other hand, fixation can be done using crosslinking reagents, such as paraformaldehyde (as suggested above). These are better at preserving cell structure, but may reduce the antigenicity of some cell components as the crosslinking will obstruct antibody binding.

I use e-bioscience and/or BD Fix/Perm kit, which consists of formaldehyde for fixation and saponin for permeabilization.

Maybe this paper will help you: http://www.jbc.org/content/273/45/29811.full

Hi, ethanol can affect the structure of proteins on cell membranes that can prevent their recognition by antibodies. Usually using ethanol is not the preferred agent to fix cells. PFA 4%, glutaraldehyde 1% or cold methanol are most commonly used to preserve membrane integrity and insure a good immunostaining. Lipidic dies are working to stain the membranes after ethanol fixation if you are looking for something non specific. Good luck! FS

EtOH certainly affects the structure of some proteins. I just tested with GFP after 70%EtOH fixing cells expressing GFP, almost 90% signals were gone. Cold methanol or 4% PFA are commonly used for immunostaining.

The ethanol will likely remove significant quantities of membrane lipids. See here for a more detailed discussion. http://lipidlibrary.aocs.org/topics/extract2/index.htm

We have used cold methanol to permeabilzing cells sufficiently to label with 30 nm nanoparticles that are completely excluded from PFA fixed cells.

If possible, try another fixation method, I would say 5-10% paraformaldehyde, especially if it is lipids that you are concerned about. The ethanol might remove some lipids from the membrane and perhaps also all sorts of lipophilic substances. Theoretically, this much ethanol might be especially harmful to the endoplasmic reticulum, of which the membrane contains very little cholesterol (which strenghtens any membrane)...

70% ethanol produce dehydratation on the cell membrane by removing water since it has more affinity

Ethanol disrupts hydrophobic and hydrogen bonding thus exposing the internal hydrophobic groups of proteins and altering their tertiary structure. Ethanol interacts more strongly with hydrophobic areas than for example methanol (so methanol would be better choice to use). Ethanol will cause distortion of nuclear and cytoplasmic detail, it will remove lipids while dehydrating the cells and also precipitate proteins on the cellular architecture.

On the other hand, fixation can be done using crosslinking reagents, such as paraformaldehyde (as suggested above). These are better at preserving cell structure, but may reduce the antigenicity of some cell components as the crosslinking will obstruct antibody binding.

I use e-bioscience and/or BD Fix/Perm kit, which consists of formaldehyde for fixation and saponin for permeabilization.

Instead of asking a generic question, you may want to ask a pointed question. What kind of read-out you want to do after 70% fixation? For example, some of the apoptotic DNA fragmentation assay may not be affected by ethanol, whereas it will affect other assays. Thus, you may want to select your application first just as immunohistochemistry, apoptosis, flow cytometry etc. Then select your fixative. That way you aim is achieved.

Ethanol at that concentration will cause significant damage to cells. Ethanol swabs are pretty effective at killing most types of cell.

Rahul, you got some very good comments above. I would just advise to check for your particular application testing in parallel ethanol, methanol and PFA. Methanol has the advantage that it's used cold, helping to preserve some membrane structures. PFA can be used at concentrations lower than 4%, but you need to extent the incubation time, and this need to be tested experimentally. You may look to a study on membrane bound IL1a done on mouse splenocytes (Villacres-Eriksson, Clin Exp Immunol 1993, 93:120), where we used 1% PFA to obtain a very mild fixation. Hope this is of help....

Thank you all for the very informative comments.

My answer is a little late but might still be of interest. Please read the article of U. Schnell (http://www.nature.com/nmeth/journal/v9/n2/full/nmeth.1855.html).

GFP labeled proteins may be found at totally different sites in your cells after fixation. The lower GFP signal may also be explained by this effect.

Can I use 90% Ethanol to fix cells, when later I have to capture RNA binding proteins from these?

Good evening resepectable everyone,

I need some advice at once very very much 

I will try an assay which is about  siımultaneous calcium  mesaurement by flow cytometry. But there is a problem. I dont have fluo-3 AM but instead I have fluo-3 impermeant potassium salt. I dont have enough time and credit to get fluo-3 AM. and ı have to try that assay. Now, what would you do if you were me ? ı have tried to permeabilise the cells with cytoperm cytofix and I think it did work but ı cant be sure to adjust an appropriate assay protocol. I mean what would you really do to measure the calcium simultaneously with fluo-3 impermeant satl. By the way, I dont have any other techniques like microinjection or scrape loading. I mean the only way is to use fluo-3 salt form. Keep forward to hearing from you honourable ladies and gentlemens.

Best regards