Paraformaldehyde (PFA) has been widely used as a cross-linking fixation agent. It has been empirically recognized in a gold standard protocol that the PFA concentration for cell fixation, C _PFA, is 4%.

Why do we use paraformaldehyde?

Paraformaldehyde causes covalent cross-links between molecules, effectively gluing them together into an insoluble meshwork. The reason cells must be fixed prior to immunostaining is quite simple. You need to permeabilize cells to allow antibodies to access intracellular structures.

Why is paraformaldehyde used instead of formaldehyde?

Paraformaldehyde (chemical name is polyoxymethylene) is a powder of polymerized formaldehyde that by itself cannot fix tissues. ... Methanol is added to slow down the polymerization to formaldehyde, which reduces the fixing power of formalin. Formalin can also be made in an alcohol-free form from powdered paraformaldehyde.