The term immunofluorescence is often confused with immunocytochemistry and immunohistochemistry. All three of these terms are frequently used interchangeably, and this can lead to confusion when looking to purchase an antibody to use in your microscopy experiment.
 

Immunocytochemistry is most commonly performed on cells that have been grown in a monolayer in a culture dish or transferred from suspension to a slide for viewing on a microscope. When working with adherent cells, the cells are often grown directly on a coverslip. Depending on the cell type, the cells may not adhere to the coverslip on their own and the coverslips will need to be coated with gelatin or either poly-L-lysine or poly-D-lysine in order to aid in this process. Suspension cells are usually fixed by adding the fixative directly to the cell culture media for approximately 15-20 minutes, washed, and resuspended in buffer and directly placed on a gelatin or poly-L/poly-D-lysine coated slide, and smeared using the edge of another slide or the side of a pipet tip. Suspension cells can also be immobilized to slides using a centrifugation method known as Cytospin™.

In immunocytochemistry, there are generally two types of fixatives used: cross-linking fixatives such as formaldehyde and paraformaldeyhde or organic solvents such as methanol or acetone. The cross-linking fixatives are usually preferred because they tend to preserve the morphology of the cell better by crosslinking proteins and forming a cytoskeletal network. However, this cross-linking of proteins can also mask epitopes and decrease the ability of an antibody to bind to its target, thereby requiring a permeabilization step so the antibody can ultimately access that target.

The most common permeabilizing reagent used for immunocytochemistry is Triton X-100, ranging from 0.1-0.5% in PBS, but saponin, digitonin, and Tween-20 can also be used. When using organic solvents as the fixative, a permeabilization step is usually not necessary.
 

Contributed by Kellie Johnson..