Immunocytochemistry (ICC) is the process of taking tissue culture cells and applying antibodies to determine the spatial and temporal features of a particular antigen (often protein). A survey of the literature reveals many different techniques. However, there is often very little comment about the individual steps of the techniques, and hopefully this page will lead to a productive discussion. Often different techniques applied to the same antibody will reveal different results.
Immunofluorescence (IF) is a subset of ICC in which the antibody localization is visualized by fluorescence. The below protocol is focused on indirect IF (the use of a secondary antibody).
The Seven Basic Steps
- Grow Cells
- Fix Cells
- Permabalize Cells
- Block Cells
- Incubate with Primary Antibody
- Incubate with Secondary Antibody
- Place on a slide
The key point to be made here is to not have too many or too few cells on the day you harvest the cells.