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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

  1. Grow Cells
  2. Fix Cells
  3. Permabalize Cells
  4. Block Cells
  5. Incubate with Primary Antibody
  6. Incubate with Secondary Antibody
  7. Place on a slide

Grow Cells

The key point to be made here is to not have too many or too few cells on the day you harvest the cells.

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