Stoichiometry of immunocytochemical staining reactions

This chapter reviewed and presented my biases about the factors that affect our ability to make quantitative measurements of epitope (for this chapter, equal to a protein) with monoclonal antibodies in a flow cytometric system. The discussion has illustrated that the chemical structure of the permeabilized cell and the affinity and specificity of the antibody are the "two" factors that are important. Pursuit of this discipline would be significantly enhanced with highly specific antibodies with a high affinity--higher than we have so far observed with the antibodies against cell cycle regulatory proteins. When moving to multiparametric space, optical systems that mask off interbeam cross talk, primary labeled antibodies, and rigorous use of fluorescence compensation is essential for the highest quality work.