Immunostaining of cytologic preparations: A review of technical problems

Immunostaining of cytologic preparations has not achieved the level of succ ess attained by the same procedure performed in paraffin sections. This def iciency has been largely attributed to the difficulties in interpretation a rising from false positivity, high background staining, and inadequacy of m aterial for immunocytochemical analysis. Many of these setbacks have now be en overcome by recent developments that include the preparation of cell blo cks, rehydration of smears, and the use of a weak fixative. Contrary to the current practice of using ethanol, cold acetone, B5, and other fixatives f or immunocytochemistry, we recommend the preparation of thin, evenly spread air-dried smears (up to 24 h) followed by fixation in 0.1% formal saline ( 2-14 h). Air-drying minimizes the loss of cells and the weak formalin prese rves cellular antigens and also removes proteinaceous fluid as well as caus ing lysis of red blood cells, both of which contribute to background staini ng in cytologic preparations. Brief post-fixation in 100% ethanol (10 M) im proves cytomorphology. Immunostaining is enhanced by the routine applicatio n of heat-induced antigen retrieval. The adoption of all these technical pr ocedures will eliminate the impediments to the realization of the full pote ntial of immunocytochemistry in diagnostic cytology.