The effect of the antibody used and method of quantification on oral mucosal vascularity

A vast literature exists on angiogenesis particularly with regard to understanding tumour growth and development. This is no less true for oral cancer, where slight variations in protocols may invalidate comparison between studies. The influence of tissue processing, and the specific antibody and method of quantification used to identify blood vessels was assessed with respect to oral mucosal vascularity. The angiogenic profile of 20 frozen sections of clinically normal oral mucosa were compared with 16 paraffin-embedded specimens. Eight of the frozen specimens had matched paraffin-embedded tissue. Vascular antigens were identified immunohistochemically using four antibodies and blood vessel density was measured by three methods. CD31 and von Willebrand's factor (vWF) counts were higher than alpha-smooth muscle actin and alphanubeta3. Results for CD31 were higher in frozen tissue than paraffin-embedded tissue. Significantly more CD31-positive vessels were found in frozen tissue in the cohort of matched samples. For paraffin-embedded tissue vWF gave higher values for vascularity than CD31. The variation in antibody reaction to these markers of blood vessels suggests that the antibody employed and method of fixation is more important than the method of quantification used. Comparison of the results for fresh frozen tissue and paraffin-processed tissue suggests that this is acceptable for vWF but not for CD31.