Abstract
We investigated the possibility of immunohistochemically identifying T- and B-lymphocytes in formalin-fixed, paraffin-embedded rat tissues as a basis for future pathological study. Spleens were removed from 8-week-old male Wister rats, and were fixed in 20% neutral-buffered formalin for 7, 30 or 90 days. After fixation, the specimens were embedded in ordinary paraffin by routine method. Paraffin sections were prepared and stained immunohistochemically using anti-CD3 (clone: G4.18) and -CD45RA (clone: OX-33) antibodies. Anti-CD3 and -CD45RA antibodies failed to stain without microwave heating pretreatment. The immunoreactivity of CD3 and CD45RA were both increased by microwave heating pretreatment. Consistent and specific immunostaining of CD3 was obtained throughout all formalin-fixation periods after 40 min or more of microwave heating pretreatment. CD45RA was clearly detected in the 7-day fixed tissues after 30 or 40 min of pretreatment. In the 30- and 90-day fixed tissues, identification of CD45RA was difficult at all lengths of microwave heating pretreatment. These findings suggest that CD3 and CD45RA are detectable in routine formalin-fixed, paraffin-embedded rat tissues although the immunoreactivity is influenced by formalin-fixation period, and that the present procedure, combining appropriate antibodies and pretreatment of microwave heating, is useful for retrospective analyses of rat lymphoid subtypes.
