Histamine monoclonal antibody for brain immunocytochemistry

Among five monoclonal antibodies (AHA-1 to 5 mAbs) prepared against glutara ldehyde (GA)-conjugated histamine (HA) in our previous study, only mAb AHA- 2 was found to detect HA specifically in rat brain neurons by an immunocyto chemistry method (ICC) using GA as a tissue fixative. All the other mAbs, e xcept for AHA-B, reacted with HA in the enterochromaffin-like cells (ECL ce lls) of rat stomach [Fujiwara et al, (1997) Histochem, Cell Biol, 107, 39-4 5], Enzyme-linked immunosorbent assay (ELISA) binding and inhibition tests demonstrated that AHA-S is specific for HA, with almost no detectable cross reaction with any other established or putative amino acid neurotransmitter s, LH-RH, TRH, or peptides with N-terminal histidines. ELISA assays also su ggested that the AHA-8 mAb recognizes a HA epitope structure different from the one recognized by the AHA-1 mAb, The immunostaining patterns with AHA- 2 mAb, as seen in the five subgroups of the tuberomammillary nuclei in the rat posterior hypothalamus, were very similar to those described by Inagaki et al, [(1988) Brain Res, 439, 402-405; (1990) Exp, Brain Res, 80, 374-380 ] and Panula et al, [(1984) Proc. Natl, Acad, Sci, USA 81, 2572-2576; (1988 ) J, Histochem, Cytochem, 36, 259-269] using polyclonal anti-HA serum. Howe ver, it was also noted that moderate numbers of immunoreactive nerve fibers projected into the median eminence. The present HA ICC method using AHA-2 mAb allows highly sensitive HA detection in brain, and thus might permit de tailed studies of HA localization hitherto impossible using previously avai lable anti-HA polyclonal antibodies produced against carbodiimide-conjugate d HA.