Extremely high titre polyclonal antisera against small neurotransmitter molecules: rapid production, characterisation and use in light- and electron-microscopic immunocytochemistry.
We have produced polyclonal antibodies against the small amino acid neurotransmitters, GABA, glutamate, glycine and taurine, with a simple new technique using antigens co-adsorbed with an adjuvant peptide to gold particles, which causes rapid and massive immune responses in all animals that we have studied. These antibodies are all of extremely high titre; they are typically used in immunocytochemistry at dilutions from 1 in 250,000 to 1 in 1,000,000 which represents an increase in titre of at least two orders of magnitude compared to standard antibody production techniques. Such very high dilutions result in minimal background labeling and a high signal-to-noise ratio when applied to sections of aldehyde-fixed, epoxy resin-embedded tissues at both light- and electron-microscopic levels. Each antibody displays minimal cross-reactivity with other neurotransmitter molecules. We suggest that our technique may be broadly applicable for raising antibodies against a wide variety of antigens of interest to neuroscientists, particularly those that normally elicit weak immune responses. The technique may also assist in clonal expansion prior to generation of monoclonal antibodies and may be viable, with modifications, for use in human immunisations.
Pow
D V
DV
Department of Physiology and Pharmacology, University of Queensland, Brisbane, Australia.
Crook
D K
DK
D000276:Adjuvants, Immunologic / Q000494:pharmacology; D000596:Amino Acids / Q000276:immunology; D000818:Animals; D000917:Antibody Formation; D000918:Antibody Specificity; D002642:Chick Embryo; D003102:Colloids; D003429:Cross Reactions; D005260:Female; D006046:Gold / Q000276:immunology; D015151:Immunoblotting; D007124:Immunoenzyme Techniques; D007150:Immunohistochemistry; D016253:Microscopy, Immunoelectron; D018377:Neurotransmitter Agents / Q000276:immunology*; D011817:Rabbits / Q000276:immunology; D051381:Rats; D012116:Resins, Plant; D012160:Retina / Q000033:anatomy & histology / Q000648:ultrastructure; D013495:Supraoptic Nucleus / Q000033:anatomy & histology; D013954:Thyroglobulin / Q000276:immunology; D016707:Tissue Fixation
D000276:Adjuvants, Immunologic; D000596:Amino Acids; D003102:Colloids; D018377:Neurotransmitter Agents; D012116:Resins, Plant; D006046:Gold; D013954:Thyroglobulin