Using as antigen a 16 amino acid peptide corresponding to the C-terminus of soluble human brain acetylcholinesterase (AChE), four monoclonal antibodies were raised by the novel procedure of in vitro immunization. MAbs BMS-5, -6, -7, and -8 all demonstrated positive binding to the isoforms of cerebrospinal fluid (CSF) and foetal bovine serum (FBS) AChE. In contrast, no binding was seen with "membrane-bound" caudate nucleus and erythrocyte AChEs. With CSF samples from Alzheimer's patients, immunoblotting of isoelectric focussing (IEF) gels with the four mAbs failed to demonstrate any positive binding to the anomalous AChE band. Immunoblotting of SDS-reduced AChE showed binding of the four mAbs with "soluble" caudate nucleus and FBS AChE, but not with erythrocyte AChE. Hence, mAbs BMS-5, BMS-6, BMS-7 and BMS-8 all bind specifically to tetrameric AChE under non-denaturing and denaturing conditions.