PRODUCTION AND CHARACTERIZATION OF SEPARATE MONOCLONAL-ANTIBODIES TO HUMAN ACETYLCHOLINESTERASE AND BUTYRYLCHOLINESTERASE

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Abstract
Butyrylcholinesterase purified from human plasma and acetylcholinesterase purified from human red blood cells were used to immunize separate groups of BALB/c mice. A solid-phase immunoadsorbance assay was developed to screen and characterize antibodies specific for the cholinesterases. Immunized spleen cells were fused with a non-Ig-secreting [mouse] myeloma cell line (FO). After 2 subcultures at limiting dilution, several clones secreting antibodies to acetylcholinesterase or butyrylcholinesterase were obtained. Selected clones were expanded as ascites tumors in immunosuppressed BALB/c mice. All tested Ig consisted of .kappa. L chains and either G1 or G2b H chains. Two-dimensional gel electrophoresis confirmed the monoclonal nature of each isolated antibody. None of the antibodies to acetylcholinesterase cross-reacted with butyrylcholinesterase and vice versa. All tested antibodies exhibited high avidity for human enzyme, independent of the tissue source (apparent dissociation constants: 1-3 nM for acetylcholinesterase antibodies; 2-13nM for butyrylcholinesterase antibodies). Treatment of enzymes with monoclonal antibodies increased the sedimentation coefficients (from 6.5 S to 12 S for acetylcholinesterase, from 11 S to 18 S or 20 S for butyrylcholinesterase). All of the monoclonal antibodies displayed marked species specificity, Several antibodies reacted only with human enzyme; others reacted with enzyme from nonhuman primates as well. A few of the butyrylcholinesterase antibodies cross-reacted weakly with enzyme from dog, cat and horse, but none reacted with the enzyme from rat, guinea pig and chicken. One acetylcholinesterase antibody cross-reacted with acetylcholinesterase of rabbit and guinea pig. The avidity, species selectivity and other properties of these antibody reagents will be useful in future studies on the regulation and disposition of cholinesterases.

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