Enzyme Linked Immunological Studies of Cholinesterases in Human Blood

Abstract

A modified Ellman method using acetylthiocholine as substrata has been used to reliably measure erythrocyte AChE activity (CV=7%). Reliable immunological quantitation of this enzyme (CV=7%) was achieved using monoclonal antisera to AChE (AE-1 and AE-2) in a fluid phase homogeneous enzyme-linked antiglobulin test (ELAT). AChE activity and antigen levels were similar for both adult male and female samples. Cord samples at term measured 60% of the adult value and reached 100% within 5 months. An overall increase in AChE activity was observed as pregnancy progresses with patients 1 to 3 days post delivery showing significantly elevated levels of activity and antigen. Investigation of haematological disorders revealed cases of reduced levels of activity and antigen in a variety of conditions. An essay of plasma ChE activity using butyrylthiocholine as substrata has been developed using a microplate Ellman technique (CV=5%). An enzyme linked immunosorbent method (ELISA) using monoclonal antibodies MAb2-l and 2-4 (CV=5%) provided a sensitive and reliable means of quantitation together with the capability of handling a large number of samples. Normal Gaussian distribution curves were obtained for both activity and antigen levels for E1u E1u, E1uE1a and E1aE1a individuals. The approximate amount that each of the genes E1a , E1a and E1f contribute to the ChE activity and antigen levels is presented allowing the effective efficiency (EE) of the gene product to be calculated. Immunological measurements have aided in the segregation of the silent gene variants representing zero, very low, low and high levels of immunoreactive protein and the identification of a new gene, E1x. Family studies have confirmed the existence of this gene. Analysis of activity and immunological data suggest that the E1h and E1t gene could be identical. A significant decrease in ChE activity occurs during pregnancy with even lower levels presenting during the first 2 to 3 days post delivery. Immunological measurements indicate similar reductions in the amount of ChE protein during these periods.