Construction of a Single Chain Variable Fragment Antibody (scFv) against Carbaryl and Its Interaction with Carbaryl.

Carbaryl is a low molecular weight insecticide that inhibits cholinesterase. Residues of carbaryl in food and the environment have damaged human health. A high-specificity scFv that can identify carbaryl is still lacking. In the present study, an anti-carbaryl scFv gene was prepared by cloning VL and VH genes from hybridoma cells secreting monoclonal antibody, then VH and VL were fused together using splicing by overlap extension (SOE) PCR with a flexible polypeptide linker connector (Gly4Ser)3, and then the scFv-pET-26b recombinant plasmid was constructed and transformed into E. coli BL21 for expression using IPTG as an inducer. The expressed recombinant protein was identified by SDS-PAGE and ELISA. The three-dimensional structure of the anti-carbaryl scFv was constructed by computer modeling, and carbaryl was docked to the scFv model to obtain the structure of the binding complex. The binding site was composed of Ala51, Ser52, Ile51, Gly54, Ser56, Arg98, and Gly100. This helps to understand the mechanism of interaction between anti-carbaryl antibody and antigen. Furthermore, it provides guidance for in vitro affinity maturation of anti-carbaryl antibody.

Night-time rise in rat pineal N-acetyltransferase due to carbaryl administration is reduced by propranolol treatment.

The purpose of the present study was to examine the effects of administration of sublethal doses of carbaryl on nighttime rat pineal melatonin synthesis in the presence and absence of propranolol, a beta-adrenergic receptor antagonist. Two groups of adult male albino rats were administered orally N-methyl-l-naphthylcarbamate (carbaryl) (8.33 mg/kg BW daily in corn oil) for six successive days; another two groups received corn oil only. On the last day of carbaryl treatment, half of the animals received an intraperitoneal injection of propranolol (20 mg/kg body weight, one hour before lights off). The other two groups were given a saline injection. Four hours after darkness onset, pineal N-acetyltransferase (NAT) and hydroxyindole-O-methyltransferase (HIOMT) activities as well as pineal concentrations of 5-hydroxytryptophan (5HTP), serotonin (5HT), 5-hydroxyindole acetic acid (5HIAA) and pineal and serum melatonin levels were measured. Nocturnal NAT activity was increased due to carbaryl administration but the pesticide was ineffective in stimulating NAT activity in rats treated with propranolol. Pineal 5HT was decreased due to carbaryl administration but 5HTP and 5HIAA levels were unaffected. Pineal and serum melatonin levels were decreased due to propranolol treatment. The results indicate that carbaryl may influence pineal NAT activity by a mechanism that involves beta-adrenergic neural transmission.

Central cholinergic-dopaminergic interaction in carbaryl-induced tremor.

Acute administration of the anticholinesterase agent carbaryl to rats produced a dose-dependent increase of tremors. The carbaryl-induced tremor was significantly reduced by prior treatment of the rats with either atropine or L-DOPA and was exacerbated by haloperidol. These results suggest the possible involvement of a central cholinergic-dopaminergic interaction in the carbaryl-induced tremor.