Enzyme Coupled Immunoassay of Insulin Using a Novel Coupling Reagent

Abstract

A novel coupling reagent, meta-maleimidobenzoyl N-hydroxysuccinimide ester was synthesized. Using this reagent, insulin was conjugated very easily with beta-D-galactosidase [EC 3.2.1.23] in neutral, aqueous solution. No reduction of the enzyme activity was observed during the coupling procedure. The competitive bindings of the conjugate and insulin to anti-insulin serum were tested. The results indicated that the conjugate has enough immune reactivity for use in enzyme coupled immunoassay. Using this assay 20-800 pg of insulin were detectable.