An enzyme immunoassay for determining plasma concentrations of didemnin B.

Didemnin A was conjugated at the amino terminus of the N-methylleucine residue, via the linkers N-succinimidyl-3-(2-pyridyldithio)-propionate and trans-1,4-maleimidomethyl-cyclohexane carboxylic acid, to keyhole limpet hemocyanin (KLH) and bovine serum albumin (BSA). The didemnin-KLH conjugates were used to hyperimmunize rabbits. The resulting high titer antisera were employed with didemnin-BSA conjugate-coated microtiter plate wells to develop an indirect competitive inhibition enzyme immunoassay (CIEIA) that was fully cross reactive with didemnin B. A CIEIA is described that is capable of detecting the drug in plasma from didemnin B-treated patients at concentrations down to 1-3 ng/ml. This simple, sensitive CIEIA has been employed to demonstrate plasma drug clearance profiles with samples from didemnin B-treated patients.