A Highly Specific Antiserum for Arginine Vasopressin1

Abstract

A high sensitivity (titer 60-150,000) radioimmunoassay for arginine vasopressin (AVP) was developed using synthetic AVP as antigen and standard. The specificity of the antiserum was characterized by measuring the relative binding affinities of 29 analogs and fragments of AVP. In general, single amino acid substitution or deletion in the AVP molecule caused a reduction in binding to the antiserum; binding was further impaired for analogs which had multiple substitutions. Ring and tail fragments of the neurohypophysial hormones did not bind to the antiserum, and the only analog which the antiserum could not differentiate from AVP was deamino-AVP. More specifically, the antiserum exhibited the following properties: position 1, no requirement for a position charge at 1/2 Cys; positions 2 and 3, requirements for .pi.-.pi. interacting pair of aromatic side chains and for Tyr at position 2; position 4, requirement for Gln (> Thr .mchgt. Val); position 8, requirement for a basic side chain of optimal length, preferably terminating with a guanidino group (Arg > Har > Hly > Orn > Lys > Leu); position 9, requirement for Gly-NH2. The antiserum exhibits a very high degree of specificity toward all the structural and conformational features of the AVP molecule, indicating its reliability for determining AVP levels in body fluids and tissues.