SOLID-PHASE EXTRACTION OF PLASMA VASOPRESSIN - EVALUATION, VALIDATION AND APPLICATION

A new solid-phase extraction method using octyl-silica columns to extract vasopressin-like immuno-reactivity from plasma has been developed. The extraction was followed by a radioimmunoassay on the vacuum-dried extracts, which were reconstituted in assay buffer. The total recovery of synthetic vasopressin was ca. 100%. Based on co-elution with synthetic vasopressin after separation by reversed-phase high-performance liquid chromatography of plasma extracts from normal Wistar and Brattleboro rats, and the cross-reactivity of the antiserum used in the radioimmunoassay system, the extracted material was found to be indistinguishable from authentic vasopressin. Unknown experimental samples were interpolated on a standard curve established in "zero" plasma (plasma derived from rats subjected to waterload) spiked with known amounts of synthetic vasopressin, and not on a standard curve established in assay buffer. The limit of detection was 1 fmol of vasopressin equivalent per millilitre. The intra- and inter-assay coefficients of variance were 10-16% and 16%, respectively. The procedure reliably showed that osmotic challenge and 24-h dehydration increased, whereas ethanol ingestion decreased vasopressin-like immuno-reactivity plasma levels in the rat, compared with normally hydrated controls.