ANALYTICAL EXTRACTION OF REGULATORY PEPTIDES FROM RAT LUNG-TISSUE

We evaluated protocols for the extraction of calcitonin gene-related p eptide, neuropeptide Y, substance P, peptide YY and beta-endorphin fro m rat lung tissue for subsequent radioimmunoassay. The effects of vary ing acidity of the extraction solution and repeating extraction on the recovery of peptide immunoreactivity and non-specific tracer-binding were compared by analysis of variance. Moreover, variability of immuno reactivity was quantified for comparison. Considering all three criter ia, the optimal acidity for extraction was: 0.1 M or 1 M acetic acid f or CGRP and beta-endorphin, 0.1 M acetic acid for NPY, 1 M acetic acid for substance P and phosphate buffer for peptide YY. Double or combin ed extraction unambiguously improved assay results only for substance P. Reversed-phase high-performance liquid chromatography of CGRP-, NPY - and SP-immunoreactivity obtained from selected extracts suggested th at differences in recovery of these peptides are not explainable by di fferential peptide fragmentation during extraction. (C) 1997 Elsevier Science Inc.