SAMPLE PREPARATION, HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC SEPARATION AND DETERMINATION OF SUBSTANCE P-RELATED PEPTIDES

This review deals with the determination of low levels of substance P and peptide fragments derived from the undecapeptide, i.e. covers the whole amount of so-called substance P-like immunoreactivity (SPLI) in biological samples. First an overview of the most currently used sampl e pretreatment procedures is given, followed by a description of the m ost effective high-performance liquid chromatographic (HPLC) separatio n methods. Special attention is paid to the choice of the appropriate column and the possible pitfalls encountered in separation of fmol amo unts of peptide material. Subsequently the most important techniques o f detection are discussed. This section primarily focuses on the coupl ing of HPLC with radioimmunoassay (RIA), which is indispensable for de tection of components in the fmol range at present. Finally, some aspe cts of preparation and chromatographic separation of radiolabelled ant igens for use in RIA are discussed.