Gauging the effects of shellfish extract matrices, salts and pH upon performance of an assay for paralytic shellfish toxins using saxiphilin

In this study, a microtitre plate binding assay using a saxiphilin isoform [3H]STX from the centipede Ethmostigmus rubripes, was challenged with differing conditions of salt concentration and identity, pH and the addition of non-toxic extracts of commercial shellfish.Elements present (Na, K, Mg, Ca, Zn, Ni, Mn, Cr, Sr, Mo, Al, V, Fe, Cu, Pb, Co) in the shellfish extracts (green lipped mussel, Perna canalicularis; Sydney rock oyster, Saccostrea glomerata; and pipis, Donax deltoides) were measured by inductively coupled plasma spectroscopy. The effects upon the assay by the four most common salts of these elements, NaCl, KCl, CaCl2 and MgCl2 were studied by varying their concentrations (0-800 mM).The effect of pH upon the assay was measured by replacing the assay buffer in standard conditions with a buffer mixture of Tris (20 mM) -Mes (10 mM) -acetic acid (10 mM), adjusted with tetramethylammonium hydroxide or HCl to indicated pH which ranged from 3.4 to 9.0. To investigate the microtitre plate saxiphilin assay's ability to deal with potential interfering compounds from shellfish extracts, and the effect of acidic pH necessary for toxin stability when extracting PSTs from shellfish.