Determination of microcystins in lake water using reusable immunoaffinity column

Abstract

A reusable immunoaffinity column for purification of microcystins in lake water was prepared by coupling anti-microcystin-LR monoclonal antibodies to immunoaffinity support. Thanks to spherical shape of the immunoaffinity support Formyl-Cellulofine® used in this study, applied solutions passed the column smoothly even when used repeatedly. Reusability of the column was examined by determining the recoveries of spiked microcystins-RR, -YR and -LR (100ng each) from lake water. After extraction with a Sep-Pak PS2® cartridge containing styrene–divinylbenzene copolymer, the extract was purified with the immunoaffinity column. The immunoaffinity column was regenerated by washing with Tris–HCl buffer containing bovine serum albumin for repeated uses. Recoveries of spiked microcystins from the first use of the column were 87–88%, and 83–88% from the second and third uses, and the recoveries gradually dropped to 63–77% from the 4–5th uses, the results of which indicated that the column could be used repeatedly for three times. The present method was applied to determine microcystins in water collected from three different lakes in Japan in 1999. In a sample from Lake Suwa, microcystins-RR and -LR were determined by high performance liquid chromatography with photodiode array detection and electrospray ionization-liquid chromatography/mass spectrometry.