Cyanobacteria produce a wide range of potent toxins, including hepatotoxic microcystins. HPLC methods for microcystin analysis and purification almost invariably include acetonitrile in the elution gradient mobile phase. The recent, acute, global acetonitrile shortage requires that adequate methods are available for microcystin analysis and purification without the need for acetonitrile. Here we present a convenient methanol-based method for effective HPLC analysis and purification of the toxins, with full separation of a range of microcystin variants.