Detection of Prorocentrum minimum (Pavillard) Schiller with an Electrochemiluminescence-Molecular Probe Assay

The occurrence of harmful algal blooms (HABs) caused by Prorocentrum minimum (Pavillard) Schiller is a crucial subject in the study of HABs. An electrochemiluminescence-molecular probe assay (ECL-MP) was developed to qualitatively and quantitatively detect P. minimum. It was based on the sandwich hybridization integrated with a nuclease protection assay (NPA-SH) and improved by electrochemiluminescence (ECL). An ECL analyzer was established in this study, and it was shown that this analyzer was stable and highly sensitive, with a detection range of 0.4 pmol to 4 nmol Ru(bpy)(3)Cl-2 center dot 6H(2)O under optimal reaction conditions of 1.0 V, 1.0 mA, 1.5 mol center dot L-1 TPrA, and pH 7.4. The optimal amount of magnetic beads for separation of labeled NPA probes in a 20-mu L hybridization mixture was 4 mu g. The ECL counts per second was linear with the number of P. minimum cells in a range of 6.25 x 10(2) to 4 x 10(4), and there was no significant difference between ECL-MP and microscopy, with a 95% confidence level (t test) when individual, mixed cultures and field samples were treated. This study provides a convenient method for fast and accurate detection of P. minimum in the marine environment.