Coupling redox and enzymic reactions improves the sensitivity of the ELISA-spot assay

A method is described to increase the sensitivity of the conventional bromo-chloro-indolyl-phosphate ELISA-spot assay. The method is based on the addition of nitro blue tetrazolium to the substrate: the hydrogen ions released during the conversion of indoxyl to indigo, reduce the tetrazolium salt to insoluble diformazan. In this paper, the nitro blue tetrazolium assay is shown (a) to be more sensitive and faster than the bromo-chloro-indolyl-phosphate assay, and (b) to disclose precisely those areas where antibody secretion has occurred.