Sensitive assay systems for detection of hemoglobin with 2,7-diaminofluorene: histochemistry and colorimetry for erythrodifferentiation

Sensitive and rapid assays, colorimetry and histochemistry, for hemoglobin in erythroid cells are established. The assays are based on pseudoperoxidase activity of hemoglobin using 2,7-diaminofluorene as a hydrogen donor for the peroxidase, instead of benzidine which is widely benzidine which is widely used for the detection of small amounts of hemoglobin but which is a potent carcinogen and has been banned from laboratory use. In the presence of hydrogen peroxide, hemoglobin catalyzes the formation of a blue compound (fluorene blue), which has a broad absorption band between 500 and 690 nm with a peak at 610 nm, from 2,7-diaminofluorene. The reagent is safe to use in the laboratory. The methods could be applied to the detection of hemoglobin in Friend erythroleukemia cells induced to cell differentiation along the erythroid pathway by dimethyl sulfoxide.