Quantifying Phenylhydrazine-Induced Heinz Body Formation in Normal Human Erythrocytes by Flow Cytometry vs. Microscopy or Spectrophotometry

Heinz bodies occur in patients experiencing specific drug-induced acute haemolytic crises, most often in those with an inherited deficiency in glucose-6-phosphate dehydrogenase activity. Laboratory methods for quantifying Heinz bodies in in vitro systems include direct microscopic counts from crystal violet-stained thin blood films or spectrophotometric measurement of free crystal violet in RBC suspensions treated for Heinz body induction. Each method is relatively laborious and prone to errors. In this study, we sought to evaluate and validate an improved flow cytometry-based method for detecting Heinz bodies in human red blood cells after in vitro exposure to phenylhydrazine using washed RBC. We report good concordance between flow cytometric counts of Heinz bodies and microscopic and spectrophotometric analyses of the same suspensions. Flow cytometry offers a less laborious, more precise, and more accurate quantification of Heinz body formation, with strong potential to improve the ease of use in experimental applications.