A simple enzymatic microdetermination of cytochrome b5 in erythrocytes

Abstract

A simple enzymic method for the microscale determination of erythrocyte cytochrome b 5 is presented. For assay of cytochrome b 5 the sodium nitrite-treated hemolysate was incubated in the presence of NADH and NADH-cytochrome b 5 reductase, and the reduction of methemoglobin was measured. The rate of methemoglobin reduction was linearly dependent on the amount of cytochrome b 5. When the red cells were stored at 4°C or methemoglobin-formed hemolysates were stored at 4°C or −20°C, there was no significant decline in the amount of cytochrome b 5 for periods of at least 2 weeks. Values for normal human subjects showed a mean of 0.70 ± 0.17 SE nmol/ml of red cells for males, 0.52 ± 0.11 nmol/ml red cells for females, and 0.48 ± 0.15 nmol/ml red cells for umbilical cord bloods.