Very High leucocytosis Interfere with colorimetric measurement of Hemoglobin

( Originally the article was published Northern Medical Journal, 2006; Vol-15 No-2, It has been modified for blog)-Helpful for laboratory practitioner
Hemoglobin estimation of a solution or a blood sample is commonly done in most cases by cyanmethhemoglobin method. But a less familiar method, which is also a photoelectric colorimetric methed,1developed by us, is followed in some areas of Bangladesh. The later method is a acid-hematin method and has found to have similar accuracy and stability. Unlike cyanmethhemoglobin method, it is very cheap and has a advantage that no biohazardous chemical have been used here2. Interestingly, during the development of the new method with a comparison to cyanmethhemoglobin method, it is observed that in both methods prepared fluid mixed with blood of high WBC count showed optical density resulting hemoglobin level that didn’t correspond with clinical condition of the patient. That means higher hemoglobin level was observed in a clinically anemic patient whose WBC count is very high e.g. in case leukemia. Moreover the blood mixed fluid, which is clear in other cases, was found to be hazy in these cases. To get rid from the problem we centrifuge the fluid and the clear supernatant fluid was used to measure the optical density and then the actual level of hemoglobin could be estimated which corresponded well with clinical condition of the patient. The deposit, after centrifugation, was examined under microscope and found plenty intact WBC. In normal cases, the deposits also shown WBC but were in very small number. Roughly it was estimated that hemoglobin level was increased by 1.2 to 1.4 gram% for 1,00,000WBC/cumm of blood. So, during estimation of hemoglobin in patient with very high WBC count (e.g leukemia) optical density should be taken from the supernatant fluid after centrifugation for few minutes (more than 3 minutes is satisfactory) of the prepared fluid.
Cyanmethaemoglobin method is followed in many countries for the last few decades, though potassium cyanide, which is used in this method, carries potential health hazard. Besides this, the limitation regarding high WBC count should be always considered. It can be easily proved in any laboratory by estimating hemoglobin of a patient having very high WBC count e.g. leukemia, by measuring absorbance before and after centrifuging the measuring fluid.

References:

1. Mujibur Rahman. A New Method for Measuring Hemoglobin. Laboratory Hematology;
2003, Vol 9, No 3: 179.
2. Barbara J Brain & Imelda Bates. Basic haematologic techniques. Dacie and Lewis Practical Haematology. Ninth Edition, Churchill Livingstone. 2001.19-46.