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Special Hemoglobin Tests

 

Isopropanol Test (IT) for Unstable Hemoglobins

The isopropanol test is performed for detection of unstable hemoglobins or hemoglobins with altered oxygen affinity. Some hemoglobin mutations result in an unstable hemoglobin that has decreased solubility and precipitates to form erythrocyte inclusions (Heinz bodies).  These inclusions reduce the deformability of the red blood cell and lead to hemolysis. Examples of unstable hemoglobins are Hb Gun Hill, Hb Leiden, and Hb Köln.

Hemolysate is incubated in a Tris buffer containing isopropanol. The less polar isopropanol buffer stresses the sulfhydryl bonds of the Hb molecule.  Normal hemoglobins will not precipitate within 20min, while unstable hemoglobins will precipitate within 5 minutes.

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(Ref: Carrel RW, Hemoglobin stability tests, Crit Rev Clin Lab Sci, 5(1):62-69,1974)