J Clin Pathol 1981;34:796-799 Zinc acetate as a precipitant of unstable haemoglobins

SUMMARY Whereas the addition of four moles of zinc acetate to one of normal human haemo-globin at pH 7-4 results in a clear solution, addition to unstable haemoglobin results in precipitate formation. Within certain limits no false-positive results are obtained with methaemoglobin, and with fetal haemoglobin. With excess zinc, normal haemoglobin also precipitates, but this process is reversible and can be used for a rapid concentration of dilute solutions of normal haemoglobin. We have recently been searching for a means to precipitate selectively very small amounts of an un-stable haemoglobin from a haemolysate as found, for example, in heterozygotes for Hb A and Hb Coventry. ' In the course of this study which is not yet finished it became apparent that zinc acetate may be a useful laboratory tool in a routine test for discovering the common unstable haemoglobins. Within certain limits it does not give false-positive results with methaemoglobin and with fetal haemo-globin. Furthermore it can be used for the reversible precipitation of normal haemoglobin from dilute solutions. Material and methods A sample (0-2 ml) of 10 g/dl haemolysate is added to 1P8 ml of a solution containing 0-35 ml of 0 003 M zinc acetate and 1-45 ml of TRIS (trimethamine) buffer 0&1 M, pH 7-4. After mixing, the tubes are incubated for 30 min at 37°C. With unstable haemo-globins an immediate flocculation is seen which has settled as a precipitate by the end of the incubation period. For routine tests this proportion of 3 5 mol zinc:1 mol Hb is preferred because slight precipi-tates (±) may appear with 0 4 ml zinc acetate and 1-4 ml of buffer, with some normal haemolysates. They can of course be disregarded, but there is no disadvantage in arranging the routine test as de-scribedhere usingO035 ml of zinc acetate and 1-45 ml of buffer (3-5 mol zinc:l mol Hb)