Recent advances in the quantitation of human fetal hemoglobins with different gamma chains

Three types of γ chain of human hemoglobin (Hb) F have been discovered. The γ and Aγ chains differ at position 136 (glycine in Gγ; alanine in Aγ) and are the products of nonallelic genes. The Tγ chain is found in rather high frequency in certain world populations, and is characterized by a threonyl residue at position 75 replacing the commonly occurring isoleucyl residue. Convincing evidence is present that the Tγ chain is the product of an allele of the Aγ chain gene, and should be renamed AγT chain. Thus, the three γ chains are GγI, AγI, and AγT. Until recently, the presence and the quantity of these chains were determined by chemical analyses of the tryptic peptides of the isolated Hb F. These procedures involve chromatographic separations or fingerprinting methods. The introduction of high pressure liquid chromatography (HPLC) has greatly facilitated the separation of these peptides. Since minute quantities are required, the latter method is ideally suited for the identification of the Hb F produced in in vitro cell colonies. Electrophoretic and chromatographic (HPLC) methods have been developed which make it possible to separate the Gγ and Aγ chains in small amounts of Hb F (less than 0.5 mg). These procedures allow accurate quantitation of the Gγ/Aγ ratio in the Hb F and may also be suitable for in vitro biosynthetic analyses. One of the HPLC procedures has been modified so that the three types of γ chain can be quantitated in one single chromatographic experiment. Examples of separations are provided, details of some of the methods are discussed, and their applicability illustrated.