Microcolumns prepared in the authors' laboratory, two commercial microchromatography kits, and electrophoresis with elution were compared for Hb A2 quantitation. Day-to-day imprecision of microchromatographic methods was similar (CV 4.7--6.6%) and somewhat less than electrophoresis with elution (CV 8.0--9.1%). Both commercial kits showed variable imprecision in different lots; one lot of Kit B gave erratic results due to resin leakage. From 49 patient specimens, Kit A microcolumns and those of the authors identified the same 14 patients with an elevated percentage of Hb A2 and showed good correlation (P = 0.90), although Kit A showed constant bias toward higher values. Electrophoresis with elution resulted in a false-positive and a false-negative value, did not correlate well with microcolumns (P = 0.78 and 0.76), and showed proportional bias toward lower values for an elevated percentage of Hb A2. Commercial kits were convenient, relatively quick, and cost-effective. Frozen, stabilized hemolysates performed well for quality control.

Blood Protein Electrophoresis↗

PubMed1977

Postsynthetic deamidation of hemoglobin Providence (beta 82 Lys replaced by Asn, Asp) and its effect on oxygen transport.

Carriers of hemoglobin Providence have three types of beta chain in their hemolysates. The two abnormal chains have asparagine (Providence N, Prov N) or aspartic acid (Providence D) at position beta 82, instead of lysine. In vitro, only two beta chains are synthesized by reticulocytes of carriers, betaA and betaProv N. In vivo studies showed that the specific activity of Providence N was initially 10-fold higher than that of Providence D; the specific activities of the two labeled hemoglobins were approximately equal 5 wk after injection of isotope. Oxygen affinity of carriers' blood was somewhat increased, but they were not polycythemic. The affinity of the purified hemoglobins Providence was decreased. Addition of 2, 3 diphosphoglycerate had little effect on the affinity of either hemoglobin component, and addition of inositol hexaphosphate produced no change in the affinity of Providence D. These studies demonstrate that Providence N is deamidated to Providence D during the life span of the erythrocyte, and suggest this finding may represent only an easily observed prototype of posttranslational modification of proteins in general. Despite and abnormal P50 of the blood, oxygen transport is probably normal in carriers of the abnormal hemoglobins.

Adolescent↗