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Hemoglobin A2 and F:
Hemolysate of whole blood is injected into an analysis stream passing through a cartridge containing diethylaminoethyl-resin using high-performance liquid chromatography (HPLC). A preprogrammed gradient controls the elution buffer mixture that also passes through the analytical cartridge. The ionic strength of the elution buffer is raised by increasing the percentage of a second buffer. As the ionic strength of the buffer increases the more strongly retained hemoglobins elute from the cartridge. Absorbance changes are detected by a dual-wavelength filter photometer. Changes in absorbances are displayed as a chromatogram of absorbances versus time.(Huismann TH, Scroeder WA, Brodie AN, et al: Microchromotography of hemoglobins. III. A simplified procedure for the determination of hemoglobin A2. J Lab Clin Med 1975;86:700-702; Ou CN, Buffone GJ, Reimer GL, Alpert AJ: High-performance liquid chromatography of human hemoglobins on a new cation exchanger. J Chromatogr 1983;266:197-205)
Hemoglobin Electrophoresis:
The CAPILLARYS System is an automated system that uses capillary electrophoresis to separate charged molecules by their electrophoretic mobility in an alkaline buffer. Separation occurs according to the electrolyte pH and electro-osmotic flow. A sample dilution with hemolysing solution is injected by aspiration. A high voltage protein separation occurs and direct detection of the hemoglobin protein fractions is at 415 nm which is specific to hemoglobins. The resulting electrophoregram peaks are evaluated for pattern abnormalities and are quantified as a percentage of the total hemoglobin present. Examples of position of commonly found hemoglobin fractions are, from cathode to anode: Hb A2', C, A2/O-Arab, E, S, D, G-Philadelphia, F, A, Hope, Bart, J, N-Baltimore and H.(Louahabi A, Philippe M, et al: Evaluation of a new Sebia kit for analysis of hemoglobin fractions and variants on the Capillarys system. Clin Chem Lab Med 2006;44[3]:340-345)
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