Test ID: SFMON    
Hemoglobin S and Hemoglobin F Quantitation for Therapeutic Monitoring, Blood

This test is performed via cation-exchange HPLC. Hemolyzed specimens are maintained at 12 degrees C +/- 2 degrees C in the automatic specimen chamber. The specimens are then sequentially injected into the analysis stream at 6.5 minute intervals for a throughput of 9 specimens per hour. Two dual-piston pumps and a preprogrammed gradient control the elution buffer mixture passing through the analytical cartridge. The ionic strength of the elution buffer mixture is increased by raising the percentage contribution of elution buffer 2. As the ionic strength of the mixture increases, more strongly retained hemoglobins will elute from the analytical cartridge.

A dual-wavelength filter photometer (415 and 690 nm) monitors the elution from the cartridge. As the hemoglobins elute from the cartridge and pass through the photometer flow cell, changes in absorbance at 415 nm are detected. The 690 nm secondary filter corrects the baseline for changes caused by the mixing of different ionic strength buffers. Changes in absorbance are monitored over time, producing a chromatogram. A built-in integrator performs reduction of the raw detector signal data collected from each analysis.

The elapsed time from the injection of the specimen to the apex of a hemoglobin peak is called the retention time. Each hemoglobin has a characteristic retention time. Windows are established from the most frequently occurring hemoglobins based on their characteristic retention time aid in the interpretation of results.(Kim, HC, Adachi K, Scwartz E: Separation of hemoglobins. In Williams Hematology. Fifth edition. Edited by E Beutler, MA Lichtman, S Coller, TJ Kipps. McGraw-Hill Inc, 1995, pp L37-L38; Ou CN, Buffone GJ, Reimer GL: High-performance liquid chromatography of human hemoglobins on a new cation exchange. J Chromatogr 1983;266:197-205)

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