|Values are valid only on day of printing.|
The beta-thalassemia short program utilizes the principles of cation-exchange high-performance liquid chromatography.
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 pre-programmed 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 buffers with different ionic strengths. Changes in absorbance are monitored versus 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 times to 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, BS Coller, TJ Kipps. McGraw-Hill Inc, 1995 pp L37-L38; Fairbanks VF: Thalassemias and related disorders. In Hemoglobinopathies and Thalassemias. Edited by BC Decker, New York, Thieme-Stratton Inc, 1980, pp 18-27; Ou CN, Buffone GJ, Reimer GL: High-performance liquid chromatography of human hemoglobins on a new cation exchanger. J Chromatogr 1983;266:197-205)
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