There are many different methods for screening for sickle cell disease. Then two most commonly used methods are isoelectric focusing and high performance liquid chromatography.
In isoelectric focusing, proteins are separated based on their specific isoelectric point. This point is determined by the pH in which the protein has no charge and does not migrate farther down the electric field. The specimen is added to a polyacrylamide gel attached to an electrical field and migrates from cathode to anode. The typical analytical run time for one test of ten samples to complete is 2.5 hours. The results are seen as sharp bands in their respective isoelectric points and are automatically calculated to show the percentages of each type of hemoglobin detected. Isoelectric focusing provides clearer banding patterns which are not affected by the analyte degrading in the blood spots. Because this method is not automated, the testing process is labor intensive and time consuming.
High performance liquid chromatography is ten times more expensive than isoelectric focusing. In this method, unknown hemoglobins are identified by comparing their retention times against a standard calibration curve. Test runs were considered normal with the presence of peaks in the zones representing HbF and HbA. Although variants are distinct, the presence of HbS in a specimen may interfere with the accurate HbA2 measurement which identifies β-thalassemia. In a recent study, automated high performance liquid chromatography was calculated to have a coefficient of variation between 0.9-1.8% for the precision of retention times and a coefficient of variation of 4.4-17.5% for the precision of quantification. The retention times were shown to be within 0.01 minutes of the specified identification window. The results of this study showed this method to be an accurate and precise assay for the detection of abnormal hemoglobins.Check out this journal article for more information
Great info, it was interesting to find out about the pros and cons of the two methods. Sickle cell screening must be accurate and precise as much as possible for the sake of those affected by the disease.
ReplyDeleteInteresting. I was wondering what the connection was between sickle cell anemia and B-thalassemia, so I looked it up. We've started discussing thalassemia in Hematology, but I haven't had a chance to look at the notes. Correct me if I'm wrong, but thalassemia is a condition in which there is excessive destruction of red blood cells, whereas sickle cell anemia only results in decreased oxygen carrying capacity. As Dr. Casanova has said, increased RBC destruction can cause tissue hypoxia, so I would assume that the same symptoms would be present with both disorders. Still, I'm a little confused as to why HbS would interfere with the measurement of HbA2.
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