Novel Test Method (Sickle Confirm) to Differentiate Sickle Cell Anemia from Sickle Cell Trait for Potential Use in Developing Countries

Abstract

The objective of this study was to develop a diagnostic testing method to detect HbS, distinguish sickle cell homozygotes from heterozygotes, and overcome testing barriers encountered in laboratories in underdeveloped countries. Blood samples positive and negative for sickle cell were subjected to the standard hemoglobin solubility test followed by a variety of centrifugation and filtration procedures. Each procedure was evaluated for the ability to remove insoluble HbS from the sample. The hemoglobin types that remain (HbA, HbA₂ and HbF) were measured spectrophotometrically or estimated visually allowing samples to be categorized into three genotypes (AA, AS and SS) as confirmed by hemoglobin electrophoresis. De-identified EDTA blood samples were obtained from Saint Louis University and Cardinal Glennon Children's hospitals and tested in the Department of Clinical Laboratory Science at Saint Louis University. The main outcome measures were turbidity of the solubility solution; color of the supernatant and the material on the surface of the solution following centrifugation; precipitate trapped on the filter paper; absorbance of the filtrate; and hemoglobin electrophoresis patterns. Centrifugation and filtration successfully separated HbS from HbA/A₂/F allowing for the differentiation of seven sickle cell homozygotes from sixteen heterozygotes with a sensitivity and specificity of 100%. This method has the potential to reliably distinguish homozygous from heterozygous sickle cell patients and it is fast, inexpensive, and simple. These characteristics make Sickle Confirm a desirable method in developing countries like Haiti and Africa where sickle cell anemia is prevalent and modern diagnostic methods like electrophoresis, HPLC and nucleic acid testing are impractical.