Improvement of alpha(0)-thalassemia screening using combined osmotic fragility, dichlorophenolindophenol and Hb H inclusion tests.
BACKGROUND
Screening for alpha(0)-thalassemia is usually done using osmotic fragility (OF) test or reduced erythrocyte indexes, both with high sensitivity, but accurate diagnosis requires PCR analysis. However, a low specificity of screening leads to unnecessary PCR workload during a massive population survey. We have established a more effective screening strategy using a combination of three simple tests.


METHODS
The study was done on 206 subjects with hypochromic microcytosis. Methods include osmotic fragility (OF) test, a dichlorophenolindophenol (DCIP) test for Hb E, a modified Hb H inclusion test, Hb and PCR analyses.


RESULTS
Initial screening with combined OF and DCIP tests identified 9 subjects with negative OF and DCIP tests (-/-), 58 with positive OF test but negative DCIP test (+/-), 13 with negative OF but positive DCIP tests (-/+) and 126 subjects with positive in both tests (+/+). Hb H inclusion was observed in 52 of 206 subjects including 1 in OF/DCIP (-/-), 31 in OF/DCIP (+/-) and 20 in OF/DCIP (+/+) groups. Among these 52 subjects, PCR analysis identified alpha(0)-thalassemia in 28 of 31 (+/-) and 16 of 20 (+/+) groups. Five of 106 subjects with negative Hb H inclusion in the (+/+) group were found to be heterozygous (3 of 5) and homozygous (2 of 5) Hb E co-inherited with alpha(0)-thalassemia.


CONCLUSIONS
Hb H inclusion is not an appropriate screening test for alpha(0)-thalassemia in a region where Hb E is common. However performance of the test in the OF/DCIP (+/-) group would enhance the specificity of screening and result in elimination of almost 50% of cases that would have required further PCR confirmation.
Chaibunruang
Attawut
A
Centre for Research and Development of Medical Diagnostic Laboratories, Thailand.
Pornphannukool
Sontong
S
Sae-Ung
Nattaya
N
Fucharoen
Goonnapa
G
Sanchaisuriya
Kanokwan
K
Fucharoen
Supan
S
D015086:2,6-Dichloroindophenol; D001205:Ascorbic Acid / Q000097:blood*; D001210:Asia, Southeastern / Q000453:epidemiology; D002353:Carrier State; D004906:Erythrocyte Count; D005838:Genotype; D006400:Hematocrit; D006447:Hemoglobin H / Q000235:genetics / Q000378:metabolism*; D006454:Hemoglobins; D006801:Humans; D008403:Mass Screening / Q000379:methods; D009996:Osmotic Fragility; D013785:Thailand / Q000453:epidemiology; D017085:alpha-Thalassemia / Q000097:blood* / Q000175:diagnosis / Q000453:epidemiology
D006454:Hemoglobins; D006447:Hemoglobin H; D015086:2,6-Dichloroindophenol; D001205:Ascorbic Acid