Abstract
A non-radioactive solid-phase minisequencing method for confirmation of abnormal hemoglobin variants causing sickle cell disease has been developed. In this method amplified 5'-biotinylated target sequences containing normal and mutation sites are immobilized onto streptavidin-coated microplates. Detection primers corresponding to target sequences are annealed immediately adjacent to the mutation site and single-step, haptenlabeled nucleotide primer extension reactions are performed. The incorporation of the labeled nucleotide is detected through immunological reaction with an enzyme-labeled anti-hapten conjugate and a substrate. The method enables confirmation of mutations of the β-globin gene variants (Hbs S, C, E, D-Punjab, O-Arab) and the α-globin gene variant (Hb G-Philadelphia). The test was evaluated using characterized dried blood spot specimens (n = 100). The advantages of the procedure are easy performance and objectiveness. The non-radioactive minisequencing assay will prove helpful for genotyping in neonatal screening for hemoglobinopathies and in prenatal and pre-implantational diagnostics.
Original language | English (US) |
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Pages (from-to) | 71-89 |
Number of pages | 19 |
Journal | Hemoglobin |
Volume | 21 |
Issue number | 1 |
DOIs | |
State | Published - Jan 1 1997 |
Externally published | Yes |
ASJC Scopus subject areas
- Hematology
- Clinical Biochemistry
- Genetics(clinical)
- Biochemistry, medical