Capillary isoelectric focusing of hemoglobin variants in the pediatric clinical laboratory.
Electrophoresis
; 18(10): 1785-95, 1997 Sep.
Article
em En
| MEDLINE
| ID: mdl-9372271
ABSTRACT
We have used capillary isoelectric focusing (cIEF) to diagnose and monitor hemoglobinopathies in children for over three years. This report describes a major revision of our original method (Clin. Chem. 1994, 40, 2288-2295) that improves the analysis of hemoglobin (Hb) variants by cIEF, especially capillary performance and quantitation precision for minor variants. The revised method uses mixed ampholytes (2% pH 6-83-10; 101), lower viscosity methylcellulose (0.375%) solution, between-sample capillary conditioning with methanol, and hemolysates prepared from red blood cells (RBC) instead of whole blood. Collectively, these changes prolonged capillary life by minimizing capillary exposure to NaOH, standardized the sample matrix, and improved the precision of peak autointegration. The between-run quantitation imprecision (% relative standard deviation) of the revised method was 0.1-3.5% for all diagnostically important major and minor Hb variants present at normal or abnormal levels. The results show the use of the revised method for (i) posttranslationally modified Hb present at low concentrations in normal blood, (ii) Hb oxidation products produced by improper sample storage, (iii) differential diagnosis of S/beta + thalassemia, G-Philadelphia trait, S/C-Harlem disease, and Hb H disease, (iv) sensitive detection of minor variants like Hb A2' as indicators of an alpha globin mutation, and (v) neonatal screening using dried blood collected on filter paper. The results show that high-efficiency separation and precise quantitation of Hb variants over a wide range of concentrations makes cIEF a comprehensive assay that can be used without adjunct analyses for the automated primary evaluation of hemoglobinopathies and thalassemias.
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Temas:
ECOS
/
Aspectos_gerais
/
Financiamentos_gastos
Bases de dados:
MEDLINE
Assunto principal:
Hemoglobinas
/
Eletroforese Capilar
/
Focalização Isoelétrica
Tipo de estudo:
Diagnostic_studies
/
Health_economic_evaluation
Limite:
Child
/
Humans
Idioma:
En
Revista:
Electrophoresis
Ano de publicação:
1997
Tipo de documento:
Article
País de afiliação:
Estados Unidos