Fast and sensitive diagnosis of thalassemia by capillary electrophoresis.
This paper demonstrates the diagnosis of beta-thalassemia by capillary electrophoresis in conjunction with laser-induced fluorescence using poly(ethylene oxide) (PEO) solutions in the presence of electroosmotic flow (EOF). During the electrophoretic separation, PEO solution entered a capillary from the anodic vial by EOF. The separation of a mixture of the polymerase chain reaction (PCR) products (330 and 334 base pairs) from a healthy person and a beta-thalassemia patient was accomplished within 15 min at 15 kV using 1.5% PEO containing 2 M urea at 30 degrees C. The electropherogram patterns instead of migration times were used to diagnose beta-thalassemia, with an accuracy of 100% for the analyses of 11 blood samples from suspected patients. After injecting a large volume of the mixture to the capillary filled with 800 mM Tris-borate buffer (pH 10.0), the DNA fragments stacked due to increases in viscosity and sieving when migrating into 1.5% PEO solution. As a result of improved sensitivity, only 15 PCR cycles were required when using 500 ng of DNA templates. The results shown in this study indicate the potential of this simple, rapid, and cost-effective method for the diagnosis of beta-thalassemia.
Chang
Po-Ling
PL
Department of Chemistry, National Taiwan University, Section 4, Roosevelt Road, Taipei, Taiwan, R.O.C.
Kuo
I-Ting
IT
Chiu
Tai-Chia
TC
Chang
Huan-Tsung
HT
D002021:Buffers; D004247:DNA / Q000032:analysis*; D019075:Electrophoresis, Capillary; D006801:Humans; D011092:Polyethylene Glycols / Q000737:chemistry; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D013696:Temperature; D013789:Thalassemia / Q000175:diagnosis*; D017086:beta-Thalassemia / Q000175:diagnosis
D002021:Buffers; D011092:Polyethylene Glycols; D004247:DNA