Clinical Chemistry
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Clinical Chemistry 53: 2205-2209, 2007. First published October 19, 2007; 10.1373/clinchem.2007.095133
This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow 095133.Supplemental Data
Right arrow All Versions of this Article:
clinchem.2007.095133v1
53/12/2205    most recent
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Web of Science (3)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Tsang, J. C.H.
Right arrow Articles by Chiu, R. W.K.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Tsang, J. C.H.
Right arrow Articles by Chiu, R. W.K.
(Clinical Chemistry. 2007;53:2205-2209.)
© 2007 American Association for Clinical Chemistry, Inc.

Technical Briefs

Mass Spectrometry–Based Detection of Hemoglobin E Mutation by Allele-Specific Base Extension Reaction

Jason C.H. Tsang1, Pimlak Charoenkwan2, Katherine C.K. Chow1, Yongjie Jin3, Chanane Wanapirak2, Torpong Sanguansermsri2, Y.M. Dennis Lo1,4 and Rossa W.K. Chiu1,4,a

1 Department of Chemical Pathology, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong SAR; 2 Faculty of Medicine, Chiang Mai University, Chiang Mai, Thailand; 3 Centre for Emerging Infectious Diseases and 4 Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong SAR;

aaddress correspondence to this author at: Department of Chemical Pathology, Rm. 38063, 1/F, Clinical Sciences Building, Prince of Wales Hospital, 30-32 Ngan Shing St., Shatin, Hong Kong SAR; fax 852 2636 5090, e-mail rossachiu{at}cuhk.edu.hk)


Abstract

Background: The specific detection of a minor population of mutant DNA molecules requires methods of high specificity and sensitivity. While the single-allele base extension reaction (SABER) was shown to be useful for the detection of certain beta-thalassemia mutations, we encountered problems with false positivity during development of SABER for the noninvasive prenatal diagnosis of the hemoglobin E (HbE) disease. Systematic optimization resulted in an alternative protocol, the allele-specific base extension reaction (ASBER).

Methods: An artificial model was established by mixing genomic DNA of HbE carriers and normal individuals. Effects of terminator concentration and annealing temperature on the nonspecificity of SABER were then studied. The use of a single relevant terminator and the other 3 types of dideoxynucleotide as competing terminators were also compared in the development of the ASBER protocol. Thirteen cases of HbE-susceptible pregnancies were tested to compare the SABER and the ASBER protocols.

Results: Decreasing the single relevant terminator concentration and increasing the annealing temperature in SABER were found to improve specificity. The use of the other 3 types of dideoxynucleotide as competing terminators was shown to offer better detection sensitivity than a single terminator in ASBER. Genotyping results were all correctly determined by ASBER, except one false-negative detection (sensitivity: 80%, specificity: 100%).

Conclusions: An alternative mass spectrometry–based protocol for noninvasive prenatal diagnosis, ASBER, has been successfully developed to allow the detection of a minor DNA population with a point mutation.




The following articles in journals at HighWire Press have cited this article:


Home page
 J. Clin. Pathol.Home page
E C W Hung, R W K Chiu, and Y M D Lo
Detection of circulating fetal nucleic acids: a review of methods and applications
J. Clin. Pathol., April 1, 2009; 62(4): 308 - 313.
[Abstract] [Full Text] [PDF]

Home page
 Proc. Natl. Acad. Sci. USAHome page
F. M. F. Lun, N. B. Y. Tsui, K. C. A. Chan, T. Y. Leung, T. K. Lau, P. Charoenkwan, K. C. K. Chow, W. Y. W. Lo, C. Wanapirak, T. Sanguansermsri, et al.
Noninvasive prenatal diagnosis of monogenic diseases by digital size selection and relative mutation dosage on DNA in maternal plasma
PNAS, December 16, 2008; 105(50): 19920 - 19925.
[Abstract] [Full Text] [PDF]

Home page
 Clin. Chem.Home page
F. M. F. Lun, R. W. K. Chiu, K. C. Allen Chan, T. Yeung Leung, T. Kin Lau, and Y. M. Dennis Lo
Microfluidics Digital PCR Reveals a Higher than Expected Fraction of Fetal DNA in Maternal Plasma
Clin. Chem., October 1, 2008; 54(10): 1664 - 1672.
[Abstract] [Full Text] [PDF]


HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Copyright © 2007 by the American Association for Clinical Chemistry.