Accurate mass measurement by electrospray ionization quadrupole mass spectrometry: detection of variants differing by <6 Da from normal in human hemoglobin heterozygotes

Rai, D.K., Griffiths, W.J., Landin, B., Wild, B.J., Alvelius, G. and Green, B.N. (2003) Accurate mass measurement by electrospray ionization quadrupole mass spectrometry: detection of variants differing by <6 Da from normal in human hemoglobin heterozygotes. Analytical Chemistry, 75 (9). pp. 1978-1982. 10.1021/ac026228h.

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DOI: 10.1021/ac026228h

Abstract

Detection of Variants Differing by <6 Da from Normal in Human Hemoglobin Heterozygotes Dilip K. Rai,* William J. Griffiths, Britta Landin, Barbara J. Wild, Gunvor Alvelius, and Brian N. Green Division of Clinical Chemistry, Department of Medical Laboratory Sciences and Technology, Huddinge University Hospital C1 74, Karolinska Institutet, SE-141 86 Stockholm, Sweden, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, SE-171 77 Stockholm, Sweden, Department of Haematological Medicine, King's College Hospital, Denmark Hill, London SE5 9RS, U.K., and Micromass UK Limited, 3 Tudor Road, Altrincham, Cheshire, WA14 5RZ, U.K. Received for review October 11, 2002. Accepted March 6, 2003. Abstract: Mass spectrometry has a basic limitation when human hemoglobin variants are analyzed, because it cannot resolve two globin chains that differ in mass by <6 Da. Several common -chain variants differ by 1 Da from normal and, hence, when present in heterozygotes, are not resolved from the normal -chain. Normal and variant chains appear together in the spectrum as a single entity, whose mass is the abundance weighted mean of the two chains. Here we show that such heterozygotes can be detected in 500-fold diluted blood by accurately measuring the mass of the -chain using an electrospray ionization quadrupole instrument and the -chain for internal mass calibration. A statistical analysis of the normal -chain mass (n = 86) showed that the standard deviation (SD) of the mean was <±0.05 Da (<±3.2 ppm). Hence, at the 95% confidence level (±2 SD), an abnormal - or -chain differing by 1 Da from normal should be detectable in a heterozygote provided its abundance is >10% of total - or -chains, respectively. Variants whose masses lay between 1 and 4 Da from normal were detected in 19 heterozygotes. Moreover, the proportion of each variant estimated from the mass change correlated with the proportion determined by cation-exchange HPLC. Variants were assigned to the - or -chain by combining the sign of the mass change with the polarity change inferred from electrophoretic data. This procedure could be used for screening clinically significant hemoglobin variants.

Item Type:Article
Additional Information:Full text available in print and electronically at the School of Pharmacy Library.
Departments, units and centres:Department of Pharmaceutical and Biological Chemistry > Department of Pharmaceutical and Biological Chemistry
ID Code:624
Journal or Publication Title:Analytical Chemistry
Deposited By:Library Staff
Deposited On:17 Jan 2007
Last Modified:19 Sep 2007 11:15

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