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Clinical Chemistry 46: 122-123, 2000;
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(Clinical Chemistry. 2000;46:122-123.)
© 2000 American Association for Clinical Chemistry, Inc.

Technical Briefs

Rapid Determination of Total Homocysteine in Blood Spots by Liquid Chromatography-Electrospray Ionization-Tandem Mass Spectrometry,

Klaus Gempel1,a, Klaus-Dieter Gerbitz1, Bruno Casetta2 and Matthias F. Bauer1

1 Institut für Klinische Chemie, Molekulare Diagnostik und Mitochondriale Genetik, Academic Hospital München-Schwabing, Kölner Platz 1, 80804 Munich, Germany;
2 P.E. Biosystems, Via Tiepolo, 24, 20052 Monza, Italy;
a author for correspondence: fax 49-(0)89-3068-3911, e-mail Klaus.Gempel@lrz.uni-muenchen.de

The determination of total homocysteine (tHcy) plays an important role in diagnosis and therapy of folate and cobalamin (vitamin B12) deficiencies. In addition, it is now widely accepted that increased tHcy is an independent risk factor for thromboembolism and cardiovascular disease, including coronary occlusive disease (1)(2)(3).

Although the causal role of Hcy in the development of vascular occlusive disease has yet to be determined, prospective intervention trials that will effectively lower total plasma Hcy currently are in progress (4). As a result, various analytical methods have been established to measure tHcy in serum or plasma (5)(6)(7)(8)(9)(10). We have established a more rapid protocol suitable for accurately measuring tHcy in hemolysates by use of liquid chromatography-electrospray ionization-tandem mass spectrometry (LC/MS/MS). The LC/MS/MS method circumvents the disadvantages of time-consuming derivatization and allows the processing of >400 samples per day.

Blood was collected in hemolysate tubes that keep the tHcy concentration constant for at least 48 h at room temperature (11). For longer time periods, the hemolysate was stored at -20 °C. Hemolysate (20 µL) was pipetted onto filter paper (Neonatal Screening Card; Schleicher & Schuell). For each sample, two spots (equivalent to 4.7 µL) were punched out and incubated for 15 min at room temperature with a mixture of 20 µL of reducing agent (500 mmol/L dithiothreitol in doubly distilled water) and of 20 µL . . . [Full Text of this Article]


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References




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A. Bowron, A. Barton, J. Scott, and D. Stansbie
Blood Spot Homocysteine: A Feasibility and Stability Study
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T. Arndt, B. Guessregen, A. Hohl, and B. Heicke
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Facts and Recommendations about Total Homocysteine Determinations: An Expert Opinion
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