Clinical Chemistry
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Clinical Chemistry 45: 1974-1980, 1999;
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Right arrow Lipids, Lipoproteins, and Cardiovascular Risk Factors
(Clinical Chemistry. 1999;45:1974-1980.)
© 1999 American Association for Clinical Chemistry, Inc.

Articles

Dynamic Reaction in a Homogeneous HDL-Cholesterol Assay Visualized by Electron Microscopy

Akira Kondo1,a, Yoshinori Muranaka2, Isao Ohta2 and Takashi Kanno1

1 Department of Laboratory Medicine and
2 Central Laboratory for Ultrastructure Research, Hamamatsu University School of Medicine, 3600 Handa-cho, Hamamatsu City, 431-3192 Japan.
a Author for correspondence. Fax 81-53-435-2794; e-mail akikondo{at}hama-med.ac.jp

Background: Measurement of HDL-cholesterol (HDL-C) by homogeneous assays with automated analyzers is replacing precipitation methods. However, in this reaction-type assay, interactions between the reagents and lipoproteins remain unknown.

Methods: Electron microscopy was used to investigate the reactions in a homogeneous HDL-C assay. Negative staining with 10 g/L uranyl acetate was performed for lipoprotein visualization by electron microscopy. Observations of the interactions between lipoproteins and the reagents of a polyanion-polymer/detergent assay were achieved by cooling the reaction mixture in ice water. This treatment also allowed observation of the time course of the reaction.

Results: In the first-reagent reaction (polyanion-polymer), every lipoprotein aggregated almost completely. In the second-reagent reaction (enzymes and detergent), only HDL in the lipoprotein aggregates was selectively resolved and reacted enzymatically. Reagent 1 contains two important substances: polyanion and synthetic polymer. Using x-ray microanalysis, we confirmed that aggregation of lipoproteins in the first reaction occurred through interaction with the phosphotungstate of the polyanion.

Conclusion: Electron microscopy morphologically revealed the dynamic reaction in a homogeneous HDL-C assay.




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Home page
 Clin. Chem.Home page
G. R. Warnick, M. Nauck, and N. Rifai
Evolution of Methods for Measurement of HDL-Cholesterol: From Ultracentrifugation to Homogeneous Assays
Clin. Chem., September 1, 2001; 47(9): 1579 - 1596.
[Abstract] [Full Text] [PDF]


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