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1
Department of Biochemistry, Hôpital Saint Antoine, 75012 Paris, France.
2
SEBIA, 23 Rue Maximillien Robespierre, 92130 Issy les
Moulineaux, France.
3
Department of Biochemistry, Hôpital de l’Archet,
06000 Nice, France.
4
Department of Endocrine and Metabolic Diseases, Hôpital Saint Antoine, 75012 Paris, France.
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Department of Biostatistics, Saint Antoine School of
Medicine, 75012 Paris, France.
6
Department of Biochemistry, Hôpital Trousseau,
75012 Paris, France.
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Department of Hepatology, Hôpital
Saint Antoine, 75012 Paris, France.
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Department of Cardiology, Hôpital
Saint Antoine, 75012 Paris, France.
a Address correspondence to this author at: Hôpital Saint Antoine, 184 Rue du Faubourg Saint Antoine, 75012 Paris, France. Fax 33-1-49-282206; e-mail pascale.benlian{at}sat.ap-hop-paris.fr
Background: Automated electrophoresis combined with enzymatic cholesterol staining might improve routine assessment of LDL- and HDL-cholesterol (LDLC and HDLC), as an alternative to the Friedewald equation and precipitation. A new method (Hydrasys; SEBIA) that adapts the cholesterol esterase/cholesterol oxidase reaction within urea-free gels was evaluated.
Methods: Fresh sera from 725 subjects (512 dyslipidemics) were analyzed by electrophoresis, in parallel with sequential ultracentrifugation, ß-quantification, calculation, and precipitation.
Results: Electrophoresis was linear up to 4 g/L cholesterol, with a detection limit of 0.042 g/L cholesterol/band. Within-run, between-run, between-batch, and between-operator imprecision (CVs) were 1.6%, 2.0%, 1.5%, and 2.7% for LDLC, and 3.9%, 4.3%, 5.5%, and 4.9% for HDLC, and remained unchanged up to 6.3 g/L plasma triglycerides (TGs). Precision decreased with very low HDLC (<0.25 g/L). Serum storage for 3–7 days at +4 or -80 °C did not interfere significantly with the assay. Agreement with ß-quantification was stable for LDLC up to 5.07 g/L (r = 0.94), even at TG concentrations >4 g/L (r = 0.91). Bias (2.88% ± 12%) and total error (7.84%) were unchanged at TG concentrations up to 18.5 g/L. Electrophoresis predicted National Cholesterol Education Program cut-points with <0.04 g/L error, exactly and appropriately classified 79% and 96% of the subjects, and divided by 2.4 (all subjects) and 5.8 (TGs >1.5 g/L) the percentage of subjects underestimated by calculation. One-half of the patients with TGs >4 g/L had LDLC >1.30 g/L. For HDLC, correlation was better with precipitation (r = 0.87) than ultracentrifugation (r = 0.76). Error (-0.10% ± 26%) increased when HDLC decreased (<0.35 g/L). Direct assessment of the LDLC/HDLC ratio detected 45% more high-risk subjects than the calculation/precipitation combination.
Conclusions: Electrophoresis provides reliable quantification of LDLC, improving precision, accuracy, and concordance over calculation, particularly with increasing plasma TGs. Implementation of methods to detect low cholesterol concentrations could extend the applications for HDLC assessment.
The following articles in journals at HighWire Press have cited this article:
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  M. Nauck, G. R. Warnick, and N. Rifai Methods for Measurement of LDL-Cholesterol: A Critical Assessment of Direct Measurement by Homogeneous Assays versus Calculation Clin. Chem., February 1, 2002; 48(2): 236 - 254. [Abstract] [Full Text] [PDF]
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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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