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Seven Direct Methods for Measuring HDL and LDL Cholesterol Compared with Ultracentrifugation Reference Measurement Procedures
Open accessAbstract
Background: Methods from 7 manufacturers and 1 distributor for directly measuring HDL cholesterol (C) and LDL-C were evaluated for imprecision, trueness, total error, and specificity in nonfrozen serum samples. Methods: We performed each direct method according to the manufacturer’s instructions, using a Roche/Hitachi 917 analyzer, and compared the results with those obtained with reference measurement procedures for HDL-C and LDL-C. Imprecision was estimated for 35 runs performed with frozen pooled serum specimens and triplicate measurements on each individual sample. Sera from 37 individuals without disease and 138 with disease (primarily dyslipidemic and cardiovascular) were measured by each method. Trueness and total error were evaluated from the difference between the direct methods and reference measurement procedures. Specificity was evaluated from the dispersion in differences observed. Results: Imprecision data based on 4 frozen serum pools showed total CVs <3.7% for HDL-C and <4.4% for LDL-C. Bias for the nondiseased group ranged from −5.4% to 4.8% for HDL-C and from −6.8% to 1.1% for LDL-C, and for the diseased group from −8.6% to 8.8% for HDL-C and from −11.8% to 4.1% for LDL-C. Total error for the nondiseased group ranged from −13.4% to 13.6% for HDL-C and from −13.3% to 13.5% for LDL-C, and for the diseased group from −19.8% to 36.3% for HDL-C and from −26.6% to 31.9% for LDL-C. Conclusions: Six of 8 HDL-C and 5 of 8 LDL-C direct methods met the National Cholesterol Education Program total error goals for nondiseased individuals. All the methods failed to meet these goals for diseased individuals, however, because of lack of specificity toward abnormal lipoproteins.
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“Most discordances in dyslipidemic samples are observed at lower concentration ranges of LDLC (< 1.8 mmol/L) and HDLC (< 1.0 mmol/L) [42].”
From Quantifying atherogenic lipoproteins for lipid-lowering strategies: Consensus-based recommendations from EAS and EFLM (2020) · cites this paper as [42]
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“Direct measurements of dLDLC and dHDLC in normolipidemic samples usually meet the National Cholesterol Education Program (NCEP) total error goals of ≤12% and ≤13%, respectively, but total error ranged from -26% to +32% for dLDLC and -20% to +36% for dHDLC in a comprehensive study of different assays in dyslipidemic samples [42].”
From Quantifying atherogenic lipoproteins for lipid-lowering strategies: Consensus-based recommendations from EAS and EFLM (2020) · cites this paper as [42]
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“It is important to note that direct LDL-C measurements also have limitations, including systematic bias and inaccuracy in patients with dyslipidaemia, especially for high TG levels [88-90].”
From 2019 ESC/EAS guidelines for the management of dyslipidaemias: lipid modification to reduce cardiovascular risk (2019) · cites this paper as [89]
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“Thus, discrepancies between homogeneous assays and β-quantification are found mostly in hypertriglyceridemic subjects, with some studies showing positive but minor biases (< 5%, on average), with no consistent pattern for the frequency of discordant LDL results with triglyceride concentration (22-24).”
ResultFan, G., Zhang, S., Wu, Q., Song, Y., Jia, A., Li, D., Yue, Y., & Wang, Q. (2022) · Clinica Chimica Acta · doi:10.1016/j.cca.2022.08.007 · Cited by 12
Measuring LDL Cholesterol: for Old and New Calculations, Is There an Optimal Formula?
Stein, E. A. (2014) · Clinical Chemistry · doi:10.1373/clinchem.2014.232793 · Cited by 12
Hsu, S. H.-J., Chen, M.-F., Chen, D., & Su, T. (2015) · Internal Medicine · doi:10.2169/internalmedicine.54.4308 · Cited by 11
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“Previous studies have highlighted the application of direct LDL-C assays to adult populations by demonstrating that they meet the performance standards set by the National Cholesterol Education Program (NCEP) and may serve to stratify patients into cardiovascular disease risk categories better than LDL-F.(4,5) For ped…”
MethodAssociation of plasma endothelial lipase levels on cognitive impairment
Yun, S., Park, J.-Y., Seo, S. W., & Song, J. (2019) · BMC Psychiatry · doi:10.1186/s12888-019-2174-8 · Cited by 10
Lipid reference values in an Irish population
Hughes, D., Crowley, J. J., O’Shea, P. M., McEvoy, J. W., & Griffin, D. G. (2020) · Irish Journal of Medical Science (1971 -) · doi:10.1007/s11845-020-02309-0 · Cited by 11
A Black and White Response to the “Gray Zone” for Fractional Flow Reserve Measurements
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“As far back as 2010, Miller et al. assessed seven distinct direct tests to quantify HDL-c and LDL-c [4].”
BackgroundNakamura, M., Koyama, I., Iso, H., Sato, S., Okazaki, M., Kayamori, Y., Kiyama, M., Kitamura, A., Shimamoto, T., & Ishikawa, Y. (2010) · Journal of Atherosclerosis and Thrombosis · doi:10.5551/jat.5470 · Cited by 10
Self-citationYeoh, E. C. K., Fang, S. C., Chang, S., Mun, S. L. K., Chi, L. S., Ying, Y. L., Wei, N. X., Tang, W. E., Ming, S. L. B., & Tavintharan, S. (2014) · Diabetes and Vascular Disease Research · doi:10.1177/1479164114547703 · Cited by 10
Hirano, T. (2024) · Journal of Atherosclerosis and Thrombosis · doi:10.5551/jat.65164 · Cited by 10
High Density Lipoproteins: Is There a Comeback as a Therapeutic Target?
von Eckardstein, A. (2021) · Handbook of experimental pharmacology · doi:10.1007/164_2021_536 · Cited by 9
Ghayad, J. P., & Barakett, V. (2021) · American Journal of Clinical Pathology · doi:10.1093/ajcp/aqab153 · Cited by 9
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Self-citationSecular Changes in the Prevalence of Cardiovascular Risk Factors in Japanese Workers, 2001–2011
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Rebholz, S. L. (2017) · Journal of Clinical and Laboratory Medicine · doi:10.16966/2572-9578.107 · Cited by 9
“LDL- and HDL-C were directly measured using second and third generation assays [7], respectively.”
Methodde Córdova, C. M. M., Helena, E. T. D. S., Galgowski, C., Figueira, V. H., Setter, G. B., Markus, M. R. P., Schmidt, C. O., Völzke, H., & Ittermann, T. (2018) · Scandinavian Journal of Clinical and Laboratory Investigation · doi:10.1080/00365513.2018.1432070 · Cited by 10
“It must be noted that different direct methods for LDL-c measurements used in many of these studies may also account for the discrepancies found [42].”
BackgroundIsmoyowati, I., Indrasanti, D., Mugiyono, S., & Pangestu, M. (2019) · Veterinary World · doi:10.14202/vetworld.2019.1689-1697 · Cited by 9
“Blood plasma was collected and stored at −20°C for further assay of total cholesterol, LDL, HDL, and triglyceride using a spectrophotometer at a wavelength of 500 nm and 340 nm using Hitachi 704 analyzer [24].”
MethodNaser, A. M., Rahman, M. M., UNICOMB, L. E., Doza, S., Selim, S., Chaity, M., Luby, S. P., Anand, S., Staimez, L. R., Clasen, T., et al. (2020) · Journal of the American Heart Association · doi:10.1161/jaha.119.014978 · Cited by 8
Ghayad, J. P., Barakett, V., & Sleilaty, G. (2022) · Laboratory Medicine · doi:10.1093/labmed/lmac049 · Cited by 9
Fusco, S. A. D., Volpe, M., Nardi, F., Matteucci, A., Aquilani, S., Marino, G., Aiello, A., & Colivicchi, F. (2025) · High Blood Pressure & Cardiovascular Prevention · doi:10.1007/s40292-025-00708-x · Cited by 9
“In the research field, enzymatic assays are often used as reference direct methods for LDL-C measurement [7], even though these methods do not meet the total error targets established by the National Cholesterol Education Program [8].”
Kitagawa, Y., Teramoto, T., & Daida, H. (2012) · Journal of Atherosclerosis and Thrombosis · doi:10.5551/jat.12682 · Cited by 8
Vasse, J., Lassartesse, A., Marmontel, O., Charrière, S., Bouveyron, C., Marrié, N., Moulin, P., & Filippo, M. D. (2023) · Clinical Chemistry and Laboratory Medicine (CCLM) · doi:10.1515/cclm-2023-0360 · Cited by 11
Nazli, S. A., Rosman, A., Kasim, N. A. M., Al-Khateeb, A. M., Ul-Saufie, A. Z., Radzi, A. B. M., Ibrahim, K. S., Kassim, S. S., & Nawawi, H. M. (2024) · Scientific Reports · doi:10.1038/s41598-024-53539-6 · Cited by 13
“The level of LDL-c is often measured using calculation; most commonly using Friedewald equation, instead of direct measurement which may give inaccurate results 84,85 .”
MethodWelge, J. A., Warshak, C. R., & Woollett, L. A. (2018) · The Journal of Maternal-Fetal & Neonatal Medicine · doi:10.1080/14767058.2018.1542679 · Cited by 9
“Cholesterol was measured in HDL using a two reagent method in which LDL, VLDL, and chylomicrons were immobilized prior enzymatic reactions with oxidase, whereas cholesterol in LDL was measured directly using enzymatic reagents that favor LDL or by the Friedewald equation [13,14].”
MethodEstrategia para el diagnóstico de las dislipidemias. Recomendación 2018
Estébanez, B. C., Mieras, M. P., Romero, C., Ramírez, J. C. V., Salán, M. E., Castro, M. J. C., García, E. R., Velilla, T. A., Calmarza, P., & Puzo, J. (2019) · Revista del Laboratorio Clínico · doi:10.1016/j.labcli.2019.03.001 · Cited by 8
Metadata
- Authors
- Miller, W. G., Myers, G. L., Sakurabayashi, I., Bachmann, L. M., Caudill, S. P., Dziekonski, A., Edwards, S., Kimberly, M. M., Korzun, W. J., Leary, E. T., Nakajima, K., Nakamura, M., Nilsson, G., Shamburek, R. D., Vetrovec, G. W., Warnick, G. R., & Remaley, A. T.
- Affiliations
- Virginia Commonwealth University (US)
- Centers for Disease Control and Prevention (US)
- Jichi Medical University (JP)
- Virginia Commonwealth University, Richmond, VA
- Pacific Biomarkers (United States) (US)
- Otsuka (Japan) (JP)
- Osaka Medical Center for Health Science and Promotion, Osaka, Japan
- Nilsson Measurement Quality, Uppsala, Sweden
- National Institutes of Health (US)
- Health Diagnostics Laboratory, Richmond, VA
- Corresponding author
- Miller, W Greg
- ORCID
- Venue
- Clinical Chemistry
- Publisher
- Oxford University Press (OUP)
- Published
- 2010-06-01
- Volume
- 56
- Issue
- 6
- Pages
- 977-986
- ISSN
- 0009-9147 (ISSN-L)
- 1530-8561
- PMC
- PMC4687457
- OpenAlex
- W2130132335
- Cited by
- 386
- References
- 23
- Citations by year
- 2026: 22 · 2025: 31 · 2024: 25 · 2023: 31 · 2022: 30 · 2021: 30 · 2020: 31 · 2019: 26 · 2018: 22 · 2017: 21 · 2016: 15 · 2015: 14 · 2014: 27 · 2013: 16 · 2012: 17
- FWCI
- 13.22
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Licence
- academic.oup.com/journals/pages/open_access/funder_policies/chorus/standard_publication_model (vor)
- Locations
- Clinical Chemistry (publishedVersion)
- PubMed (publishedVersion)
- PubMed Central (acceptedVersion)
- Funders
- Merck
- Pfizer
- Abbott
- NIH
- Topics
- Health Sciences › Health Professions › Medical Laboratory Technology › Clinical Laboratory Practices and Quality Control
- Health Sciences › Medicine › Cardiology and Cardiovascular Medicine › Lipoproteins and Cardiovascular Health
- Physical Sciences › Physics and Astronomy › Instrumentation › Scientific Measurement and Uncertainty Evaluation
- Keywords
- HDL cholesterol, LDL cholesterol, direct measurement method, method comparison, total error, imprecision, trueness, specificity, dyslipidemia, cardiovascular disease
- MeSH terms
- Blood Chemical Analysis, Humans, Cholesterol, HDL, Cholesterol, LDL, Reference Standards, Sensitivity and Specificity, Ultracentrifugation
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