Cardiovascular disease (CVD) is a leading cause of death and a major cause of disability in the United States.1 More than half of all men and women over the age of 45 will develop heart disease in their lifetime. Identifying high risk individuals and implementing timely interventions to delay the progression of CVD has a major public health impact in terms of health care expenses and quality of life.

Total cholesterol (TC) and total triglycerides (TG) measurements continue to be used extensively as the basis of CVD risk assessment. Risk calculations also incorporate the levels of high density lipoproteins (HDL) and low density lipoproteins (LDL), as these are considered the main lipoprotein particles that facilitate cholesterol transport and fatty acid metabolism. Traditionally, HDL and LDL levels are measured based on total cholesterol content (HDL-C and LDL-C). In recent years, large population studies showed that HDL and LDL particle number (HDL-P and LDL-P) more significantly correlate with CVD risk than HDL-C and LDL-C levels.2, 3 One explanation of this mainly epidemiologic phenomenon appears to be the size polydispersity of HDL and LDL particles. At the same HDL-C and LDL-C levels, a higher number of small HDL particles is anti-atherogenic because it enhances cholesterol clearance from peripheral tissues and arterial plaques, while a higher number of small LDL particles is pro-atherogenic because it increases the development of arterial plaques. Another theory is that the HDL-P and LDL-P correlation to CVD risk originates from up and down regulation of various metabolic pathways. These pathways are responsible for elevated circulation or clearance of small HDL and LDL particles, where size is a reflection of significant variations in apolipoprotein composition and a marker of metabolic irregularities (ApoA-I, A-II, A-IV, B-100, C-I, C-II, C-III, E and more).

In spite of growing evidence that particle number is a potentially stronger CVD risk predictor,4 the more general use of clinical assays for HDL-P, LDL-P, and apolipoprotein biomarkers is impaired by lack of satisfactory inter-laboratory comparison and characterized, traceable reference standards. Using its longstanding leadership position and expertise in standardizing biomarkers for cardiovascular and other chronic diseases, the CDC is developing a comprehensive method to measure lipoprotein particle size and composition for CVD risk assessment. This method is based on preparative sub-fractionation of HDL and LDL particles based on size. The method uses asymmetric flow field-flow fractionation, which is a unique size separation technique allowing the preparative collection of intact lipid particles and their size characterization by multiple techniques, in a manner that is traceable to universal particle/molecule size calibrators. The size characterization of the HDL and LDL sub-species is coupled with targeted quantitative analysis of apolipoproteins by mass spectrometry, giving particle size distribution profiles of specific apolipoproteins. Primarily, this method workflow can be used for characterization of particle size distribution and apolipoprotein composition of reference materials. Furthermore, this method can be applied in a high throughput manner during epidemiological studies and contribute to improved interpretation of the correlation between HDL-P, LDL-P levels and CVD risk by health professionals.


  1. Murray, C. J. L., Lopez, A. D., The Lancet 1997, 349. 1498-1504, DOI:
  2. Otvos, J. D., Jeyarajah, E. J., Cromwell, W. C., The American Journal of Cardiology 2002, 90. 22-29, DOI:
  3. Bradley, R. D., Oberg, E. B., Integrative medicine (Encinitas, Calif.) 2008, 7. 18-23.
  4. El Harchaoui, K., van der Steeg, W. A., Stroes, E. S. G., Kuivenhoven, J. A., Otvos, J. D., Wareham, N. J., Hutten, B. A., Kastelein, J. J. P., Khaw, K.-T., Boekholdt, S. M., Journal of the American College of Cardiology 2007, 49. 547-553, DOI: