August 2008: Volume 34, Number 8

CDC Cholesterol Standardization Program Celebrates 50 Years of Service
From Modest Beginnings to Landmark Improvements in Healthcare
By Gerald R. Cooper, MD, PhD   

In September, the CDC will mark the 50th anniversary of its cholesterol standardization program. Through the efforts of many investigators, this program has developed tools to insure accuracy and has earned recognition as a quality approach to improve lab testing. Here is a look back on how this program evolved, as well as a look into the future.

From Modest Beginnings

The need for a cholesterol reference laboratory originally stemmed from the 1957 Conference on Longitudinal Cardiovascular Studies. Cardiovascular researchers had exchanged serum specimens and found disagreement among the cholesterol measurements performed in each investigator’s respective laboratory. As a result, clinicians and laboratorians agreed that cholesterol measurements had to be standardized to obtain measurement results that were reproducible within and comparable between laboratories. CDC was asked to develop a cholesterol reference laboratory and serve as a neutral party to help standardize cholesterol measurements among cardiovascular epidemiologic laboratories.

By the end of that year, CDC had established a Cholesterol Methodology Development Laboratory and Standardization Office staffed by: Eloise Eavenson, PhD, chemist; A. John Schneider, MD, medical officer; Myron Willis, PhD, statistician; and Gerald R. Cooper, PhD, medical director. Their plan was to develop suitable serum-based reference materials for cholesterol and then design a program that would focus on standardizing cardiovascular epidemiologic laboratories. Among the program’s major objectives were: determining whether lyophilized human serum could be used as a standard reference material for total cholesterol analyses; preparing highly purified cholesterol for use as the primary standard; and measuring and assessing the significance of differences among labs that determined total cholesterol measurements. To accomplish these objectives, the group prepared and distributed lyophilized serum specimens of different cholesterol content so that they could collect and evaluate analytical data for various labs.

The First Cooperative Cholesterol Standardization Program

In 1958, the agency initiated the first Cooperative Cholesterol Standardization Program (CCSP) with seven cardiovascular epidemiologic laboratories. The CCSP was organized in three phases for participating laboratories. During the first phase, participating laboratories used blind, randomized duplicates to measure within-laboratory precision. In phase two, a lab’s techniques had to be deemed acceptable, and the lab had to produce comparable coefficients of variation (CVs). The third phase measured variability in cholesterol concentrations reported on human serum pools. Since a cholesterol reference method was not yet available, the goal was to assist the labs in maintaining their cholesterol serum levels as determined on quality-control serum pools and on the pools of the CCSP. The average standard deviation of the first CCSP reported results was 10.6 mg/dL (0.27 mmol/L) for pools in the 150–225 mg/dL (3.88–5.82 mmol/L) range. Six of the seven laboratories used modifications of the Abell-Kendall (A-K) method. The seventh laboratory used an alcohol-ether extraction and ferric chloride-acetic acid-sulfuric acid color-developing reagent procedure.

After an extensive evaluation and comparison of five cholesterol methods was conducted, CDC selected the A-K procedure as the cholesterol reference method for the CCSP on the basis that a CV of 1% was demonstrated, the procedure and reagents appeared valid, and fewer problems were encountered in daily use.

National Heart Institute Collaboration

CDC’s program got another boost when Jim Watt, MD, Director of the National Heart Institute, attributed the failure of heart disease studies to questionable cholesterol measurements. In 1961, he initiated a collaboration with CDC to expand the CCSP standardization services to all epidemiologic lipid labs supported by NHI. By 1965, the CCSP provided services to at least 65 participating cardiovascular labs in the U.S. and 18 labs in other countries.

CDC Becomes a WHO Collaborating Center

Another landmark in the cholesterol standardization program occurred in 1962 when WHO appointed CDC as a Collaborating Center for Reference and Research in Blood Lipids. The objective was to offer cholesterol standardization assistance to international research labs supporting WHO studies of CVD. This designation opened the door for CDC to broaden its standardization influence around the globe.

The Program Grows

From 1966 through 1968, CDC implemented an Experimental Triglyceride Standardization Program with 19 laboratories. The program goal was to determine the precision and accuracy that participating labs attained using different methods for measuring triglyceride concentrations. The first reference method, developed in cooperation with Hugh Lofland, MD, of Wake Forest University School of Medicine, was a method that used silicic acid-chloroform extraction, chemical hydrolysis, and color development by metaperiodate-arsenite-chromotropic acid reagent. All results were to be reported in mmol/L to prevent errors resulting from different primary standards. From the reported triglyceride values, the researchers estimated that the maximum allowable limit for reported means should be ±0.1 mmol/L and 5% for reported analytical CV. The initial Cooperative Triglyceride Standardization Program in 1968 had 105 participating laboratories and was modeled after the CCSP. By 1974, the cholesterol and triglyceride standardization efforts were combined into a Cooperative Cholesterol and Triglyceride Standardization Program (CCTSP), and the same human serum pools were used for both analyses.

During the 1970s, medical interest in the role of HDL-C as a risk factor for CVD skyrocketed and the need for accurate measurement of HDL-C became apparent. In 1981, CDC added standardization for HDL-C and instituted a reference method for it. To reflect the broader emphasis on total cholesterol, triglycerides, and HDL-C, as well as the close collaboration between CDC and NHLBI, the name of the program was changed in 1984 to the CDC-NHLBI Lipid Standardization Program.

The First Reference System in Clinical Chemistry

Another milestone in the program’s history occurred in 1975. That year, AACC formed a Cholesterol Reference Method Study Group to select and evaluate potential reference methods for cholesterol measurement. The group recommended that CDC study the A-K method and an enzymatic method as two candidate reference methods and that NIST develop a definitive method. After extensive studies were completed, the A-K method was selected as the reference method and isotope dilution-mass spectrometry was chosen as the definitive method. The CDC A-K reference method, the NIST ID-MS definitive method, the NIST SRM 911 pure cholesterol primary standard, and the serum reference materials prepared by CDC and offered by NIST, became the components of the National Reference System for Cholesterol, the first officially recognized reference system in clinical chemistry.

Where to from Here?

In recent years, the number of candidate risk factors for CVD and its complications has grown considerably. These biomarkers are termed emerging or novel risk factors because they are associated with an increased risk for CVD, but their causative, independent, quantitative contributions to the disease are not yet fully documented, nor have standardized methods for measurement been developed.

Today, CDC is focusing its efforts on biomarkers that have been identified by experts as important to cardiovascular laboratory medicine. Among the leading candidates are cTn I, BNP, urine albumin, cystatin C, CRP, apo AI, and apo B. Building on a 50-year history of successful accomplishments that have made it a global leader in clinical laboratory standardization, CDC plans to pursue collaborative efforts with other agencies and organizations to develop standardization programs for these important biomarkers of the future.

Gerald Cooper is Medical Research Officer in the Clinical Chemistry Branch in the CDC’s Division of Laboratory Sciences.