doi: 10.1111/j.1753-0407.2010.00109.

Journal of Diabetes 3 (2011) 36

COMMENTARY

Analytical goals for HbA1c: Are HbA1c results good enough for optimal use?

In the mid-to-late 1990s, results from the Diabetes Control and Complications Trial (DCCT) and the United Kingdom Prospective Diabetes Study (UKPDS) showed conclusively that intensive glycemic control signicantly reduced the risk and progression of long-term complications of diabetes.1,2 These results highlighted the use of HbA1c for monitoring of glycemic control in diabetes and set the stage for the use of specic diabetes treatment goals using HbA1c. Since that time, most clinical diabetes organizations have included specic goals for diabetes treatment in terms of HbA1c. Since small differences in %HbA1c translate into large differences in the rate of progression of complications, it is extremely important for HbA1c results to be accurate and precise, with harmonization between methods and laboratories. The National Glycohemoglobin Standardization Program (NGSP), along with manufacturers, have improved the quality of HbA1c methods over the last 1520 years, but there is still a need for even greater accuracy and precision with some available methods. The International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) provides a denitive reference method for HbA1c that facilitates traceability to a true accuracy base. There is a tight linear correlation between results of the NGSP and IFCC networks; however, the actual numbers reported by the NGSP and IFCC are different, which has led to controversy over which numbers should be reported by laboratories. In 2007 and again in 2010, consensus statements recommended the reporting of both numbers worldwide: NGSP in %HbA1c and IFCC in mmol mol Hb. Importantly, the relationship between the NGSP and IFCC laboratory networks has been established in a master equation: NGSP% = (0.0915 IFCC [mmol mol] + 2.15), which continues to be monitored.3 The two groups now serve complementary purposes: the IFCC provides manufacturers with secondary reference materials with which to anchor their methods and the NGSP provides certication with dened acceptable performance limits that are based on clinical requirements. Many countries have already

recommended the reporting of both numbers, but the USA will continue to report only the NGSP %HbA1c according to American Diabetes Association (ADA) recommendations. Regardless of which number is reported (NGSP or IFCC), it is essential that HbA1c be measured with precision and accuracy to enable healthcare providers to reliably distinguish optimal from suboptimal glycemic control. The recent recommendation to use HbA1c for the diagnosis of diabetes further emphasizes the need for optimal assay performance.4,5 Current assay performance In order to improve method performance for HbA1c, the NGSP and the College of American Pathologists (CAP) have been progressively tightening their acceptance limits for method certication and laboratory performance, respectively. HbA1c assay performance has improved considerably over the past 10 years, with overall coefcients of variation (CVs) (across different methods in different laboratories) now approximating 4%. In the normal range, the decrease has been from CVs of 7% down to <4% over the past 10 years.6 Between-laboratory CVs for some methods are <2%. Figure 1 shows the mean 2SD for each method on the 2010 survey compared to the CAP limits. Clearly some methods are better than others, but 95% of laboratories are reporting results within 8% of the NGSP target (CAP acceptance limit).7 This CAP limit will be tightened in 2011 to 7% in order to motivate manufacturers and laboratories to improve further. Are HbA1c results good enough for optimal use of the test? Many people believe that performance goals for HbA1c testing should be based on clinical requirements. There are two important questions for a physician to consider when using HbA1c measurements to assess a patients glycemic status. One question is whether the patients glycemic control is stable, improving or deteriorating,
3

2011 Ruijin Hospital, Shanghai Jiaotong University School of Medicine and Blackwell Publishing Asia Pty Ltd

Commentary

Figure 1 Mean and 2SD bars for each method group in the 2010 College of American Pathologists (CAP) GH2-B survey for the mid-level HbA1c sample. The dotted line shows the National Glycohemoglobin Standardization Program (NGSP) target value and the shaded area indicates the 8% acceptable CAP limit.

and the second is how the HbA1c result compares to the individuals target HbA1c (the current ADA recommendation is 7% with the qualication that lower is better if feasible). In answer to the rst question, many physicians have suggested that 0.5% HbA1c is a clinically signicant change. Importantly, treatment guidelines and algorithms from various clinical organizations recommend evaluating new treatment regimens in terms of whether HbA1c is lowered by 0.5% points or more.8,9 Therefore, it is important to be sure that a change of this magnitude is statistically signicant and not due to analytical variation. Taking a statistically signicant difference of 0.5% HbA1c at a HbA1c concentration of 7% as our goal for HbA1c measurement, one can use the reference change value (RCV), to calculate an appropriate goal in terms of a methods CV.10 For sequential results to be signicantly different, the numbers must differ by more than the combined variation inherent in the two results. Assuming a within-subject biological variation of <1%,11 if the analytical CV of a HbA1c method is 2% (feasible for many commercially available HPLC systems), then the RCV (95% probability) is <0.5% HbA1c (0.43%). We can conclude that, at least when HbA1c is measured in an accredited laboratory with an accurate and precise assay method, physicians can be reasonably (95%) certain that a difference of 0.5% HbA1c between successive patient samples represents a statistically signicant change in glycemic control. If the laboratory is using a method that demonstrates a within-laboratory CV of >2%, as is still the case with some assay methods, there is less condence that the difference of 0.5% HbA1c is signicant. In the situation where a physician wants to look at the difference between a patients result and a goal of 7% HbA1c, both the bias and variability (% CV),
4

in other words the total error, of the method must be taken into account. For example, if a method has zero bias, a CV of 3.5% is required to have 95% condence that the HbA1c result for a patient with a true result of 7% will read between 6.5% and 7.5% (7%). If there is a bias of 0.2% HbA1c, the CV requirement would tighten to 2.3%. While we have made considerable progress in improving HbA1c method performance, it is clear that further improvement is necessary in order for the test to be used optimally. Hopefully, further tightening of NGSP and CAP limits will lead to better accuracy and precision of HbA1c methods. HbA1c interferences Another area of concern, other than precision and accuracy, is the potential for erroneous HbA1c results due to interference. HbA1c can be accurately assessed in the vast majority of subjects, but in cases where the subject may have a medical condition that can interfere with HbA1c, or if a hemoglobin variant may be present, the results should be interpreted with caution. Conditions such as hemolytic anemia, renal failure or polycythemia can alter the red cell lifespan, thereby falsely lowering or increasing HbA1c results. Severe iron-deciency anemia has also been reported to interfere with HbA1c results to some degree.12 The effects of common hemoglobin variants on HbA1c results have been studied extensively1315 and these ndings are summarized on the NGSP web site.16 Hemoglobin variants usually occur in the heterozygous form; for these individuals and individuals with elevated HbF interferences are method-specic and HbA1c can be measured accurately if an appropriate assay method is used. HbAS and HbAC (two of the most common variants in the world) interfere with some immunoassays, although the most widely used

2011 Ruijin Hospital, Shanghai Jiaotong University School of Medicine and Blackwell Publishing Asia Pty Ltd

Commentary

immunoassays are now free of this interference. HbAE and HbAD interfere with some HPLC methods, but do not affect immunoassays. With ion-exchange methods, chromatograms generally show aberrant peaks, indicating the presence of a variant and thus enabling detection of unacceptable HbA1c results prior to their being reported. For subjects with HbF >10%, a method that does not show interference from this variant should be used. As with any test, results that are inconsistent with the clinical presentation should be investigated further. For individuals with certain homozygous or double heterozygous variants (e.g. HbSS, HbCC, and HbSC) that cause a shortened red cell lifespan, the HbA1c results will be inaccurate regardless of the assay method, and alternative methods for assessing glycemic control must be used. Other factors have been reported to inuence HbA1c results. Reports have indicated that HbA1c increases slightly with age, even after adjusting for blood glucose levels,17 but there is some question as to whether this necessitates age-specic treatment goals. It has also been reported that HbA1c results vary slightly among ethnic groups,18,19 but there is some controversy over the clinical signicance of these differences and whether they are due to differences in mean blood glucose or other factors. Summary The NGSP has been highly successful in terms of reducing the variability of HbA1c results among methods and laboratories by harmonizing results to clinical trials that established the relationships between HbA1c and the risks for diabetic complications. The IFCC reference method and materials facilitate metrological traceability of HbA1c results to a higher order reference system. The issue of which numbers are to be reported in clinical settings (NGSP% or IFCC mmol mol Hb) is being decided on a country-by-country basis, but there is an established equation where results can easily be converted between the two systems. Nonetheless, there is still room for improvement. While many assay methods demonstrate performance that is sufcient to meet clinical needs (in terms of reliably distinguishing differences of 0.5% HbA1c), some still do not. Moreover, optimal assay performance is more crucial than ever, given the recent recommendation to use HbA1c for the diagnosis of diabetes. Future reductions in the NGSP and CAP acceptance limits are likely, and the impact of these measures on the quality of HbA1c results will continue to be evaluated. Additionally, the NGSP will continue to evaluate the impact of potential interferences on HbA1c results,

with the goal of encouraging the development and use of methods that are free of common interferences. Taken together, these measures will hopefully ensure that HbA1c testing is sufciently reliable to meet clinical needs. Randie R. Little1,2 and Curt L. Rohlng1 Departments of 1Pathology & Anatomical Sciences, and 2 Child Health, University of Missouri School of Medicine, Columbia, Missouri, USA Email: littler@health.missouri.edu References
1. DCCT Research Group. The effect of intensive treatment of diabetes on the development and progression of long term complications in insulin dependent diabetes mellitus. N Engl J Med. 1993; 329: 97786. 2. UK Prospective Diabetes Study (UKPDS) Group. Intensive blood-glucose control with sulphonylureas or insulin compared with conventional treatment and risk of complications in patients with type 2 diabetes (UKPDS 33). Lancet. 1998; 352: 83751. 3. Geistanger A, Arends S, Berding C et al. Statistical methods for monitoring the relationship between the IFCC reference measurement procedure for hemoglobin A1c and the designated comparison methods in the United States, Japan, and Sweden. Clin Chem. 2008; 54: 137985. 4. International Expert Committee. International Expert Committee report on the role of the A1c assay in the diagnosis of diabetes. [see comment]. Diabetes Care. 2009; 32: 132734. 5. American Diabetes Association. Standards of medical care in diabetes. Diabetes Care. 2010; 33: S1161. 6. Little RR, Rohlng CL, Sacks DB, for the National Glycohemoglobin Standardization Program Steering C. Status of hemoglobin A1c measurement and goals for improvement: from Chaos to order for improving diabetes care. Clin Chem. DOI: 2010.148841. 7. College of American Pathologists. Surveys 2010 GH2-A Glycohemoglobin Participant Summary. Vol. College of American Pathologists, Notheld (IL), 2010. 8. Nathan DM, Buse JB, Davidson MB et al. Medical management of hyperglycemia in type 2 diabetes: a consensus algorithm for the initiation and adjustment of therapy: a consensus statement of the American Diabetes Association and the European Association for the Study of Diabetes. Diabetes Care. 2009; 32: 193 203. 9. National Institute for Health and Clinical Excellence (NICE). Type 2 diabetes: newer agents for blood glucose control in type 2 diabetes. Available from: http:// www.nice.org.uk/nicemedia/live/12165/44318/44318.pdf (Accessed September 2010).

2011 Ruijin Hospital, Shanghai Jiaotong University School of Medicine and Blackwell Publishing Asia Pty Ltd

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10. Fraser CG, Harris EK. Generation and application of data on biological variation in clinical chemistry. Crit Rev Clin Lab Sci. 1989; 27: 40937. 11. Rohlng C, Wiedmeyer HM, Little R et al. Biological variation of glycohemoglobin. Clin Chem. 2002; 48: 11168. 12. Tarim O, Kucukerdogan A, Gunay U, Eralp O, Ercan I. Effects of iron deciency anemia on hemoglobin A1c in type 1 diabetes mellitus. Pediatr Int. 1999; 41: 35762. 13. Little RR, Rohlng CL, Hanson S et al. Effects of hemoglobin (Hb) E and HbD traits on measurements of glycated Hb (HbA1c) by 23 methods. Clin Chem. 2008; 54: 127782. 14. Mongia SK, Little RR, Rohlng CL et al. Effects of hemoglobin C and S traits on the results of 14 commercial glycated hemoglobin assays. Am J Clin Pathol. 2008; 130: 13640. 15. Rohlng CL, Connolly SM, England JD et al. The effect of elevated fetal hemoglobin on hemoglobin A1c

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results: ve common hemoglobin A1c methods compared with the IFCC reference method. Am J Clin Pathol. 2008; 129: 8114. NGSP. HbA1c assay interferences. Available from: http://www.ngsp.org/interf.asp (accessed November, 2010). Pani LN, Korenda L, Meigs JB et al. Effect of aging on A1c levels in individuals without diabetes: evidence from the Framingham Offspring Study and the National Health and Nutrition Examination Survey 20012004. Diabetes Care. 2008; 31: 19916. Herman WH, Ma Y, Uwaifo G et al. Differences in A1c by race and ethnicity among patients with impaired glucose tolerance in the Diabetes Prevention Program. Diabetes Care. 2007; 30: 24537. Herman WH, Dungan KM, Wolffenbuttel BH et al. Racial and ethnic differences in mean plasma glucose, hemoglobin A1c, and 1,5-anhydroglucitol in over 2000 patients with type 2 diabetes. J Clin Endocrinol Metab. 2009; 94: 168994.

2011 Ruijin Hospital, Shanghai Jiaotong University School of Medicine and Blackwell Publishing Asia Pty Ltd