What factors influence the accuracy of the Hemoglobin A1c (HbA1c) test?

Factors Influencing HbA1c Accuracy

HbA1c accuracy is primarily compromised by conditions affecting red blood cell lifespan, hemoglobin variants, and laboratory/technical factors, with shortened RBC survival causing falsely low values and prolonged survival causing falsely elevated results. 1, 2

Red Blood Cell Turnover Disorders

Conditions Causing Falsely Low HbA1c

Any condition that shortens RBC survival or decreases mean RBC age produces falsely low HbA1c results regardless of the assay method used. 1, 2 The most clinically significant conditions include:

• Hemolytic anemia of any etiology falsely lowers HbA1c by reducing the time RBCs are exposed to glucose 1, 3, 4
• Recovery from acute blood loss decreases mean RBC age, artificially lowering HbA1c 1, 2
• Pregnancy alters RBC turnover, lowering usual HbA1c concentrations even in women without diabetes 2
• Chronic kidney disease can shorten erythrocyte lifespan through uremic toxins and other mechanisms, resulting in falsely low values 1, 2, 5

Conditions Causing Falsely Elevated HbA1c

• Iron deficiency anemia prolongs erythrocyte lifespan and exposes cells to glucose for longer periods, causing falsely elevated HbA1c levels 1, 2
• Occult malignancies affecting bone marrow function can alter red blood cell production and lifespan, leading to falsely elevated HbA1c 2

Hemoglobin Variants and Structural Abnormalities

HbA1c cannot be measured and should not be reported in individuals with homozygous hemoglobin variants such as HbSS or HbEE; glycated proteins like fructosamine or glycated albumin must be used instead. 1, 2

• Hemoglobin variants (HbS, HbC) can result in falsely elevated or falsely lowered HbA1c levels depending on the specific assay method used 1, 2
• Laboratories should be aware of potential interferences, including hemoglobin variants, and consider these in their particular patient population when selecting assay methods 1, 2
• A hemoglobin variant or interference should be suspected if the HbA1c reading is surprisingly high or low, or significantly changed coincident with a change in laboratory method 2

Age and Race-Related Variations

• Age-related increases in mean HbA1c occur in people without diabetes at approximately 0.1% per decade after age 30 years 1, 2
• African Americans with prediabetes have HbA1c values that are 0.4% higher than white persons for any given level of glycemia, unexplained by measured differences in glycemia, clinical factors, or sociodemographic factors 1, 2
• The American Diabetes Association recommends that a new diagnosis of diabetes be based on a confirmatory fasting blood glucose level of at least 126 mg/dL if the initial HbA1c value is 6.5% to 6.9% in populations where racial/ethnic differences may be relevant 1

Medications and Nutritional Factors

• Vitamins C and E are reported to falsely lower HbA1c results, possibly by inhibiting glycation of hemoglobin 1, 2
• Oral hypoglycemic agents (metformin or sulfonylureas) may alter the relationship between blood glucose and HbA1c levels, although clinical significance remains unclear 1, 2
• Hypertriglyceridemia, hyperbilirubinemia, chronic alcoholism, chronic ingestion of salicylates, and opiate addiction are reported to interfere with some assay methods, falsely increasing results, though these studies are old and findings may not pertain to modern methods 1, 2

Laboratory and Technical Factors

Measurement Precision

• A single HbA1c measurement, even from a high-quality laboratory, has a margin of error such that the true value falls within a range defined by the coefficient of variation 1, 2
• Sequential HbA1c values within 0.5% do not statistically differ from one another unless the assay coefficient of variation is less than 3%, ideally less than 2% 2
• Comparing HbA1c tests performed in different clinical laboratories introduces additional error due to interlaboratory variation 1, 2

Sample Handling

• Whole blood samples are generally stable for up to 1 week at 4°C 1, 2
• Samples stored at -70°C or colder are stable long term (at least 1 year), but specimens are not as stable at -20°C 1, 2
• Improper handling of specimens, such as storage at high temperatures, can introduce large artifacts that may not be detectable depending on the assay method 1

Clinical Management Algorithm

When to Suspect HbA1c Inaccuracy

HbA1c should be interpreted with caution when there is a discrepancy with self-monitored blood glucose readings, especially in patients with suspected occult disease 2

Diagnostic Workup for Discordant Results

When HbA1c appears inconsistent with clinical presentation:

1. Immediately repeat the HbA1c test using the same laboratory method and simultaneously obtain fasting plasma glucose to confirm true glycemic status 6
2. Obtain complete blood count to assess for anemia 2, 6
3. Order iron studies to evaluate for iron deficiency 2, 6
4. Check reticulocyte count to assess RBC turnover 6
5. Consider hemoglobin electrophoresis if hemoglobin variant is suspected 6

Alternative Glycemic Markers

In clinical settings where abnormalities in red blood cell turnover, hemoglobin variants, or other interfering factors compromise interpretation, assays of other glycated proteins such as fructosamine or glycated albumin may be used, although they reflect a shorter period of average glycemia (2-3 weeks) than HbA1c 1, 2, 6

• Fructosamine and glycated albumin are unaffected by RBC disorders and provide reliable glycemic assessment when HbA1c is unreliable 6, 4
• Glucose testing will be necessary for screening, diagnosis, and management in individuals with disorders that affect red blood cell turnover, as HbA1c may provide spurious (generally falsely low) results regardless of the method used 1, 2

Absolute Contraindications to HbA1c Use

Mandatory use of glucose criteria only (not HbA1c) is required for:

• Pregnant women 2, 6
• Patients with hemolytic anemia of any cause 6
• Recent blood transfusion recipients 6
• Chronic kidney disease with shortened RBC lifespan 6
• Known homozygous hemoglobin variants 1, 2, 6

Common Pitfalls to Avoid

• Treatment decisions based solely on a single HbA1c measurement without consideration of other clinical data, such as glucose monitoring results, may lead to unnecessary initiation or intensification of therapy 2
• Differences in mean red cell half-life, which may range from approximately 48 to 68 days, may explain some of the interindividual variability in the relationship between measured average glucose and HbA1c levels 1
• Ensure laboratory uses NGSP-certified methods traceable to DCCT reference standards to maintain accuracy and comparability 6, 7