Retrospective Analysis of Point-of-Care andLaboratory-Based Hemoglobin A1c Testing

Jennifer L. Clark1 and Lokinendi V. Rao1*

Background:Glycemic control is essential to diabeticmanagement, and hemoglobin A1c (HbA1c) has longbeen used for this purpose. Though laboratory-based testing is standard, point-of-care (POC) systemsprovide rapid results in clinic, allowingmore timely patient management. A negative bias with POC testinghas been observed, and our aim is to further characterize these discrepancies at our institution.Methods: A medical record search identified patients who underwent laboratory-based and/or POCHb A1c testing (DCA Vantage™) at our medical center from July 2015 to April 2016. Patients whounderwent both tests within 30 days were grouped by age, sex, and test interval (same day, <1 day, ≤15days, or ≤30 days). Mean laboratory-based and POC values were compared using the paired t-test.Correlation statistics were determined using the Deming regression.Results: In total, 40503datapointsweregathered fromthedatabase, comprising28555 laboratory-basedHbA1c tests and 11948 POC-basedHbA1c tests. A total of 28292uniquepatientswere identified, ofwhich 493underwent both testswithin30days.WhileDCAand laboratory-based testingwashighly correlated, therewasameannegative bias of 0.18%with POC testing. Biaswas greater for women [0.17% higher (95% CI, 0.063%–0.284%), P = 0.002] and children aged 0–13 years [0.52% higher (95% CI, 0.141%–0.891%), P = 0.007].Conclusions: There is a consistent negative bias with POC testing, most pronounced in the female andpediatric populations. Further studieswill determinewhat variables contribute to thisdiscrepancyandhowclinical management is modified. POC testing using the DCA Vantage should be interpreted cautiously.

IMPACT STATEMENTThe aim of our study was to evaluate the degree of negative bias in point-of-care (POC) hemoglobin A1c (Hb

A1c) testing, an observation that has beenwidely reported, in the diabetes clinic at our institution. On the basisof our data, we were able to identify specific groups within the diabetic population most at risk for discrepanttesting that could adversely affect clinical management. Given these findings, clinicians may more criticallyevaluate POC Hb A1c values, particularly in these vulnerable populations, before making major modificationsto a patient's treatment plan.

1Department of Pathology, UMass Memorial Medical Center, University of Massachusetts Medical School, Worcester, MA.*Address correspondence to this author at: Hospital Labs, Pathology, UMass Memorial Medical Center, One Biotech, 365 Plantation St.,Worcester, MA 01605. Fax 774-442-9604; e-mail lokinendi.rao@umassmemorial.org.DOI: 10.1373/jalm.2016.021493© 2017 American Association for Clinical Chemistry2Nonstandard abbreviations: Hb A1c, hemoglobin A1c; POC, point-of-care.

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Copyright 2017 by American Association for Clinical Chemistry.

Diabetes is the sixth leading cause of disease-related death in the United States, contributing toa wide variety of comorbidities including heart dis-ease, kidney disease, and blindness (1). Glycemiccontrol is central to the successful management ofthese patients (2). Hemoglobin A1c (Hb A1c),2 orglycohemoglobin, measurements have long beenused as an indicator of glycemic control in diabeticpatients, representing themeanblood glucose lev-els over the previous 120 days, the mean lifespanof the red cell (2–4). When increased, glycemic con-trol may be enhanced through changes in diet, life-style, andmedication regimens with repeat testingevery 3–6 months to monitor progress (5).Laboratory-based Hb A1c levels are generally

preferred, with a goal <7.0% in most diabetic pa-tients (5). However, test results are normally ob-tained after patients have already left their officevisit. Point-of-care (POC) testing systems, whichcan provide a result while the patient is still in theoffice, have been designed to bridge that gap, pro-viding real-time feedback to the patient and physi-cian and allowing more efficient and effectivemanagement decisions (6–8). Despite the demon-strated benefits of POC testing in this setting, con-cerns remain regarding the accuracy of finger-stickHb A1c testing (9, 10). Though studies have showngood correlation overall between laboratory-based and POC measurements, a systematic neg-ative bias has been frequently observed with thefinger-stick testing (9, 11–13).The aim of the current study is to address ob-

served bias in our own laboratory and identify par-ticular groups most at risk for falsely decreasedPOC Hb A1c levels.

MATERIALS AND METHODS

A retrospective medical record search was per-formed to identify all patients who underwent lab-oratory-based and/or POC Hb A1c testing at ourinstitution during a 10-month timeperiod from July2015 to April 2016. Those patients undergoing

both tests within 30 days or less were identified.The presence or absence of a hemoglobinopathywas not known. Groups were compared based ontest interval (sameday, within 1 day, within 15 days,or within 30 days), age, and sex. To correlate thevalues of the 2 tests within these groups, correla-tion coefficient (R), slope, and y-intercept were cal-culated based on the Deming regression. Todetermine the statistical significance of the meandifference between laboratory-based and POC HbA1c values between groups, the P value was calcu-lated using a Student t-test.Laboratory-based testing was performed using

the Roche Tina-quant Gen.2 Hb A1c turbidimetricinhibition immunoassay on the Integra 800 ana-lyzer (Roche Diagnostics). Hb A1c determination isbased on turbidimetric inhibition immunoassay ofhemolyzed whole blood samples. Hb A1c in thesample reacts with anti–Hb A1c antibody, forming asoluble antigen-antibody complex. Polyhaptensreact with excess anti–Hb A1c, and the formation ofan insoluble antibody-polyhapten complex is tur-bidimetrically determined. In the central labora-tory, 2 levels of QCmaterial are run twice daily. Theprecision at level 1 (Hb A1c concentration of 5.5%)QC material was <1%, and precision at level 2 (HbA1c concentration of 9.9%) was 0.7%, respectively.POC testing was performed using DCA Vantage

(Siemens Medical Solutions Diagnostics, Tarrytown,NY), which is based on latex agglutination inhibitionimmunoassay methodology. The system consists ofa spectrophotometric andprecalibrated, unitized re-agent cartridge containing both wet and dry re-agents. The immunological reaction uses a mono-clonal antibody, and light scattering is quantifiedfrom absorbancemeasured at 530 nmwith simulta-neous total hemoglobin evaluation using potassiumferricyanide.

RESULTS

A total of 40503 data points were gatheredfrom the laboratory database, comprising 28555

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laboratory-based Hb A1c tests (70.5%) and 11948POC Hb A1c tests (29.5%). Of the laboratory-basedtests, 24356 (85.3%) were performed on samplesfrom outpatients, and approximately 30% of testvalues fell within normal limits. All POC testing wasperformed in the diabetes clinics, and the majority(93.8%) of tests yielded increased Hb A1c levels.Patient sex was relatively equal in the dataset with51% females and 49% males. Patient age rangedfrom 1 year, 11 months, to 105 years, with a me-dian age of 57 years.Within the total dataset, 28292 unique patients

were identified, and many of which had multipletests during the study period. Those patients whounderwent both laboratory-based andDCA testingwithin 30 days or less were identified, consisting of499 instances. Six patients with both laboratory-based and DCA values >14% were eliminated,since this is the limit of the reportable range for theDCA test, leaving 493 in the study group. Withinthis group, 246 (49.9%) were female and 247(50.1%) were male. Ages ranged from 2 years, 7months, to 89 years, with amedian age of 57 years.Only 21 (4.2%) patients in this group exhibited HbA1c levels within the normal range during the studyperiod. Both tests were obtained on the same dayin 145 (29.4%) instances, within 1 day in 245(49.7%) instances, and within 15 days in 372(75.5%) instances. The mean interval betweentests was 8 days overall, with children aged 0–10years demonstrating the longest mean test inter-val at 18 days.

The mean laboratory-based and DCA values,ranges, correlation coefficient (R), slope of regres-sion line, y-intercept, and P values are summarizedin Table 1 for each test interval group. Correlationcoefficients between laboratory-based and DCAvalues always exceeded 0.95 with slopes rangingfrom 1.018 to 1.090 and y-intercept ranging from−0.34 to −0.90. Based on the paired t-test, therewas a statistically significant difference betweenthe laboratory-based and DCA values in each testinterval group (P <0.001 for all groups). The meanlaboratory-based and DCA values within Hb A1cranges are shown for each test interval group inFig. 1. Mean laboratory-based values were higherthanmean DCA values in the range of 6.0%–11.0%with outliers in the very low and very high ranges.Overall, DCA values were on average 0.18% (actualbias) lower than laboratory-based values. The ac-tual bias for each patient andmean bias are shownfor each test interval group in Fig. 2. The actualdifference in reported test results varies up to 1–2points (more toward negative side), and this bias isgreater at higher Hb A1c levels. This result could beattributable to poor peripheral circulation in theuncontrolled diabetic population with higher HbA1c levels. Most assays should fall within <0.3%bias according to the National GlycohemoglobinStandardization Project (NGSP) (14). In our study,14.6%of cases showedanoverestimateofHbA1c onDCA testingby≥0.3%, and41.8%of cases showedanunderestimate of Hb A1c on DCA testing by ≥0.3%.Overall, 56.4% of patients had discrepant DCA test

Table 1. Comparison of laboratory-based and POC Hb A1c tests performed within 30 days.a

Testinginterval n

Meanlab-basedHb A1c, %

Range oflab-basedHb A1c, %

MeanPOC

Hb A1c, %

Range ofPOC

Hb A1c, % R Slopey-

Intercept PA Same day 145 8.56 5.2–14.0 8.40 5.1–14 0.96 1.076 −0.81 <0.001b

B ≤1 day 245 8.49 5.2–14.0 8.36 4.2–14 0.97 1.090 −0.90 <0.001b

C ≤15 days 372 8.49 4.8–15.1 8.33 4.2–14 0.97 1.039 −0.49 <0.001b

D ≤30 days 493 8.44 4.7–15.1 8.25 4.2–14 0.95 1.018 −0.34 <0.001b

a R, slope, and y-intercept were calculated by using the Deming regression. The P value was calculated by using the paired t-test for mean lab-basedand POC-based Hb A1c tests. A–D indicate corresponding graph in Fig. 1.b Significance.

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results exceeding the maximum allowable error.These results are summarized in Table 2.In addition to identifying statistically significant

test discrepancies, we determined the extent ofclinically significant discrepancies in our data setusing the generally accepted threshold of 0.5%points Hb A1c (15). Overall, we found that the DCAvalue underestimated the Hb A1c by ≥0.5% in 106cases (21.5%). Of these, 63.2% were female. Inter-estingly, the DCA value overestimated the Hb A1cby ≥0.5% in 42 cases, and 61.9% of these weremale. Altogether, clinically significant discrepan-cieswere found in 148 cases (30.0%). These resultsare summarized in Table 2.

Mean laboratory-based Hb A1c values werehigher than DCA values in all age-groups (Fig. 3),and this result was statistically significant basedon the paired t-test (0–13 years: P = 0.018, >13years: P <0.001). The difference between testswas largest in children aged 0–13 years, whodemonstrated higher laboratory-based valuesthan DCA values by a mean of 0.69% (actual bias)compared to the >13 years of age population,who demonstrated laboratory-based values just0.18% higher. The mean test discrepancy (labo-ratory-based value minus DCA value) betweenthe 0–13 years of age and >13 years of age pop-ulations was statistically significant (P = 0.007)

Fig. 1. Mean laboratory-based and POC-based Hb A1c values at different level intervals (range from4.1% to 14%) for patients undergoing both tests.The overall mean test values are shown at each Hb A1c range for same-day testing (A), testing interval ≤1 day (B), testinginterval ≤15 days (C), and testing interval ≤30 days (D). Hb A1c ranges are based on laboratory-based value. Error barsrepresent SE. Statistical significance is indicated as follows: *P <0.05, **P <0.01, ***P <0.001.

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based on a Student t-test. These results are sum-marized in Table 3.The 2 tests were also compared based on sex (Fig.

4). Females demonstrated mean laboratory-based

values of 8.51% and mean DCA values of 8.24%,with a mean difference of 0.27%. Males demon-strated mean laboratory-based values of 8.36%and mean DCA values of 8.26%, with a mean

Fig. 2. Actual bias between laboratory-based and DCAHb A1c testing for each test interval group: sameday testing (A), testing within 1 day (B), testing within 15 days (C), and testing within 30 days (D).

Table 2. Number of laboratory-based Hb A1c tests under- or overestimated by the POC test using0.3% laboratory bias threshold and 0.5% clinically significant bias threshold out of 493 total cases.

≥0.3% Discrepancy ≥0.5% DiscrepancyPOC underestimated Hb A1c 206 (41.8%) 106 (21.5%)POC overestimated Hb A1c 72 (14.6%) 42 (8.5%)Total discrepant 278 (56.4%) 148 (30.0%)

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difference of 0.10%. The test discrepancy (laboratory-based value minus DCA value) between males andfemales was statistically significant (P = 0.002)based on a Student t-test, as was the difference

between laboratory-based and DCA values withinthe male and female groups (females: P <0.001,males: P = 0.005) based on the paired t-test. Theseresults are summarized in Table 3.

DISCUSSION

We show in the current study that there is aconsistent negative bias with finger stick DCA test-ing when compared to laboratory-based testing.This discrepancy on average is 0.18% and is statis-tically significant. This discrepancy certainly carriesclinical weight, since small changes in Hb A1c levels,even those below the accepted 0.5% clinicallythreshold, may trigger modifications in clinicalmanagement (15). A number of patients in our

Fig. 3. Mean laboratory-based and DCA Hb A1cvalues by patient age.The mean test values (A) and mean difference betweentest values (B) are shown for patients age 0–13 years and>13 years. The mean difference was calculated by sub-tracting the DCA value from the laboratory-based value.Error bars represent SE. Statistical significance is indi-cated as follows: *P <0.05, **P <0.01, ***P <0.001.

Table 3. Summary of laboratory-based and POC Hb A1c tests by sex and age.

nMean lab-based

Hb A1c, %Range of lab-based

Hb A1c, %Mean POCHb A1c, %

Range ofPOC Hb A1c, % Pa

0–13 years 11 8.14 4.7–12.0 7.45 4.5–9.7 0.018b

>13 years 482 8.45 4.8–15.1 8.27 4.2–14 <0.001b

Male 247 8.36 4.7–14.3 8.26 4.5–14 0.005b

Female 246 8.51 4.8–15.1 8.24 4.2–14 <0.001b

a P value was calculated by using the paired t-test.b Significance.

Fig. 4. Mean laboratory-based and DCA Hb A1cvalues by sex.The mean test values are shown for females and males.Error bars represent SE. Statistical significance is indi-cated as follows: **P <0.01, ***P <0.001.

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studyactuallydemonstrateddiscrepancies in labora-tory-based and DCA testing greatly exceeding 1%point, an unacceptable deviation from the generallyaccepted laboratory bias threshold <0.3% in the ac-tual numbers reported. Overall, more than half ofpatients showed a discrepancy of 0.3% or greaterbetween the 2 tests, and approximately 20% of pa-tients included in the study showed a clinically signif-icant discrepancy based on a threshold of 0.5% forclinical significance. The clinical ramificationsof thesediscrepancies are not known and deserve consider-ation in further studies.We did identify 2 groups at risk for potentially

clinically significant discrepancies on finger-stickHb A1c testing. Females demonstrated a greaterdiscrepancy than males in this study with a meannegative bias of nearly 0.3% on DCA testing, whichwas statistically significant. Though this discrep-ancy is still small, less than the 0.5% generally con-sidered to be clinically significant, this group wasclearly more prone to test discrepancies andtherefore had increased risk of inadequate glyce-mic management (16). Notably, the majority of pa-tients whose DCA test underestimated the Hb A1cby 0.5% or greater were female. The most strikingresult in this study was the highly clinically signifi-cant discrepancy seen in children aged 0–13years. In this group, there was a mean negativebias approaching 0.7% on DCA testing. Such a

discrepancy would certainly affect clinical manage-ment in this particularly vulnerable population.These results may indicate that the finger-stick tech-nique used on women and children by some phle-botomists is not ideal for POC Hb A1c testing or thatthere is some physiological difference in pediatricand female populations that affects Hb A1c levels inthe subcutaneous capillary beds. Further studies willbe needed to answer this question.There are several limitations to this study, due in

particular to its retrospective nature. Ideally, thestudywould be doneprospectively with all patientsundergoing both tests on the same day by thesame phlebotomist with a standardized finger-stick technique. In addition, more young childrenshould be enrolled in subsequent studies, as thepediatric population in the current study was quitelimited. It is important to note that these data per-tain to testing done at a single institution and maynot represent diabetic populations in all locales.However, we believe our results may be general-ized tomost diabetic patients seeking care at large,urban tertiary care centers using the same testingmethods. In conclusion, POC testing should be in-terpreted cautiously in the context of clinical deci-sion-making, particularly in pediatric and femalepopulations. In addition, care should be taken touse adequate finger-stick technique when per-forming POC tests in these groups.

Author Contributions: All authors confirmed they have contributed to the intellectual content of this paper and havemet the following4 requirements: (a) significant contributions to the conception and design, acquisition of data, or analysis and interpretation of data; (b)drafting or revising the article for intellectual content; (c) final approval of the published article; and (d) agreement to be accountable forall aspects of the article thus ensuring that questions related to the accuracy or integrity of any part of the article are appropriatelyinvestigated and resolved.

Authors’ Disclosures or Potential Conflicts of Interest: No authors declared any potential conflicts of interest.

Role of Sponsor: No sponsor was declared.

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