Clinica Chimica Acta 476 (2018) 67–74

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Clinica Chimica Acta

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Recognition of rare hemoglobin variants by hemoglobin A1c measurement T

procedures
Sydney W. Stricklanda, Sean T. Campbella, Randie R. Littleb, David E. Brunsa,
⁎
Lindsay A.L. Bazydloa,
a
Department of Pathology, University of Virginia School of Medicine, Charlottesville, VA, United States
b
Department of Pathology & Anatomical Sciences, University of Missouri School of Medicine, Columbia, MO, United States

A R T I C L E I N F O A B S T R A C T

Keywords: Background: Unrecognized hemoglobinopathies can lead to measured hemoglobin A1c (Hb A1c) concentrations
Hemoglobin variants that are erroneous or misleading. We determined the effects of rare hemoglobin variants on capillary electro-
Hemoglobin A1c phoresis (CE) and HPLC methods for measurement of Hb A1c.
Diabetes Methods: We prospectively investigated samples in which Hb A1c was measured by CE during a 14-month period.
Ion-exchange HPLC
For samples in which the electropherograms suggested the presence of rare hemoglobinopathies, hemoglobin
Capillary electrophoresis
variants were identified by molecular analysis or by comparison with electropherograms of known variants.
Boronate affinity chromatography
When sample volume permitted, Hb A1c was measured by 2 HPLC measurement procedures and by boronate
affinity HPLC.
Results: Hb A1c was measured by CE in 33,859 samples from 26,850 patients. 15 patients (0.06%) were iden-
tified as having rare hemoglobinopathies: Hbs A2 prime, Agenogi, Fannin-Lubbock I, G Philadelphia, G San Jose,
J Baltimore, La Desirade, N Baltimore, Nouakchott, and Roanne. Among 6 of these samples tested by 2 ion-
exchange HPLC methods, the rare Hb was detected by both HPLC methods in only one sample, and none were
detected by boronate affinity HPLC. The mean of the Hb A1c results of 2 HPLC methods differed from the result of
the CE method by 0.7–2.2% Hb A1c in samples with variant hemoglobins versus < 0.2% Hb A1c in samples
without variants.
Conclusion: Measurement procedures differ in the ability to detect the presence of rare Hb variants and to
quantify Hb A1c in patients who harbor such variants.

1. Introduction HPLC, immunoassay, or boronate affinity HPLC. As HPLC measurement

Hemoglobin A1c (Hb A1c) is formed by non-enzymatic glycation of
the N-terminal valine of the hemoglobin beta chain [1], and the rate of
the glycation reaction is proportional to the concentration of glucose
[2,3]. Increased Hb A1c reflects increased mean plasma glucose con-
centrations over the preceding months [4] and is correlated with in-
creased rates of occurrence of microvascular complications of diabetes
[5]. In 2009, after years of clinical use to monitor glycemic control in
patients with diabetes, Hb A1c was recommended as a diagnostic cri-
terion for diabetes [6], placing increased demands for accuracy on
methods to quantify Hb A1c [7].
Numerous studies have shown that the results of Hb A1c measure-
ment procedures are altered in patients who have common hemoglobin
variants including Hbs C, D, E, and S [10–13]. These studies have been
conducted almost exclusively with measurement procedures that used
procedures have evolved, resolution of Hb A from the common he-
moglobin variants has improved, minimizing their impact on Hb A1c
quantification. > 1000 less common hemoglobin variants have been
recognized [13], however, and little is known about their effect on
measurements of Hb A1c. In a previous short report, we described an
unexpectedly high prevalence of “rare” hemoglobinopathies among
samples submitted for measurement of Hb A1c [14].
A recently introduced Hb A1c measurement procedure uses capillary
electrophoresis (CE) [16], a technique that has high resolving power for
Hbs. Indeed, CE is used in a related measurement procedure to identify
Hb variants [17,18]. The relatively new CE approach for Hb A1c mea-
surement, although potentially attractive, was used by only 1.2% of
clinical laboratories in the U.S. participating in the proficiency testing
program of CAP in December 2016 [19]. Few studies have examined
the ability of this CE measurement procedure for Hb A1c to resolve rare

Abbreviations: Hb A1c, hemoglobin A1c; CE, capillary electrophoresis; Hb, hemoglobin

⁎
Corresponding author at: Department of Pathology, PO Box 800168, University of Virginia School of Medicine, Charlottesville, VA 22908, United States.
E-mail address: lal2s@hscmail.mcc.virginia.edu (L.A.L. Bazydlo).

https://doi.org/10.1016/j.cca.2017.11.012
Received 29 September 2017; Received in revised form 10 November 2017; Accepted 13 November 2017
Available online 14 November 2017
0009-8981/ © 2017 Elsevier B.V. All rights reserved.
S.W. Strickland et al.
hemoglobin variants or the effects of such variants on the measured
concentration of Hb A1c [20,21].
2. Material and methods
Hb A1c was measured by CE, within 12 h of collection, by use of the
Sebia Capillarys 2 Flex Piercing system, per manufacturer's instructions.
The method identifies the presence of hemoglobins A, F, A2 and A1c,
and electrophoretically separates each form by charge and size within
the capillary. Detection is performed with an absorbance reading at
415 nm and the software calculates Hb A1c% as Hb A1c/(Hb A1c + Hb
A0), but will not calculate the Hb A1c% when there are aberrant peaks
from specific Hb variants (not including common Hb variants: Hbs C, D,
E, and S) or 1 of the primary peaks (A, F, A2 or A1c) is missing or if Hb F
is > 15%. Between October 2015 and December 2016, we identified all
samples for this study as those that were flagged during routine use of
the CE method as having “Atypical Profiles” and for which no Hb A1c%
was reported, thus suggesting that an uncommon Hb variant might be
present. The medical laboratory scientists held these electropherograms
for review. For each patient, we searched the medical record to de-
termine if a Hb A1c result had been reported from the Tosoh G7 method,
which had been used to measure Hb A1c prior to October 2015. During
the 14-month study period, the CVs of the Sebia CE measurement
procedure were 6.1% and 1.6% at concentrations of 5.3% Hb A1c and
8.2% Hb A1c, respectively. Results of CAP proficiency-testing samples
differed from the results of the reference method by a mean of 0.17%
(range 0.07%–0.43%) of the reference method result. A retrospective
audit was conducted upon completion of the study in which one of the
authors (SWS) reviewed all electropherograms from 1 month (March
2017) that were identified by the software as having an “Atypical
Profile”. No additional electropherograms suspicious for hemoglobi-
nopathies were found, suggesting that the protocol for identifying
samples for study had been successful.
For study samples of adequate volume, Hb A1c was measured also by
the Tosoh G8 (ion exchange HPLC) and the Trinity Biotech ultra2
(boronate affinity HPLC) methods at the Diabetes Diagnostics
Laboratory at the University of Missouri. Samples were also sent to
Mayo Medical Laboratories for Hb A1c testing via the Bio-Rad Variant II
Turbo (ion-exchange HPLC) Hb A1c method procedure and for mole-
cular testing to identify unknown hemoglobin variants. The ion ex-
change HPLC methods separate Hb species (labile and stable Hb A1c, Hb
68
Clinica Chimica Acta 476 (2018) 67–74
Fig. 1. Patient Flow Chart. Between September 2015–November
2016, 36 individuals were identified as having “Atypical
Profiles” as reported by the SEBIA CAPILLARYS system and
could not be reported.
A, and Hb F) based primarily upon charge alone, in comparison to CE
that separates based upon size and charge. Detection is performed with
an absorbance reading at 415 nm and the software calculates Hb A1c as
Hb A1c/“other Hb species”, which can include Hb A1a, Hb A1b, Hb A0,
and/or Hb F and varies based upon vendor and methodology used.
2.1. Patients and blood samples
Blood samples from patients at the University of Virginia were
collected in tubes containing K3EDTA (BD Vacutainer). The patient
demographics of the hospital are: 78.6% White, 17.1% Black, 3.2%
“other”, 1% Asian, and 0.1% American Indian, which closely reflects
demographics of Charlottesville city and contiguous counties (not
shown). The study protocol was approved by the Institutional Review
Board at the University of Virginia.
3. Results
During the 14-month study period, Hb A1c was measured by CE in
33,859 samples from 26,850 patients, including 19,601 patients (73%)
who self-identified as white, 5907 (22%) as black, and 1342 (5%) as
“other”. The CE measurement procedure flagged approximately 90
samples per month as having a Hb variant that was recognized by the
analyzer and reported results for Hb A1c; most of these samples con-
tained the common Hb variants S (sickle trait), C and E; the relative
prevalences of these common variants were consistent with the ex-
pected prevalences in the patient population. Samples from 36 patients
were flagged as having an “atypical profile”, and no Hb A1c result was
clinically reportable. Among these 36 patients, 4 patients were lost to
follow-up or were deceased and could not be further investigated
(Fig. 1). In 15 patients, no Hb A1c result was reported because of re-
cognized interferences (such as Hb F > 15% [16]) or the presence of
homozygous Hb S or Hb C (Fig. 1). The 17 remaining electro-
pherograms strongly suggested the presence of rare hemoglobin var-
iants, for an overall prevalence of suspected rare hemoglobinopathies of
0.06%. Among these 17 patients, 9 patients self-identified as white, 6 as
black, and 2 as “other”, yielding prevalence rates of suspected he-
moglobinopathies of 0.04%, 0.1%, and 0.15%, respectively, in the 3
self-identified groups.
Among the 17 patients in whom hemoglobinopathies were sus-
pected, 11 had been tested in previous months by the Tosoh G7 HPLC
S.W. Strickland et al. Clinica Chimica Acta 476 (2018) 67–74

Table 1
Detection of hemoglobin variants by CE- and HPLC-based measurement procedures.

Hb variants found by Capillarys2 Mutation Number of patients Presence of variant detected by Presence of variant detected by Presence of variant detected by
identified† Tosoh G7?c Tosoh G8? Variant II Turbo?

Hb Fannin-Lubbock I β-G119D 1 No No No
Hb La Desirade β-A129V 4 No Yes/No No
Hb Roanne α-D94E 1 No No No
Hb G-San Jose β-E7G 1 No – –
Hb A2 prime δ-G16R 1 No – Yesb
Hb G-Philadelphia/Hb AS α-N268K 1 Yes Yesa Yesa
Hb J-Baltimore β-G16D 3 No Noa Yesa
Hb N-Baltimore/Hb AS β-K95E 1 Yesa Noa Yesa
Hb Agenogi β-E91K 1 – – –
Hb Nouakchott α-P114L 1 – – –

a
Indicates data from previous report [16].
b
Indicates data from previous report [19].
c
Includes data from a previous report [14].

Fig. 2. Expected electropherogram and chromatogram for Hb A1c in a

sample with no hemoglobin variant. A: Expected pattern of Sebia
Capillarys electropherogram and B: Expected pattern of Tosoh G8
chromatogram.

69
S.W. Strickland et al.
Fig. 3. Hb A1c analysis on samples from 2 patients with confirmed Hb La Desirade. Panels A and C: Sebia Capillarys electropherograms; Panels B and D: Tosoh G8 chromatograms. Panels
A and B: Patient 1; Panels C and D, Patient 2. Arrows indicate peaks not seen in samples from patient lacking hemoglobin variants.
method for Hb A1c quantification. As previously described [14], the
HPLC method reported Hb A1c results for 10 of the 11 patients (91%)
with no indication that a hemoglobin variant was present. The sample
that the Tosoh G7 HPLC method identified as containing an Hb variant
was recognized as Hb G-Philadelphia/Hb AS (Table 1).
Samples from 13 of the 17 patients with suspected hemoglobino-
pathies were available for molecular testing. Sequencing identified
hemoglobinopathies in all 13 (Table 1). Two of the 17 patients with
suspected hemoglobinopathies were lost to follow-up, but for another 2
of the 17 samples, the original capillary electropherograms matched the
capillary electropherogram of one of the confirmed hemoglobino-
pathies for a total of 15 patients with confirmed or presumptively-
identified hemoglobinopathies. Among the 15 patients, 10 distinct
molecular variants were identified (Table 1).
3.1. Resolution of hemoglobins by CE and HPLC measurement procedures
The hemoglobinopathies that were detected by CE were incon-
sistently recognized by other methods. CE electropherograms and the
Tosoh G8 HPLC elution patterns are shown in Figs. 2–4: Fig. 2 shows
results for a sample without a Hb variant, and Figs. 3 and 4 show results
for samples with Hb variants. Biorad Variant II Turbo HPLC chroma-
tograms are shown in Fig. 5. For samples without Hb variants, CE
achieved better baseline resolution of the hemoglobin peaks (Fig. 2A)
than was seen with the HPLC methods (e.g., Fig. 2B). Samples that
contained Hbs La Desirade and Roanne were also tested by a gel-elec-
trophoresis method for detection of hemoglobinopathies; it failed to
identify their presence (not shown).
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Clinica Chimica Acta 476 (2018) 67–74
The CE electropherograms for samples from 2 patients with Hb La
Desirade (Fig. 3A and C) showed peak doublets at Hb A0, Hb A1c and
“Other Hb A” (arrows). The patterns were identified by the analyzer's
software as atypical and no results were reported for Hb A1c. On HPLC
analyses (Tosoh G8), extra peaks near the Hb A0 and Hb A1c peaks were
seen in the sample from patient #1 (Fig. 3B; arrows), but the sample
from patient #2 had no HPLC doublet for the Hb A0 peak and only a
small doublet for the Hb A1c peak (Fig. 3D, arrow). The Tosoh G8 (and
the Variant II Turbo) gave no indication that this was an atypical
chromatogram, and the %Hb A1c was calculated.
The CE profile for Hb Roanne showed a variant peak eluting in the
same location as Hb F (Fig. 4A; arrow). No extra peak was seen in the
Tosoh G8 chromatogram (Fig. 4B) nor the Variant II Turbo chromato-
gram, and this sample would have been reported as having a normal
profile.
Hb Fannin-Lubbock I showed an atypical profile on CE analysis with
a variant peak eluting between the Hb A1c and Hb A0 peaks (Fig. 4C;
arrow); this peak hampered the software's ability to calculate Hb A1c.
Neither the Tosoh G8 HPLC nor the Variant II Turbo chromatograms
revealed the presence of a hemoglobin variant (Fig. 4D).
Blood containing Hb N-Baltimore/Hb AS was not identified as
having a variant by the Tosoh G8, but was identified as non-reportable
by the Variant II Turbo (Table 1).
In summary, among samples from the studied patient population in
which a variant had been detected by the CE method, the 3 con-
temporary HPLC measurement procedures for Hb A1c detected the
presence of hemoglobin variants in only 2 of 17 chromatograms (1 of
11 for Tosoh G7, 1 of 3 for Tosoh G8 and 0 of 3 for Variant II Turbo).
S.W. Strickland et al.
Fig. 4. Hb A1c analysis on samples from patients with confirmed Roanne and Hb Fannin-Lubbock I. A: Hb Roanne, Sebia Capillarys electropherogram; B: Hb Roanne sample, Tosoh G8
chromatogram; C: Hb Fannin-Lubbock I, Sebia Capillarys electropherogram; D: Hb Fannin-Lubbock I, Tosoh G8 chromatogram. Arrows in panels A and C indicate peaks not seen in
samples from patient lacking hemoglobin variants.
The boronate method does not recognize the presence of hemoglobi-
nopathies since separation of glycated Hb is based on the structure of
the glucose bound to hemoglobin and not based on the properties of the
hemoglobin itself.
Other published reports are available on HPLC results for 3 of the
hemoglobin variants [20,22]: Hbs J-Baltimore, G-Philadelphia/Hb AS,
and N-Baltimore/Hb AS. The published results are shown in par-
entheses in Table 1 with citation of the published reports. All 3 have
been characterized by the Variant II Turbo which detected the presence
of all 3, and 2 were studied by the Tosoh G8 which detected 1 of the 2.
Capillary electropherograms for additional hemoglobin variants
found in this study are shown in Fig. 6 and were also analyzed in a
second laboratory giving the same electropherogram results.
3.2. Quantitative results reported for Hb A1c% by CE and compared
methods
The Hb A1c results for samples that contained Hb variants varied
among methods. Quantitative Hb A1c results for samples containing Hb
Roanne and Hb Fannin-Lubbock I, are presented in Table 2 along with
results for a sample without a variant hemoglobin. Results for the
sample without an identified Hb variant differed by a maximum of
0.3% among the four methods. For these hemoglobinopathy samples
(and for a third Hb, Hb La Desirade), all CE electropherograms were
identified by the CE procedure's software as “Atypical Profile” and, in
some cases, the software provided no Hb A1c result. By contrast, the
HPLC instruments flagged none of the HPLC chromatograms as atypical
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Clinica Chimica Acta 476 (2018) 67–74
and provided Hb A1c results for the samples, including the samples from
2 patients with Hb La Desirade (shown in Table 1).
For the sample that contained Hb Roanne, the CE result was higher
by 0.8% Hb A1c than the results of the ultra2 (Table 2). The Hb A1c
result for the sample containing Hb Fannin-Lubbock I was flagged by
the CE analyzer, but the quantitative result that it calculated was 1.4%
Hb A1c higher than results from the ultra2 (See Discussion). Results also
differed among the G8, ultra2, and Variant II Turbo when a hemoglobin
variant was present (Table 2).
4. Discussion
According to the WHO, approximately 7% of the worldwide popu-
lation is affected by a hemoglobin variant [23]. Many of these variants
are clinically silent and are detected only fortuitously. Identification of
variants is critical for interpretation of Hb A1c results for several rea-
sons: First, Hb variants may produce altered red cell survival [2], thus
making even analytically accurate Hb A1c results clinically misleading.
Second, unresolved peaks can contribute to falsely increased or de-
creased Hb A1c results in both HPLC and CE methodologies: Peaks that
overlap with the Hb A0 peak or with the Hb A1c peak prevent proper
integration of the peaks of interest and compromise calculations of the
peak area, thereby affecting the reported Hb A1c%. Third, hemoglobin
variants may affect hemoglobin glycation [24], further increasing the
need for identification of the presence of these variants.
This study shows that the presence of Hb variants is not reliably
detected by contemporary ion-exchange HPLC-based (or boronate-
S.W. Strickland et al.
Fig. 5. Biorad Variant II HPLC Chromatogram with Hb La Desirade and Hb Fannin Lubbock. A: Expected pattern of Biorad Variant II chromatogram. B: Hb Fannin-Lubbokc sample Biorad
Variant II chromatogram; C: Hb La Desirade sample Biorad Variant II chromatogram.
based) methods for measuring Hb A1c. The analytical resolution of CE
allowed identification of 10 patients who had hemoglobin variants that
had gone undetected by an HPLC method, and Hb variants were not
reliably detected by 2 additional HPLC-based measurement procedures
used during this study. The HPLC methods were unable to efficiently
resolve the Hb variants; and, despite potential interference from
shoulder peaks in some cases (e.g., Fig. 3), the HPLC measurement
procedures nonetheless reported results for Hb A1c. The HPLC chro-
matograms of other Hb variants (e.g., Hb Roanne and Hb Fannin-Lub-
bock I) showed no sign of the variant, most likely due to lack of re-
solution of these Hbs from Hb A0.
The quantitative results of the various methods deserve further at-
tention. The Hb A1c result in the CE method derives from the following
ratio: Hb A1c peak/(Hb A1c peak + Hb A0 peak). In the Fannin-Lubbock
I patient sample, this calculation resulted in a reported Hb A1c% of
9.3%, whereas results of the other methods ranged from 7.1–7.9%Hb
A1c (Table 2). However, if the CE calculation had included the aberrant
peak for Hb Fannin-Lubbock I (which was resolved from the Hb A0
peak as indicated by the arrow in Fig. 4C) as part of the Hb A0 peak, the
Hb A1c% result would have been 7.2%, a result consistent with the
other methods. This could suggest that the Hb Fannin-Lubbock I peak
on the HPLC chromatogram is hidden within the large A0 peak
(Fig. 4D) and is contributing to a decreased Hb A1c% by increasing the
area of the “A0” peak. Moreover, it suggests that neither method se-
parated Hb A1c from a presumed Hb Fannin-Lubbock I Hb that is gly-
cated on its N-terminal valine. A similar situation was observed for Hb
Roanne, which showed a slightly lower Hb A1c% for the HPLC-based
method than for the CE method. Another possible explanation of the
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Clinica Chimica Acta 476 (2018) 67–74
data for these 2 hemoglobins is that the calculation of Hb A1c% by the
CE (Sebia) software included glycated Hb variant (not visible on the
electropherogram; presumably integrated with the Hb A1c peak) with
the Hb A1c while the software fully resolved the nonglycated Hb variant
as part of the Hb A0, thus the result was an overestimate of Hb A1c% by
the CE method.
This study has several strengths, notably that each of the 10 rare
hemoglobin variants was definitively identified by sequencing. By
analyzing samples both by CE and by representative comparison
methods, the study revealed that CE recognized the presence of he-
moglobinopathies that the other studied methods did not recognize as
present. A limitation of the study is that it did not prospectively com-
pare the frequencies of identification of hemoglobin variants by CE and
by other methods in a series of patients that were all tested by both
methods to ascertain whether CE is more sensitive than other methods
in detection of hemoglobin variants. The observed frequency of detec-
tion of rare hemoglobinopathies, however, appears to be near the ex-
pected prevalence of rare hemoglobinopathies in this population. As
reported by Huisman in a study of hemoglobinopathies in the
Southeastern United States, the prevalence of rare hemoglobin variants
with known phenotypes was 0.012% [26], similar to the prevalence of
0.06% in our study population [14] in the same part of the U.S. Al-
though this difference can be expected to occur by chance, a possible
explanation for the higher prevalence in the present study than in the
previous study [14] is that the high resolution of CE allowed identifi-
cation of hemoglobin variants that were not recognized as being present
by the screening gel-electrophoresis method used in the earlier (1980)
study. This is supported by our observation that two Hb variants that
S.W. Strickland et al. Clinica Chimica Acta 476 (2018) 67–74

Fig. 6. Rare Hemoglobin variants identified by CE A: Capillary electropherogram of sample with Hb Nouakchott. Arrows represent extra peaks present in the patient sample. B: Capillary
electropherogram of sample with Hb A2 prime. Arrows represent aberrant peaks present in the patient sample. Representative of either a homozygous A2 prime mutation or a patient that
is heterozygous for A2 prime mutation and delta thalassemia. C: Capillary electropherogram of sample with Hb G Philadelphia/AS. Arrows represent extra peaks present in the patient
sample [18]. D: Capillary electropherogram of sample with Hb N-Baltimore/Hb AS. Arrows represent aberrant peaks present in the patient sample. Peak 1: Hb-N Baltimore, Peak 2: other
HbA, Peak 3: unknown, Peak 4: Hb AS, Peak 5: putative A2.

Table 2 confirmed hemoglobinopathy patients and for 1 presumptively identi-

Hb A1c concentrations reported by 4 measurement procedures. fied patient. Although 4 of these patients were anemic, the anemia was
mild (Hb concentrations of 9.2–11.0 g/dL) and was not being treated.
%Hb A1c
One of these variants, Hb La Desirade, is usually discovered only as an
CE Ion-exchange HPLC Boronate HPLC incidental finding and was not revealed by the Tosoh G7, the Variant II
Turbo, or the boronate-affinity methods for measurement of Hb A1c,
CapillaryS2 G8 Variant II Turbo Ultra2 making it likely that more individuals harbor the variant than are re-
Hb A 6.6 6.8 6.9 6.6 cognized clinically. Anemias that are associated with hemoglobino-
Hb Roanne 6.6a (5.3b) 5.8 5.9 5.8 pathies may reflect shortened red cell survival, which can decrease Hb
Hb Fannin-Lubbock I 9.3a (7.2b) 7.1 7.9 7.9 A1c% by shortening the time the RBCs are exposed to circulating glu-
a
cose. In such cases the reported Hb A1c% could be clinically misleading
No result would be reportable because of non-Hb A peak > 15% (Hb Roanne) or
if interpreted without knowledge of the presence of the variant.
lack of baseline resolution from Hb A (Hb Fannin-Lubbock I).
b
Results in parentheses: for comparison with HPLC results, the variant peak was in-
cluded with the A0 peak in the calculation of “Hb A1c%”. 5. Conclusions

were resolved by the CE measurement procedure were not detected by a
gel-electrophoresis-based method for hemoglobinopathy detection.
Moreover, the most common of the rare hemoglobinopathies that were
identified in the earlier study were Hb A2 prime, G-Philadelphia and N-
Baltimore, all of which were also identified in our study in the South-
eastern United States. These observations suggest that measurement of
Hb A1c by the CE method used here identifies a high proportion of the
patients with the rare hemoglobinopathies that are encountered in the
geographic area studied. Thus, other methods, if tested in parallel with
the CE method, would be unlikely to identify appreciably more patients
who have variant Hbs than were found by the CE method.
Rare hemoglobinopathies are unlikely to be diagnosed clinically. In
the present study, the presence of a hemoglobin variant was unknown
in all patients prior to testing, despite the fact that most had a long
association with medical facilities. Three of the hemoglobin variants
identified by CE in the present study — Hbs Roanne, La Desirade, and
Nouakchott — are typically silent clinically, and are associated only
with mild anemia [27–29]. These 3 variants accounted for 5 of our
Rare Hb variants occur with considerable frequency and their de-
tection by contemporary HPLC measurement procedures for Hb A1c is
unreliable. These variants can interfere analytically in measurements of
Hb A1c and may produce misleading results if associated with decreased
red cell survival or decreased rates of glycation. The resolving power of
the studied CE measurement procedure for quantification of Hb A1c
appears sufficient to recognize the presence of a range of rare he-
moglobinopathies in addition to the more common Hbs C, D, E, F, and
S.
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