Comparison of High-Performance Liquid Chromatography and Capillary Electrophoresis in the Screening of Haemoglobinopathies
DOI:
https://doi.org/10.51253/pafmj.v74i2.7452Keywords:
Beta-thalassaemia, Capillary electrophoresis, Haemoglobinopathies, High-Performance liquid chromatographyAbstract
Objective: To compare the results of High-Performance Liquid Chromatography and Capillary Electrophoresis in the
screening of haemoglobinopathies.
Study Design: Cross-sectional study.
Place and Duration of Study: Department of Haematology, Armed Forces Institute of Pathology, Rawalpindi Pakistan, from May 2020 to Nov 2020.
Methodology: Ninety(90) newly diagnosed patients with haemoglobinopathies were included. Patients with a history of recent blood transfusion (<4 weeks) were excluded. Venous blood samples were taken in EDTA tubes, separated into two aliquots and evaluated in parallel on High-Performance Liquid Chromatography and Capillary Electrophoresis analyzers.
Results: Nine different haemoglobinopathies were encountered in the study sample. The most common condition was beta thalassaemia trait found in 50(55.6%) subjects, while Haemoglobin (Hb) (D) homozygous was the least common, detected in only 1(1.1%) subjects. A significant difference was observed between values in cases of Hb(D) Iran trait; Mean Hb(A2) percentage detected by HPLC was 41.96±1.46% compared to 1.5±0.79% detected by CE (p-value<0.001). CE detected a mean of 41.4±1.01% of Hb(D)Iran in the patients of the Hb(D) Iran trait, while High-Performance Liquid Chromatography did not detect this Hb variant (p-value<0.001). A significant difference was also found in cases of Hb(E) trait in percentages of Hb(A2) and Hb(E) variants. High-Performance Liquid Chromatography detected a mean of 31.43±2.84% of Hb(A2) compared to 4.5±0.5% detected by CE (p-value=0.003).
Conclusion: Capillary electrophoresis is capable of identifying haemoglobin variants Hb(D) Iran and Hb(E) in cases of Hb(D) Iran trait and Hb(E) trait/Hb(E) homozygous, which HPLC does not pick up.
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Nisha TR, Bindu CS. HPLC based evaluation of
Haemoglobinopathies in a tertiary care setting in North Kerala. J
Med Sci Clin Res 2018; 6(1): 32027-32032.
https://doi.org/10.18535/jmscr/v6i1.102.
Shabbir S, Nadeem M, Sattar A, Ara I, Ansari S, Farzana T, et al.
Type and frequency of haemoglobinopathies, diagnosed in the
area of Karachi, in Pakistan, Cogent Med 2016; 3(1): 1-6.
https://doi.org/10.1080/2331205X.2016.1188875.
Piel FB. The present and future global burden of the inherited
disorders of hemoglobin. Hematol Oncol Clin North Am 2016;
(2): 327-341. https://doi.org/10.1016/j.hoc.2015.11.004.
Ghafoor M, Sabar MF, Sabir F. Prevention programmes and
prenatal diagnosis for beta thalassemia in Pakistan: A narrative
review. J Pak Med Assoc 2021; 71(1): 326-331.
https://doi.org/10.47391/jpma.665.
Karadag ME, Akbulut ED, Avci E, Oguz EF, Kader S, Abuşoğlu
G. et al. Evaluation of four different HPLC devices for
hemoglobinopathy screening. Turk J Biochem 2021; 46(1): 39–44.
Canatan D, Delibas S. Report on ten years’ experience of
premarital hemoglobinopathy screening at a center in Antalya,
southern Turkey. Hemoglobin 2016; 40(4): 273-276.
https://doi.org/10.3109/03630269.2016.1170030.
Chakravarthy SN, Ramanathan RS, Smitha S, Valappil V,
Durairaj M. Hide and seek: Efficacy of HPLC and capillary zone
electrophoresis as screening tools for hemoglobin disorders. Int J
Path Lab 2016; 2(1): 6.
Frömmel C. Newborn Screening for Sickle Cell Disease and
Other Hemoglobinopathies: A Short Review on Classical
Laboratory Methods-Isoelectric Focusing, HPLC, and Capillary
Electrophoresis. Int J Neonatal Screen 2018; 4(4): 39.
https://doi.org/10.3390/ijns4040039.
Greene DN, Pyle AL, Chang JS, Hoke C, Lorey T. Comparison of
SebiaCapillarys Flex capillary electrophoresis with the Bio Rad
Variant II high pressure liquid chromatography in the
evaluation of haemoglobinopathies. Clin Chim Acta 2012;
(15-16): 1232-1238. https://doi.org/10.1016/j.cca.2012.03.027.
Sangkitporn S, Sangkitporn SK, Tanjatham S, Suwannakan B,
Rithapirom S, Yodtup Y. et al. Multicenter validation of fully
automated capillary electrophoresis method for diagnosis of
thalassemias and haemoglobinopathies in Thailand. Southeast
Asian J Trop Med Public Health 2011; 42(4): 1224-1232.
Cotton F, Changying L, Fontaine B. Evaluation of a capillary
electrophoresis method for routine determination of
hemoglobins A2 and F. Clin Chem 1999; 45(2): 237-243.
Bakshi NA, Gulbranson R, Garstka D. Serum free light chain
(FLC) measurement can aid capillary zone electrophoresis in
detecting subtle FLC-producing M proteins. Am J Clin Pathol
; 124(2): 214-218.
https://doi.org/10.1309/xe3u-dark-w1b9-emwm.
Bossuyt X, Lissoir B, Marien G. Automated serum protein
electrophoresis by Capillarys. Clin Chem Lab Med 2003; 41(5):
-710. https://doi.org/10.1515/cclm.2003.107.
Keren DF, Hedstrom D, Gulbransan R, Ou CN, Bak R.
Comparison of sebiacapillarys capillary electrophoresis with the
primus high-pressure liquid chromatography in the evaluation
of haemoglobinopathies. Am J Clin Pathol 2008; 130(5): 824-31.
https://doi.org/10.1309/ajcpqy80hzwhhgzf.
Mohanty PK, Meher S, Dehury S, Bhattacharya S, Das K, Patel S,
et al. Compound heterozygote of Hb D Iran [HBB: c. 67G> C, β
(B4) Glu> Gln] with β0-thalassemia [cds 41/42 (-CTTT)] from
Eastern India Hematol Transfus Cell Ther 2018; 40(1): 82-85.
https://doi.org/10.1016%2Fj.bjhh.2017.09.001.
Chopra P, Bhardwaj S, Negi P, Arora A. Comparison of Two
High-Pressure Liquid Chromatography Instruments Bio-Rad
Variant-II and Tosoh HLC-723G11 in the Evaluation of
Haemoglobinopathies. Indian J Hematol Blood Transfus 2020;
(4): 725-732.
https://doi.org/10.1007/s12288-020-01298-5.
Olivieri NF. The beta-thalassemias. N Engl J Med 1999; 341(2):
-109. https://doi.org/10.1056/nejm199907083410207.
Chhotray GP, Dash BP, Ranjit M. Spectrum of haemoglobinopathies in Orissa, India. Hemoglobin 2004; 28(2): 117-122.
https://doi.org/10.1081/hem-120034244.
Daniel YA, Turner C, Haynes RM. Quantification of hemoglobin
A2 by tandem mass spectrometry. Clin Chem 2007; 53(8): 1448-
https://doi.org/10.1373/clinchem.2007.088682.
Paleari R, Ceriotti F, Harteveld CL, Strollo M, Bakker-Verweij G,
Ter Huurne J, et al. Calibration by commutable control materials
is able to reduce inter-method differences of current highperformance methods for HbA2. Clin Chim Acta 2018; 477: 60-
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