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COMPARISON STUDY OF THE EOSIN-5'-MALEIMIDE BINDING TEST, OSMOTIC FRAGILITY TEST ANDCRYOHEMOLYSIS TEST IN THE DIAGNOSIS OF HEREDITARY SPHEROCYTOSIS
Author(s):
Esin Özcan
,
Esin Özcan
Affiliations:
Pediatric Hematology,Dr. Behcet Uz Children's Hospital,Izmir,Turkey
Yesim Oymak
,
Yesim Oymak
Affiliations:
Pediatric Hematology,Dr. Behcet Uz Children's Hospital,Izmir,Turkey
Tuba Hilkay Karapınar
,
Tuba Hilkay Karapınar
Affiliations:
Pediatric Hematology,Dr. behcet Uz Children's Hospital,Izmir,Turkey
Salih Gözmen
,
Salih Gözmen
Affiliations:
Pediatric Hematology,Dr. Behcet Uz Children's Hospital,Izmir,Turkey
Sultan Aydın Köker
,
Sultan Aydın Köker
Affiliations:
Pediatric Hematology,Dr. Behcet Uz Children's Hospital,Izmir,Turkey
Neryal Muminoğlu
,
Neryal Muminoğlu
Affiliations:
Pediatric Hematolgy,Dr. Behcet Uz Children's Hospital,Izmir,Turkey
Sultan Okur
,
Sultan Okur
Affiliations:
Pediatric Hematogy,Dr. Behcet Uz Children's Hospital,Izmir,Turkey
Raziye Canan Vergin
Raziye Canan Vergin
Affiliations:
Pediatric Hematology,Dr. Behcet Uz Children's Hospital,Izmir,Turkey
(Abstract release date: 05/18/17) EHA Library. Özcan E. 05/18/17; 182556; PB1842
Esin Özcan
Esin Özcan
Contributions
PDF Abstract
Abstract

Abstract: PB1842

Type: Publication Only

Background
The primary lesion in HS is loss of membran surface area due to defects of the membran protein. Cryohemolysis test and osmotic fragility (OF) test are used for screening. However no test for HS is 100% reliable. The eosin-5'-maleimide (EMA) binding test based on flow cytometry. Eighty percent of the fluorescent-labelled EMA binds to band 3 protein which is lost in HS due to protein 4.1, spectrin and ankyrin deficiency. Thus measurement of the flourescent EMA test detects all the different forms of HS.

Aims
In this study we aimed to evaluate the concordence of EMA binding test with other diagnostic parameters for HS.

Methods

The patients with HS were diagnosed according to clinical findings for hemolytic anemia, splenomegaly and spherocytes in peripheral blood. Hemogram, reticulocyte count, total/direct bilirubin, spherocytes in blood smear (BS), EMA binding test, OF test, and cryohemolysis test were obtained from pateints and control groups. Correlation between EMA, OF and cryohemolysis tests were evaluted.

Results

Twenty-five male, 17 female HS patients aged between 1.0-19.0 years and 38 male, 47 female healthy controls were evaluated. There were no differences between both groups in terms of age and sex (Table 1). The median (range) values of hemoglobin (%), reticulocyte count (%), mean corpuscular volume (fl), MCHC (%) and total bilirubin level were seen in table 1. Besides MCV values there were differences between groups in terms of these parameters (Table 1). The median MCF of HS patients was significantly lower than that of healthy controls while cryohemolysis and osmotic fragility were higher in HS patients than healthy controls (Table1). There were moderate concordence between cryohemolysis and EMA test (r=-0.355, p<0.001). The sensitivity of EMA was 92.86%, specifity was 82.35%, PPV was %72.22, NPV was %95.89. EMA was superior diagnostic test to osmotic fragility. (sensitivity; %83.33, specifity;%76.47, PPV;%63.64 and NPV;%90.28). The sensitivity of cryohemolysis test was 90.48%, specificity was 94.12%, PPV was %88.37, NPV was %95.24.
Table 1: Comparison of Clinicaland Laboratory Findings in Hereditary Spherocytosis groups and Healthy Controls
Hereditary Spherocytosis
(n =42)
Healthy Controls
(n = 85)
P Value
Sex, M/F
25/17
38/47
>0.05
Age, median (range), year
8.5 (1.0-19.0)
12.0 (1.0-20.0)
0.416
Hemoglobin, median (range), g/dL
10.8 (5.8-14.7)
14.2 (11-17.2)
< 0.001
Reticulocyte, median (%)
6.1 (1.6-38.0)
1.1 (0.1-4.1)
<0.001
MCV, median (range),fL
77.5(73-90)
82(77-94)
0.994
MCHC, median (range), g/dL
33.0(27.0-37.0)
32.0 (27.0-35.0)
< 0.001
Total bilirubin,
median (range), mg/dL
3.3 (1.0-16.0)
0.9 (0.2-2.9)
< 0.001
EMA (%), median (range)
18.38 (8-48)
26.43 (12-45)
< 0.001
Cryohemolysis (%),
median (range)
37.0 (2.0-83.0)
3.8 (2.0-14.0)
< 0.001
OF (%)
62.2
37.8
< 0.001
M/F; Male/Female, FC EMA;flowcytometric eosin-5'-maleimide, OF;osmotic fragility, HS; hereditary spherocytosis, MCHC; mean corpuscular hemoglobin concentration, MCV; mean corpuscular volume, BS; bloodsmear

Conclusion

In this study EMA-FC was more sensivitive and specificity than osmotic fragility. However specificity and PPV of cryohemolysis was higher than other test. Also we showed moderate concordance cryohemoysis and EMA test.
Although high sensitivity and specifity of EMA test there were need to use other tests together with family history of patient, physical examination, evaluation of blood smear and several tests for HS diagnosis.

Session topic: 27. Enzymopathies, membranopathies and other anemias

Keyword(s): Hemolytic anemia, Band 3, Ankyrin, Hereditary spherocytosis

Abstract: PB1842

Type: Publication Only

Background
The primary lesion in HS is loss of membran surface area due to defects of the membran protein. Cryohemolysis test and osmotic fragility (OF) test are used for screening. However no test for HS is 100% reliable. The eosin-5'-maleimide (EMA) binding test based on flow cytometry. Eighty percent of the fluorescent-labelled EMA binds to band 3 protein which is lost in HS due to protein 4.1, spectrin and ankyrin deficiency. Thus measurement of the flourescent EMA test detects all the different forms of HS.

Aims
In this study we aimed to evaluate the concordence of EMA binding test with other diagnostic parameters for HS.

Methods

The patients with HS were diagnosed according to clinical findings for hemolytic anemia, splenomegaly and spherocytes in peripheral blood. Hemogram, reticulocyte count, total/direct bilirubin, spherocytes in blood smear (BS), EMA binding test, OF test, and cryohemolysis test were obtained from pateints and control groups. Correlation between EMA, OF and cryohemolysis tests were evaluted.

Results

Twenty-five male, 17 female HS patients aged between 1.0-19.0 years and 38 male, 47 female healthy controls were evaluated. There were no differences between both groups in terms of age and sex (Table 1). The median (range) values of hemoglobin (%), reticulocyte count (%), mean corpuscular volume (fl), MCHC (%) and total bilirubin level were seen in table 1. Besides MCV values there were differences between groups in terms of these parameters (Table 1). The median MCF of HS patients was significantly lower than that of healthy controls while cryohemolysis and osmotic fragility were higher in HS patients than healthy controls (Table1). There were moderate concordence between cryohemolysis and EMA test (r=-0.355, p<0.001). The sensitivity of EMA was 92.86%, specifity was 82.35%, PPV was %72.22, NPV was %95.89. EMA was superior diagnostic test to osmotic fragility. (sensitivity; %83.33, specifity;%76.47, PPV;%63.64 and NPV;%90.28). The sensitivity of cryohemolysis test was 90.48%, specificity was 94.12%, PPV was %88.37, NPV was %95.24.
Table 1: Comparison of Clinicaland Laboratory Findings in Hereditary Spherocytosis groups and Healthy Controls
Hereditary Spherocytosis
(n =42)
Healthy Controls
(n = 85)
P Value
Sex, M/F
25/17
38/47
>0.05
Age, median (range), year
8.5 (1.0-19.0)
12.0 (1.0-20.0)
0.416
Hemoglobin, median (range), g/dL
10.8 (5.8-14.7)
14.2 (11-17.2)
< 0.001
Reticulocyte, median (%)
6.1 (1.6-38.0)
1.1 (0.1-4.1)
<0.001
MCV, median (range),fL
77.5(73-90)
82(77-94)
0.994
MCHC, median (range), g/dL
33.0(27.0-37.0)
32.0 (27.0-35.0)
< 0.001
Total bilirubin,
median (range), mg/dL
3.3 (1.0-16.0)
0.9 (0.2-2.9)
< 0.001
EMA (%), median (range)
18.38 (8-48)
26.43 (12-45)
< 0.001
Cryohemolysis (%),
median (range)
37.0 (2.0-83.0)
3.8 (2.0-14.0)
< 0.001
OF (%)
62.2
37.8
< 0.001
M/F; Male/Female, FC EMA;flowcytometric eosin-5'-maleimide, OF;osmotic fragility, HS; hereditary spherocytosis, MCHC; mean corpuscular hemoglobin concentration, MCV; mean corpuscular volume, BS; bloodsmear

Conclusion

In this study EMA-FC was more sensivitive and specificity than osmotic fragility. However specificity and PPV of cryohemolysis was higher than other test. Also we showed moderate concordance cryohemoysis and EMA test.
Although high sensitivity and specifity of EMA test there were need to use other tests together with family history of patient, physical examination, evaluation of blood smear and several tests for HS diagnosis.

Session topic: 27. Enzymopathies, membranopathies and other anemias

Keyword(s): Hemolytic anemia, Band 3, Ankyrin, Hereditary spherocytosis

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