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Annals of Clinical & Laboratory Science 32:37-43 (2002)
© 2002 Association of Clinical Scientists

Rapid, Sensitive Diagnosis of Hemolytic Anemia Using Antihemoglobin Antibody in Hypotonic Solution

Wonbae Lee1, Yonggoo Kim1, Jihyang Lim1, Myungshin Kim1, Eun Jung Lee1, Ahwon Lee1, Kyo Young Lee1, Chang Suk Kang1, So-Young Kim1, Kyungja Han1 and Soo Hwan Pai2
1 Departments of Clinical Pathology and Pediatrics, Catholic University Medical College, Seoul, Korea
2 Department of Clinical Pathology, College of Medicine, Inha University, Inchon, Korea

Address correspondence to Kyungja Han, M.D., Department of Clinical Pathology, Catholic University Medical College, St. Mary’s Hospital, Youngdeungpo-gu, Youido-dong 62, Seoul, South Korea 150-713; tel 82 2 3779 1297; fax 82 2 783 6648; e-mail hankja{at}cmc.cuk.ac.kr.

We have developed a new and simple flow cytometric method to detect damaged red blood cells (RBCs) using anti-Hb in hypotonic solution. We studied a total of 200 patients, including 62 patients with schistocytosis, 8 postsplenectomy patients, and 108 healthy controls. Peripheral blood (2 µl) was stained with phycoerythrin-conjugated (PE) antihemoglobin antibody (anti-Hb) in 0.6% (w/v) NaCl solution, and analyzed by flow cytometry omitting the washing step. The proportion of RBCs stained by anti-Hb was 0.55% (SD ±0.23%) in normal controls and was significantly higher in patients with schistocytosis (2.95±2.95%, p <0.001). Six of 108 blood samples from normal controls and 60 of 62 samples from schistocytosis patients showed >=1.01% stained RBCs (ie, values > mean+2SD of normal controls). The number of schistocytes counted by microscopic examination correlated with the proportion of RBCs stained by anti-Hb (r = 0.637, p <0.001). The proportions of stained RBCs in blood samples with malaria, spherocytosis, and elliptocytosis were also significantly higher than in normal controls. However, the results in postsplenectomy and iron-deficiency anemia (IDA) patients were not significantly different from the normal controls; the number of schistocytes in postsplenectomy patients was not related to the proportion of RBCs stained by anti-Hb. Based on these findings, flow cytometry of damaged RBCs using anti-Hb in hypotonic solution is a simple, sensitive, and accurate method to detect active hemolysis.

Keywords: hemolytic anemia, antihemoglobin antibody, flow cytometry




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Home page
 Annals of Clinical & Laboratory ScienceHome page
K. Han, Y. Kim, S.-Y. Chung, M. Kim, and J. Lim
How to Establish the Cut-off Values when Estimating Fragmented RBCs by Flow Cytometry
Ann. Clin. Lab. Sci., October 1, 2002; 32(4): 404 - 405.
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