USEFULNESS OF TWO NEW METHODS,-FLOW CYTROMETRY (FC) AND CRYOHEMOLYSIS (CH) TESTS-FOR THE DIAGNOSIS OF HEREDITARY SPHEROCYTOSIS (HS): COMPARISION WITH STANDARD SCREENING TESTS.

CRISP, R; DONATO, H; VENEGAS, B; CHAMORRO ME; SOLARI, L; VOTA, D; GAMELLA, D; MIGUEZ, G; SCHVARTZMAN, G; VITTORI, D; CALDAROLA, S; ALFONSO, G; NESSE, A

Copenhagen

Congreso; 13th Congress of the European Hematology Association; 2008

European Hematology Association

<!-- /* Style Definitions */ p.MsoNormal, li.MsoNormal, div.MsoNormal {mso-style-parent:""; margin:0cm; margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:12.0pt; font-family:"Times New Roman"; mso-fareast-font-family:"Times New Roman";} @page Section1 {size:595.3pt 841.9pt; margin:70.85pt 3.0cm 70.85pt 3.0cm; mso-header-margin:35.4pt; mso-footer-margin:35.4pt; mso-paper-source:0;} div.Section1 {page:Section1;} --> 0622 USEFULNESS OF TWO NEW METHODS - FLOW CYTOMETRY (FC) AND CRYOHEMOLYSIS (CH) TESTS - FOR THE DIAGNOSIS OF HEREDITARY SPHEROCYTOSIS (HS): COMPARISON WITH STANDARD SCREENING TESTS L. Crisp,1 C. Donato,2 B. Venegas,1 E. Chamorro,3 A. Solari,1 D. Vota,3 D. Gammela,1 G. Miguez,1 A. Schvartzman,2 D. Vittori,3 S. Caldarola,1 G. Alfonso,1 A. Nesse3 1Hospital Nacional A.Posadas, BUENOS AIRES; 2Consultorios de Hematología Infantil, BUENOS AIRES; 3Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, BUENOS AIRES, Argentina Background. CH and FC tests are recently developed methods for the diagnosis of HS. Their usefulness seems to be better than standard screening methods. No experience concerning these tests has been reported in our country. Furthermore, the prevalence of each protein deficiency underlying HS in argentine patients is unknown. Aims. a) To compare usefulness of CH and FC with standard tests; b) To establish the prevalence of each protein deficiency in Argentina. Methods. Nineteen patients (aged 3 months to 77 years) previously diagnosed with HS by standard tests, 20 asymptomatic family members, and 106 normal controls have been included so far. Patients, family members, and controls were studied by complete blood cells count, blood smear, osmotic fragility (OF), autohemolysis (AH), CH, eosin-5’-maleimide FC, and denaturing electrophoresis (SDS-PAGE); OF, AH, and SDS-PAGE were not performed in 72 normal controls. For FC, the reduction of the mean channel fluorescence (MCF) reading and the increase of the coefficient of variation (CV) were obtained using mean values for simultaneously performed normal controls. The cut-off values for CH and FC were determined through the analysis of receiver operating characteristics (ROC) curves. For FC and SDS-PAGE, each assay was set up together with 6 normal controls. Results. In patients, percentages of positive tests were: OF 79%, AH 59%, CH 78%, FC 89%, and SDS-PAGE 58%. Only 6 (32%) were positive for all the tests. Considering only CH and FC, at least one of them was positive in 100% of patients. For CH, the mean normal value was 1.47±0.56%, and the established cut-off value was 2.8%; mean value in patients was 8.55±8.45%. The CH showed sensitivity of 78% and specificity of 96%. Results of FC were evaluated either through MCF decrease or CV increase: the established cut-off values were 19% for MCF (sensitivity 74%, specificity 97.5%), and 12% for CV (sensitivity 74%, specificity 97.5%). No correlation was demonstrated between MCF and CV changes. In 2 patients, test was positive only through the CV change. In 6 cases (5 of them splenectomized), MCF changes were greater than CV changes. A cut-off value of 25% for each parameter determined a specificity of 100%. FC was positive in 7 of 8 patients without demonstrable protein deficiency (with CH positive in 3); in the remaining one, only CH was positive. SDS-PAGE detected the following deficiencies: spectrin (6), ankyrin (5), protein 4.1 (2), protein 4.2 (1). Seven patients showed a single deficiency whereas 4 showed 2 deficiencies. Band 3 deficiency was demonstrated in one asymptomatic family member in whom CH was the only positive test. Conclusions.CH and FC showed high specificity and sensitivity for diagnosis, similar to OF and higher than SDS-PAGE. Main advantages of CH and FC over standard tests are smaller amount of sample required and shorter performing time; moreover, CH is a very simple test not requiring special equipment. The simultaneous readings of MCF and CV improve the diagnostic value of FC compared with CH. The observed predominant spectrin deficiency agrees with reports from other Latin American countries.