Evaluating The Value Of Flow Cytometry For The Differential Diagnosis Between Refractory Cytopenia With Multiple Dysplasia And Aplastic Anemia

Objective:①evaluating the value of flow cytometry(FCM) for the differential diagnosis between myelodysplasia(MDS) subtype refractory cytopenia with multiple dysplasia(RCMD) and aplastic anemia(AA),②proposing the best antibodies combination using in the diagnosis of MDS and the differential diagnosis between RCMD and AA,③indicating the attention shoud be payed in the analysis of flow cytometric results.Methods:We analysed the flow cytometric datas of bone marrow samples from 168 cases of RCMD,in which 39 cases were hypoplastic MDS(hypo-RCMD),and 77 cases of AA, retrospectively and in blind,then we compared the flow cytometric results with comprehensive diagnosis(gold standard) to evaluate the diagnosis values.Results:①In the evaluation of single immunophenotypes,in RCMD and AA group,the specificity of the immunophenotypes in the surface of granulocytes and myeloblasts was high (range 75%-100%),but the sensitivity was very low(range 5.4%-32.1%),the sensitivity of CD34+ cells≥1%and myeloblasts≥3%was also low,which were 50%and 42.3%, respectively;in hypo-RCMD and AA group,the results were similar to those in RCMD and AA group.②In parallel tests,in RCMD and AA group,the sensitivity and specificity of the combination of CD34+ cells≥1%,myeloblasts≥3%,loss of CD13 in myeloblasts and abnormal expression of CD117 in granulocytes were higher than other combinations,which were 63.7%and 92.2%,respectively;in hypo-RCMD and AA group,the sensitivity and specificity of the combination of CD34+ cells≥1%,myeloblasts≥3%,increased intensity of CD33 in granulocytes and abnormal expression of CD117 in granulocytes were higher than other combinations,which were 61.5%and 92.2%,respectively.③In the scoring method, scoring with eight markers,which were CD34+ cells≥1%,myeloblasts≥3%,abnormal expression of CD117 in granulocytes,loss of CD13 in myeloblasts,increased intensity of CD33 in granulocytes,loss of CD13 in granulocytes,loss of CD10 in granulocytes,and decreased SSC in granulocytes.The sensitivity and specificity were both high if we defined that the score equal or higher than 1.5 was RCMD and the score lower than 1.5 was AA,in RCMD and AA group which were 64.9%and 93.5%,respectively;in hypo-RCMD and AA group which were 61.5%and 92.2%,respetively.④The best antibodies combination using for the differential diagnosis between RCMD and AA or other pancytopenia diseases was IgG1-FITC/IgG1-PE/IgG1-PC5/IgG1-ECD,HLA-DR/CD117/CD34/CD45,CD56/CD33/CD6 4/CD45,CD7/CD10/CD19/CD45,CD16/CD11b/CD13/CD45,CD4/CD8/CD3/CD45,cIgG1-FI TC/cIgG1-PE/cIgG1-PC5/cIgG1-ECD,cCD79a/cMPO/cCD3/CD45.⑤Pay attention to the following when analysing immunophenotypes with flow cytometry:1) the pattern of fluorescence distributions to distinguish positive from negtive population;2) when evaluating the changes in the fluorescence intensity of antigen expression,comparing with normal cells; 3) calculating the numbers of CD34+ cells through CD34 versus CD45 pattern for well;4) The numbers of myeloblasts in bone marrow are few,so it must be very earnest and elaborative when analysing the immunophenotypes in the surface of myeloblasts.Conclusions:①The value of single immunophenotypes for differential diagnosis between RCMD and AA is low,the specificity is high,but the sensitivity is low.②Comparing with single immunophenotypes,parallel tests can elevate the diagnostic sensitivity obviously and not greatly degrade the specificity.When combining CD34+ cells≥1%,myeloblasts≥3% and abnormal expression of CD117 in granulocytes those three markers with other markers, the sensitivity and specificity are both well,so the three markers are important.③We suggest to distinguish RCMD from AA with scoring method which is preciser.④The new antibodies combination we proposed is used for the differential diagnosis between RCMD and AA or other pancytopenia diseases,but it should be evaluated in the further prospective researchs.