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Detection of t(9;22) b2a2 fusion transcript by flow cytometry
Author(s) -
Ranjbaran R.,
Okhovat M. A.,
Abbasi M.,
Moezzi L.,
Aboualizadeh F.,
Amidzadeh Z.,
Golafshan H. A.,
BehzadBehbahani A.,
Sharifzadeh S.
Publication year - 2016
Publication title -
international journal of laboratory hematology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.705
H-Index - 55
eISSN - 1751-553X
pISSN - 1751-5521
DOI - 10.1111/ijlh.12515
Subject(s) - flow cytometry , microbiology and biotechnology , chromosomal translocation , fluorescence in situ hybridization , k562 cells , biology , in situ hybridization , fluorescence microscope , jurkat cells , fluorescence , cell culture , cytoplasm , cell , messenger rna , immunology , t cell , chromosome , gene , biochemistry , immune system , physics , genetics , quantum mechanics
Summary Introduction We assessed the feasibility of flow cytometry–fluorescent in situ hybridization technique in the detection of translocated mRNA in the cytoplasm of human peripheral blood nucleated cells. It is assumed that this assay can be applied as a diagnostic method in the detection of chromosomal translocation which commonly occurs in hematologic malignancies. Methods KCL ‐22 cell line and white blood cells from 21 CML patients were recruited in the study. Cells were isolated and fixed. After permeabilization, cells were resuspended in hybridization buffer and probes were added to the mixture. Subsequently, cells were washed and analyzed on the flow cytometer instrument. The flow cytometry results were compared with qRT ‐ PCR and fluorescent microscope outcomes. Results Using the current principle, 97 ± 2.1% of the KCL ‐22 cells were labeled with b2a2 mRNA ‐specific probes. In addition, seven patients were recognized positive for t(9;22) b2a2. The percentage of cells containing abovementioned translocation in these patients was varied from 3.26% to 97.8%. There was no false‐positive result in negative controls (K562 with BCR ‐ ABL 1 b3a2, NB 4, and Jurkat cell lines) along with blood samples of normal controls. All the results obtained by flow‐ FISH were confirmed by qRT ‐ PCR and fluorescent microscope. Conclusion This strategy benefits from appropriate specificity, sensitivity, rapidity, and ability in the determination of malignant cell percentage. Therefore, it can improve traditional time‐consuming and labor‐intensive FISH method.