SHORT COMMUNICATION: DEVELOPMENT OF FLOW CYTOMETRY-FLUORESCENT IN SITU HYBRIDIZATION (FLOW-FISH) METHOD FOR DETECTION OF PML/RARA CHROMOSOMAL TRANSLOCATION IN ACUTE PROMYELOCYTIC LEUKEMIA CELL LINE

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ZAHEDIPOUR FATEMEH; RANJBARAN REZA; BEHZAD BEHBAHANI ABBAS; TAVAKOL AFSHARI KHALIL; OKHOVAT MOHAMMAD ALI; TAMADON GHOLAMHOSSEIN; SHARIFZADEH SEDIGHEH

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Journal: Avicenna Journal of Medical Biotechnology Year:2017 | Volume:9 | Issue:2 Page(s): 104-108

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Title

SHORT COMMUNICATION: DEVELOPMENT OF FLOW CYTOMETRY-FLUORESCENT IN SITU HYBRIDIZATION (FLOW-FISH) METHOD FOR DETECTION OF PML/RARA CHROMOSOMAL TRANSLOCATION IN ACUTE PROMYELOCYTIC LEUKEMIA CELL LINE

Author(s)

Keywords

ACUTE PROMYELOCYTIC LEUKEMIA

FLOW CYTOMETRY

FLUORESCENT IN SITU HYBRIDIZATION

PML-RARALPHA

Abstract

Background: ACUTE PROMYELOCYTIC LEUKEMIA (APL) is a subclass of acute myeloid leukemia. The chromosomal aberration in 95% of APL cases is t (15; 17) (q22; q21), which prevents cell differentiation. Characterization of the underlying molecular lesion is valuable in determining optimal treatment strategy. The goal of this study was to develop a new and powerful Flow- FISH technique to detect the long isoform (L) of PML-RARa fusion transcript in NB4 cell line.Methods: To achieve the best condition for fixation, two different fixatives including 2% paraformaldehyde and 75% ethanol were used.0.2% Triton X-100 and 0.2% saponin were used for the permeabilization step.In hybridization, a wide range of times and temperatures were used and probe was designed in FRET system. Results were confirmed by fluorescent microscope assay and reverse transcription PCR.Results: In the present study, a novel technique was successfully optimized that combines in situ hybridization with FLOW CYTOMETRY to detect the presence of PML-RARa transcript. Using standard fixation and permeabilization protocol of 2% PFA and 0.2% saponin gave the best fluorescent results in FLOW CYTOMETRY. Also, results indicated that the optimum time and temperature for hybridization was 2hr at 42°C. The results of reverse transcription PCR and fluorescent microscopy confirmed the presence of PML-RARa transcript.Conclusion: The concordance between the results of Flow-FISH and those of two other techniques including reverse transcription PCR and FISH indicated that this method would be applicable as a diagnostic test for APL in clinical samples and MRD monitoring.

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APA: Copy

ZAHEDIPOUR, FATEMEH, RANJBARAN, REZA, BEHZAD BEHBAHANI, ABBAS, TAVAKOL AFSHARI, KHALIL, OKHOVAT, MOHAMMAD ALI, TAMADON, GHOLAMHOSSEIN, & SHARIFZADEH, SEDIGHEH. (2017). SHORT COMMUNICATION: DEVELOPMENT OF FLOW CYTOMETRY-FLUORESCENT IN SITU HYBRIDIZATION (FLOW-FISH) METHOD FOR DETECTION OF PML/RARA CHROMOSOMAL TRANSLOCATION IN ACUTE PROMYELOCYTIC LEUKEMIA CELL LINE. AVICENNA JOURNAL OF MEDICAL BIOTECHNOLOGY (AJMB), 9(2), 104-108. SID. https://sid.ir/paper/313980/en

Vancouver: Copy

ZAHEDIPOUR FATEMEH, RANJBARAN REZA, BEHZAD BEHBAHANI ABBAS, TAVAKOL AFSHARI KHALIL, OKHOVAT MOHAMMAD ALI, TAMADON GHOLAMHOSSEIN, SHARIFZADEH SEDIGHEH. SHORT COMMUNICATION: DEVELOPMENT OF FLOW CYTOMETRY-FLUORESCENT IN SITU HYBRIDIZATION (FLOW-FISH) METHOD FOR DETECTION OF PML/RARA CHROMOSOMAL TRANSLOCATION IN ACUTE PROMYELOCYTIC LEUKEMIA CELL LINE. AVICENNA JOURNAL OF MEDICAL BIOTECHNOLOGY (AJMB)[Internet]. 2017;9(2):104-108. Available from: https://sid.ir/paper/313980/en

IEEE: Copy

FATEMEH ZAHEDIPOUR, REZA RANJBARAN, ABBAS BEHZAD BEHBAHANI, KHALIL TAVAKOL AFSHARI, MOHAMMAD ALI OKHOVAT, GHOLAMHOSSEIN TAMADON, and SEDIGHEH SHARIFZADEH, “SHORT COMMUNICATION: DEVELOPMENT OF FLOW CYTOMETRY-FLUORESCENT IN SITU HYBRIDIZATION (FLOW-FISH) METHOD FOR DETECTION OF PML/RARA CHROMOSOMAL TRANSLOCATION IN ACUTE PROMYELOCYTIC LEUKEMIA CELL LINE,” AVICENNA JOURNAL OF MEDICAL BIOTECHNOLOGY (AJMB), vol. 9, no. 2, pp. 104–108, 2017, [Online]. Available: https://sid.ir/paper/313980/en

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