Evaluation of a new flow cytometry based method for detection of BCR-ABL1 fusion protein in chronic myeloid leukemia

Swati Dasgupta; Ujjal K Ray; Arpita Ghosh Mitra; Deboshree M. Bhattacharyya; Ashis Mukhopadhyay; Priyabrata Das; Sudeshna Gangopadhyay; Sudip Roy; Soma Mukhopadhyay

doi:10.5045/br.2017.52.2.112

Quick links

Most Cited Most Read Ahead of Print Archives Go to E-submission

Original Article

Split Viewer

Blood Res 2017; 52(2):

Published online June 22, 2017

https://doi.org/10.5045/br.2017.52.2.112

Evaluation of a new flow cytometry based method for detection of BCR-ABL1 fusion protein in chronic myeloid leukemia

Swati Dasgupta¹, Ujjal K Ray², Arpita Ghosh Mitra⁴, Deboshree M. Bhattacharyya¹, Ashis Mukhopadhyay³, Priyabrata Das¹, Sudeshna Gangopadhyay¹, Sudip Roy⁴, and Soma Mukhopadhyay¹

Author Info. +

¹Department of Molecular Biology and Hematology, Netaji Subhas Chandra Bose Cancer Research Institute, West Bengal, India.

²Department of Pathology, Netaji Subhas Chandra Bose Cancer Research Institute, West Bengal, India.

³Department of Hemato-Oncology, Netaji Subhas Chandra Bose Cancer Research Institute, West Bengal, India.

⁴Department of HLA & Molecular Lab, Medica Superspeciality Hospital, West Bengal, India.

Correspondence to : Soma Mukhopadhyay, Ph.D. Department of Molecular Biology and Hematology, Netaji Subhas Chandra Bose Cancer Research Institute, 16 A Park Lane, Kolkata 700016, India. s.amukhopadhyay@gmail.com

Received: September 27, 2016; Revised: November 17, 2016; Accepted: January 6, 2017

Abstract

Go to

Abstract

Background

Philadelphia chromosome, a hallmark of chronic myeloid leukemia (CML), plays a key role in disease pathogenesis. It reflects a balanced reciprocal translocation between long arms of chromosomes 9 and 22 involving BCR and ABL1 genes, respectively. An accurate and reliable detection of BCR-ABL fusion gene is necessary for the diagnosis and monitoring of CML. Previously, many technologies, most of which are laborious and time consuming, have been developed to detect BCR-ABL chimeric gene or chromosome.

Methods

A new flow cytometric immunobead assay was used for detection of BCR-ABL fusion proteins and applicability, sensitivity, reliability, efficacy and rapidity of this method was evaluated.

Results

From February 2009 to January 2014, a total 648 CML patients were investigated for the status of BCR-ABL1 protein. Among them, 83 patients were enrolled for comparative study of BCR-ABL1 positivity by three routinely used procedures like karyotyping, and quantitative real time PCR (RT-PCR) as well as immunobead flow cytometry assay. BCR-ABL protein analysis was found consistent, more sensitive (17% greater sensitivity) and reliable than the conventional cytogenetics, as flow cytometry showed 95% concordance rate to RT-PCR.

Conclusion

BCR-ABL fusion protein assay using a new flow cytometric immunobead might be useful in the diagnosis and monitoring CML patients.

Keywords Chronic myeloid leukemia, BCR-ABL1, Philadelphia chromosome, Flow cytometry

Article

Original Article

Blood Res 2017; 52(2): 112-118

Published online June 22, 2017 https://doi.org/10.5045/br.2017.52.2.112

Evaluation of a new flow cytometry based method for detection of BCR-ABL1 fusion protein in chronic myeloid leukemia

Swati Dasgupta¹, Ujjal K Ray², Arpita Ghosh Mitra⁴, Deboshree M. Bhattacharyya¹, Ashis Mukhopadhyay³, Priyabrata Das¹, Sudeshna Gangopadhyay¹, Sudip Roy⁴, and Soma Mukhopadhyay¹

¹Department of Molecular Biology and Hematology, Netaji Subhas Chandra Bose Cancer Research Institute, West Bengal, India.

²Department of Pathology, Netaji Subhas Chandra Bose Cancer Research Institute, West Bengal, India.

³Department of Hemato-Oncology, Netaji Subhas Chandra Bose Cancer Research Institute, West Bengal, India.

⁴Department of HLA & Molecular Lab, Medica Superspeciality Hospital, West Bengal, India.

Correspondence to:Soma Mukhopadhyay, Ph.D. Department of Molecular Biology and Hematology, Netaji Subhas Chandra Bose Cancer Research Institute, 16 A Park Lane, Kolkata 700016, India. s.amukhopadhyay@gmail.com

Received: September 27, 2016; Revised: November 17, 2016; Accepted: January 6, 2017

Abstract

Background

Methods

A new flow cytometric immunobead assay was used for detection of BCR-ABL fusion proteins and applicability, sensitivity, reliability, efficacy and rapidity of this method was evaluated.

Results

Conclusion

BCR-ABL fusion protein assay using a new flow cytometric immunobead might be useful in the diagnosis and monitoring CML patients.

Keywords: Chronic myeloid leukemia, BCR-ABL1, Philadelphia chromosome, Flow cytometry

Abstract

Table 1 . Specific primer sequences for detection of BCR-ABL1 RQ-PCR..

Table 2 . MFI values of all CML patients (N=648)..

Table 3 . Comparison of BCR-ABL1 status results using karyotyping, flow cytometry and RQ-PCR..

Table 4 . Variable transcripts in 83 CML patients detected by nested RT-PCR..

Table 5 . Comparison of mean MFI values in different response groups using karyotyping..

^a)Ph positive metaphase cells are absent. ^b)Presence of Ph positive metaphase cells. ^c)Presence of Ph positive metaphase cells 36–65%. ^d)Presence of Ph positive metaphase cells..

Table 6 . Comparison of mean MFI values in different response groups using RQ-PCR (N=68)..

^a)No transcript has been detected. ^b)BCR-ABL/ABL ratio of different transcripts : 0.005–0.01%. ^c)BCR-ABL/ABL ratio of different transcripts ≤0.1%. ^d)BCR-ABL/ABL ratio of different transcripts >0.1%..

Table 7 . Comparison between flow cytometry and karyotyping against RQ-PCR..