Optimization of methods for the detection of BCR-ABL activity in Philadelphia-positive cells.

Hamilton, Ashley, Alhashimi, Fatma, Mysina, Svetlana, Jorgensen, Heather G and Holyoake, Tessa L (2009) Optimization of methods for the detection of BCR-ABL activity in Philadelphia-positive cells. Experimental hematology, 37 (3). pp. 395-401. ISSN 1873-2399

Full text not available from this repository.

Search Google Scholar

Abstract

OBJECTIVE

The recent success in treating chronic myeloid leukemia (CML) with tyrosine kinase inhibitors (TKI), such as imatinib mesylate (IM), has created a demand for reproducible methods to accurately assess inhibition of BCR-ABL activity within CML cells, including rare stem and progenitor cells, either in vitro or in vivo. The purpose of this study was to develop an enzyme-linked immunosorbent (ELISA) method to measure total tyrosine phosphorylation (P-Tyr) in small samples of cells that express BCR-ABL and to compare to more established methods.

MATERIALS AND METHODS

The assay was first validated in BCR-ABL wild-type and mutant vs BCR-ABL-negative cell lines. P-Tyr levels were then measured by ELISA in primary CD34(+) CML cells treated with IM.

RESULTS

In vitro exposure to TKI resulted in decreases in the level of P-Tyr, in both BCR-ABL-positive cell lines and primary CD34(+) CML samples, which were comparable to the reduction in P-Tyr by flow cytometry and phosphorylation of CrkL by either Western blot or flow cytometry.

CONCLUSION

We have developed an accurate ELISA method to measure BCR-ABL activity within Ph(+) cells, which is comparable to other in vitro BCR-ABL assessment techniques in terms of sensitivity and could be adapted for high throughput.

Item Type: Article
Subjects: Sciences > Biomedical Sciences
Depositing User: Svetlana Mysina
Date Deposited: 08 Oct 2018 09:54
Last Modified: 09 Oct 2018 10:44
URI: http://sure.sunderland.ac.uk/id/eprint/10013

Actions (login required)

View Item View Item