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High purity of tumour samples is a necessity for accurate genetic and …


Biology Articles » Biotechnology » Red Biotechnology » Negative selection of chronic lymphocytic leukaemia cells using a bifunctional rosette-based antibody cocktail

Abstract
- Negative selection of chronic lymphocytic leukaemia cells using a bifunctional rosette-based antibody cocktail

Negative selection of chronic lymphocytic leukaemia cells using a bifunctional rosette-based antibody cocktail

Salim Essakali1,2, Dennis Carney1,3, David Westerman1,3, Peter Gambell1,3, John F Seymour1,3 and Alexander Dobrovic1,3

1Department of Pathology, Peter MacCallum Cancer Centre, St Andrews Place, Melbourne, Victoria 3002, Australia

2Universitaetsklinik Duesseldorf, Molekulare Pathologie, Moorenstr. 5, 40215 Duesseldorf, Germany

3Department of Pathology, University of Melbourne, Parkville, Victoria 3010, Australia

BMC Biotechnology 2008, 8:6. [Open Access]

 

Abstract

Background

High purity of tumour samples is a necessity for accurate genetic and expression analysis and is usually achieved by positive selection in chronic lymphocytic leukaemia (CLL).

Results

We adapted a bifunctional rosette-based antibody cocktail for negative selection of B-cells for isolating CLL cells from peripheral blood (PB). PB samples from CLL patients were split into aliquots. One aliquot of each sample was enriched by density gradient centrifugation (DGC), while the other aliquot of each sample was incubated with an antibody cocktail for B-cell enrichment prior to DGC (RS+DGC). The purity of CLL cells after DGC averaged 74.1% (range: 15.9 – 97.4%). Using RS+DGC, the purity averaged 93.8% (range: 80.4 – 99.4%) with 23 of 29 (79%) samples showing CLL purities above 90%. RNA extracted from enriched CLL cells was of appropriately high quality for microarray analysis.

Conclusion

This study confirms the use of a bifunctional rosette-based antibody cocktail as an effective method for the purification of CLL cells from peripheral blood.

 


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