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Optimization of rVAR2-Based Isolation of Cancer Cells in Blood for Building a Robust Assay for Clinical Detection of Circulating Tumor Cells.

International journal of molecular sciences (2020-04-05)
Nicolai T Sand, Tobias B Petersen, Sara R Bang-Christensen, Theresa D Ahrens, Caroline Løppke, Amalie M Jørgensen, Tobias Gustavsson, Swati Choudhary, Thor G Theander, Ali Salanti, Mette Ø Agerbæk
ABSTRACT

Early detection and monitoring of cancer progression is key to successful treatment. Therefore, much research is invested in developing technologies, enabling effective and valuable use of non-invasive liquid biopsies. This includes the detection and analysis of circulating tumor cells (CTCs) from blood samples. Recombinant malaria protein VAR2CSA (rVAR2) binds a unique chondroitin sulfate modification present on the vast majority of cancers and thereby holds promise as a near-universal tumor cell-targeting reagent to isolate CTCs from complex blood samples. This study describes a technical approach for optimizing the coupling of rVAR2 to magnetic beads and the development of a CTC isolation platform targeting a range of different cancer cell lines. We investigate both direct and indirect approaches for rVAR2-mediated bead retrieval of cancer cells and conclude that an indirect capture approach is most effective for rVAR2-based cancer cell retrieval.

MATERIALS
Product Number
Brand
Product Description

BRAND® 96-well microplate, U-bottom, round bottom, non-sterile
Sigma-Aldrich
Dulbecco′s Phosphate Buffered Saline, Modified, without calcium chloride and magnesium chloride, liquid, sterile-filtered, suitable for cell culture
Sigma-Aldrich
Chondroitinase ABC from Proteus vulgaris, lyophilized powder, 0.3-3 units/mg solid
Sigma-Aldrich
Heparan sulfate proteoglycan, ≥400 μg/mL glycosaminoglycan
Sigma-Aldrich
Biotin-maleimide, ≥95% (TLC), powder
Sigma-Aldrich
Decorin from bovine articular cartilage, salt-free, lyophilized powder, sterile-filtered
Sigma-Aldrich
Anti-V5 antibody produced in rabbit, IgG fraction of antiserum

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