br Determination of Ab MNP Si binding to CFSC stained
3.4. Determination of Ab/MNP-Si binding to CFSC-stained GS 9620 and separation from whole blood
To evaluate the performance of Ab/MNP-Si in the blood and the ability of the nanoparticle-silane-antibody conjugate to separate cancer cells, after cell staining, diﬀerent amounts of CFSE-stained SK-BR-3 cells (100 to 4 × 105) were added to 1 mL of fresh human whole blood; separation was done with a magnet and the cells were detected by fluorescence microscopy. As can be seen in Fig. 9, Ab/MNP-Si were generally competent to separate stained cells from whole blood; how-ever, when the number of cells was decreased from 4 × 105 to 100, the number of detected cells obviously decreased, while no cells could be detected without the process of magnetic separation or in the control sample (Fig. 9e and f). Fig. 10 shows the fluorescence microscopy images for the separation of 100 cancer cells in diﬀerent forms.
The most noteworthy point in this experiment in comparison with previous studies is the detection of 100 cancer cells in more than 6 billion sets of blood cells (RBCs + WBCs) with a ratio of 1 cancer cell to 60 million blood cells, whereas in another study , 300 cancer cells could be detected in more than 4.4 billion sets of blood cells with a ratio of 1 cancer cell to 10 million blood cells. The ratio obtained in the present study explicitly documents the significant ability of MNPs to identify and isolate cancer cells. Our results collectively show that Herceptin-coated MNP-Si can be used to identify and/or enrich rare circulating tumor cells.
We developed a new system based on Fe3O4MNPs coated with APTES and conjugated with anti-HER2 antibody to isolate and enrich tumor cells. Several tests confirmed appropriate silane and Herceptin bonding on MNPs without significant eﬀects on their magnetic prop-erties. Data from flow cytometry and fluorescence microscopy showed the Ab/MNP-Si nanoparticles to eﬃciently locate and separate low number of HER2-expressing tumor cells (SK-BR-3), from PBMCs and from fresh human whole blood, even when the concentration of the target cells was very low (with a ratio of 1 cancer cell to 60 million blood cells). These results collectively show that Herceptin-coated MNP-Si can be used to identify and/or enrich low number of tumor cells.
We thank K. Shashok (AuthorAID in the Eastern Mediterranean) for improving the use of English in parts of the manuscript.
Appendix A. Supplementary data
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