A Novel Cell Sorting System Based On Au@Fe₃O₄ Nanocomposite Particles

Cell sortingtechnology has been widely applied inthe fields of cell biology,disease diagnosis andtreatment.The magnetic activated cell sorting?MACS?system,in particularly,have attracted ever-increasing attention due to the noticeable advantages such as simple operation,low price,and wide range of applications,which might be better than traditional methods.The most critical factors in MACSarethe type and performances of magnetic particles it used.Compared with micron-scale particles,nanoparticles can bestable forcaptured cells and compatible to flow cytometry analysis,thus favored by most research researchers.However,there are always a few of problems which has not been well solved yet.For instance,the magnetic response of nanoparticles is relatively weak,so that rapid separationcannot be realizedby using ordinary permanent magnets when particles carrying cells.Currently,a special equipment called magnetic separation column,which is usually expensive,single-used and easy to be blocked by target cells,seems have to be used for separation of nano-scale particles.In this study,several types of nanoparticles with different structure and composition were assessed systematically.We hope to find a suitable type of nanoparticle which will have great potential to develop a simple,fast,and high-efficientof cell sorting system under the conventional external magnetic field.First,four types of magnetic particles including the classical core-shell structure nanoparticles Fe₃O₄ @AuGoldMag NPs?50 nm?and Fe₃O₄ @Au GoldMag NPs?30 nm?,the new petal-type nanoparticlesAu@Fe₃O₄GoldMag NPs?30 nm?and the polymer-coated nanoparticlesMono--SA?50 nm?were prepared,and modifiedsurface withstreptavidin?SA?.Then,the physical and chemical properties of these particles were characterized and the immobilization efficiency of SA were estimated aiming to screen out the particles suitable for cell sorting system.Finally,with CD4+T cells as the model,the cell sorting systemwas stablished and envaluated based on the principle ofindirect cell sorting method.During this process,we optimized the amount of capture antibody and magnetic particles,total number of cellsused and the type of magnetic separator.According to the results,the flower-like Au@Fe₃O₄GoldMag NPs?30 nm?waschosen forestablishment of cell sorting system.When 3.5×106 of peripheral blood mononuclear cells?PBMCs?,60 ?L of capture antibody,65 pg of magnetic nanoparticles,and a DynalMag-Spin magnetic separator were used,our method can successfully separate CD4+T cells with the recovery of 93.34%and the purity of 95.21%.Another cell type CD8 + T cells were also successfully separated with a recovery rate of 94.13%and a purity of 95.86%by using this method,which indicated that the flower-like gold magnetic particles can be used as a universal carrier for human peripheral blood immune cell sorting.In addition,with the breast cancer cells line MCF-7 as a circulating tumor cells?CTC?model,our system can capture about 25 cancer cells in 100%fetal bovine serum?FBS?environment?there are about 1-50 cells/mL in the peripheral blood of patients with early cancer?with the recovery rate of 88 ± 2.5%,which provides the valuable theoretical and experimental basis for the rare cells enrichment.