Streptavidin Magnetic Particles Based CD4 Cells Immunomagnetic Separation

Immunomagnetic cell sorting is based on the specific reaction between the antigens and antibodies. The magnetic nanoparticles as a carrier to label the antibodies, subsequently, the specific binding with antigens occurs on the surface of the target cells and then the target cells are separated with the aid of high-gradient magnetic field. Nowadays, immunomagnetic separation is highly valuable for research and biomedicine application. Several magnetic separation commercial kits have been developed by different overseas companies, such as Miltenyi and Dynal Beads. Thus, it is extraordinarily meaningful to develop a protocol of immunomagnetic cell separation by using China self-made magnetic particles immediately.Based on the method of direct and indirect cell sorting, this study was developed a protocol for cell sorting by using immunomagnetic particles. Five different self-made magnetic particles (xMagTM Isothiocyanate MicroBeads, GoldMag(?)AS MicroBeads, GoldMag(?)CS MicroBeads, xMagTM Protein G MicroBeads, Streptavidin MicroBeads) were used as carriers for sorting different lymphocytes from human peripheral blood samples. The sorted target cells were then quantified by flow cytometry. Finally, the streptavidin-modified magnetic microspheres were chosen for coupling with biotinylated antibody and subsequent selective isolation of target cells from complex samples based on highly specific antigen-antibody reactions. The method was optimizing according to binding ability, temperature and time and also compared with other commercial products depends. Although all these five self-made magnetic particles could be used in immunomagnetic cell separation, results showed that using 1/2 streptavidin loaded SA magnetic particle was preferred. As to sorting CD4 cells, the dosage of biotin-antibody is 0.4μg and magnetic particle is 16μg that would be suitable when the total number of cells are 1×106. The sorting solution was cooling down first, and incubated for 15 min at 4℃. The both sorting efficiency and purity can reach up to 90% without damage the cells. Moreover, more kinds of cells, such as CD4 cells, CD8 cells, CD16 cells, CD19 cells, also could be successfully separated by SA magnetic particles. Compared with MACS Streptavidin MicroBeads which has a high reputation in the field of cell sorting, all results had showed there was no obvious difference between these two products. This work laid a good groundwork for more lymphocytes separation.