The invention discloses an improved high-efficiency human autologous (
Regulatory T cell) separation and amplification method. The improved high-efficiency human autologous separation and amplificationmethod comprises the following steps: S1, treating 5000mL of PB (
Peripheral Blood) through a
blood cell separator (single sampling
machine) in 1 to 2 cycles, obtaining PBMCs (
Peripheral Blood Mononuclear Cells) in order magnitude of 10*9 to 10*10, sieving separated PBMCs through a nylon net of which the
pore diameter is 30 mum, and removing a
cell cluster in suspension cells; S2, labeling non-CD4+ cells; S3, removing the non-CD4+ cells through
negative selection of an LD
separation column; S4, labeling cd25+ cells on an
antibody; S5, positively selecting the CD25+ cells through an MS column.According to the improved high-efficiency human autologous separation and amplification method disclosed by the invention, 5000mL of
peripheral blood can be treated through the
blood cell separator (single acquisition
machine) in 1 to 2 cycles, the PBMCs (
Peripheral Blood Mononuclear Cells) in order magnitude of 10*9 to 10*10 can be obtained, and mononuclear cells of which the number is 100 timesor above of the number of the mononuclear cells obtained through a traditional
blood sampling method can be easily obtained; other blood components are in
direct feedback, so that waste is avoided, and the influence on
total blood component and
blood cell count of a patient is more slight.