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Method for preparing CD3+CD56+cells through high-killing K562cells from perinatal placental blood

A placental blood and perinatal technology, applied in the field of preparation of CD3+CD56+ cells, can solve the problems of poor stability of blood components, long cell preparation cycle, lack of collection process, etc., and achieves shortened preparation cycle, high proliferation potential, The effect of increasing the number of cell proliferation

Active Publication Date: 2013-06-26
JIANGSU HEZE STEM CELL GENE ENG CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0016] In view of the defects in the prior art, such as poor stability of blood components, lack of safety in the collection process, long cell preparation cycle, and low proliferation potential, the present invention provides a high-killing blood cell derived from perinatal placenta. CD3 of K562 cells + CD56 + cell preparation method

Method used

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  • Method for preparing CD3+CD56+cells through high-killing K562cells from perinatal placental blood
  • Method for preparing CD3+CD56+cells through high-killing K562cells from perinatal placental blood
  • Method for preparing CD3+CD56+cells through high-killing K562cells from perinatal placental blood

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preparation example Construction

[0055] CD3 of the present invention derived from high killing K562 cells in perinatal placental blood + CD56 + The preparation method of cells comprises the following steps:

[0056] a. Collect perinatal placental blood in an airtight container. First, within 10 seconds after the newborn is delivered, clamp it with two hemostatic forceps at a distance of 5 to 8 cm from the umbilicus of the newborn, ligate two places between the two clamps, cut the umbilical cord of the newborn from the middle, and take the newborn away. Normal treatment. The perinatal placental blood was collected from the filling part of the umbilical vein far away from the umbilical cord with gauze and disinfected carefully, and was anticoagulated with an anticoagulant containing heparin sodium. For example, a special blood collection bag containing heparin sodium anticoagulant can be used for blood collection. The specific operation is to insert the needle of the blood collection bag into the umbilical ...

Embodiment 1

[0060] First, within 10 seconds after the newborn is delivered, two hemostats are clamped at a place 6cm away from the umbilicus of the newborn, two places are ligated between the two clamps, the umbilical cord of the newborn is cut from the middle, and the newborn is taken away. Process normally. Disinfect by rubbing carefully with gauze from the umbilical vein engorgement away from the end of the umbilical cord. After disinfection, a special blood collection bag containing heparin sodium anticoagulant was used to collect perinatal placental blood. The specific operation is to insert the needle into the umbilical vein of the sterilized part, and draw perinatal placental blood through the umbilical vein. After the collection, close the blood collection bag tube, and quickly shake the collected blood with the anticoagulant in the bag to avoid coagulation. Then put the blood collection bag and the biological ice bag together in a special incubator, keep the temperature in the ...

Embodiment 2

[0074] In order to show the difference, the scheme in Example 1 is hereinafter referred to as this scheme; and the scheme in which SCF is not added to the culture medium and other conditions are the same as those in Example 1 is called the control scheme. The following CD3 obtained through this scheme and the control scheme + CD56 + The cells were tested for killing tumors.

[0075] Determination of CD3 with CCK-8 (Cell Counting Kit-8) reagent + CD56 + cytotoxicity of tumor cells. CCK-8 reagent contains WST-8 (water-soluble tetrazolium salt), which can be reduced to a highly water-soluble yellow formazan product (Formazan) by dehydrogenase in the mitochondria of cells. The amount of formazan produced is directly proportional to the number of viable cells. The light absorption value was measured at a wavelength of 450nm by an enzyme-linked immunosorbent assay instrument, which can indirectly reflect the number of living cells.

[0076] CD3 to be harvested + CD56 + The c...

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Abstract

The invention provides a method for preparing CD3+CD56+cells through high-killing K562 cells from perinatal placental blood. The method comprises the step of: separating mononuclear cells from the perinatal placental blood so as to carry out induction differentiation to form CD3+CD56+cells. By utilizing the method, the preparation period of the cells can be shortened, and moreover, the multiplication capability of the cells is improved.

Description

technical field [0001] The present invention relates to a kind of cell culture and immunotherapy, especially to a kind of CD3 that kills K562 cells + CD56 + Cell preparation method. Background technique [0002] CD3 + CD56 + Cells are a type of immune cells induced by various cytokines in vitro. It is obtained from mononuclear cells in human blood after being co-cultured with various cytokines for a period of time in vitro. Since this cell also expresses CD3 + (CD3 + It is a T cell surface specific protein, only present on the surface of T cells) and CD56 + (CD56 + NK cell surface-specific protein) two membrane protein molecules, so it is also called NK cell (natural killer cell, natural killer cell)-like T lymphocyte (NKT cell), which has strong anti-tumor activity of T lymphocytes and The non-MHC (major histocompatibility complex major histocompatibility complex) restriction tumor killing advantage of NK cells. Compared with LAK cells (lymphokine activated killer...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0783
Inventor 刘乐锋王溢文魏昌盛林强刘洋
Owner JIANGSU HEZE STEM CELL GENE ENG CO LTD
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