Method for extracting hematopoietic stem cells

Abstract

The invention discloses a method for extracting hematopoietic stem cells, and belongs to the technical field of the extraction of stem cells. The method comprises the steps that a placenta is flushedfrom the exterior with normal saline, 30 mL of an anticoagulant / vasodilator solution is injected at the room temperature; one end of a perfusate needle is inserted into the umbilical artery of the placenta, and the other end of the perfusate needle is inserted into a perfusion container; one end of a reflux liquid needle is inserted into the umbilical vein of the placenta, and the other end of thereflux liquid needle is inserted into the perfusion container to form a perfusion circuit; long-term lavage is carried out for 6 hours with a cell lavage fluid at a flow velocity of 5 drops per minute, and the first-time lavage fluid is collected; lavage is carried out for 30 minutes with the cell lavage fluid at a flow velocity of 5 drops per minute, and the second-time lavage fluid is collected; the first-time lavage fluid and the second-time lavage fluid are mixed to collect the cells, and DMSO is added and mixed with a dextran cell cryoprotectant for cryopreservation. The method for extracting the hematopoietic stem cells has the advantages that the viability is high, and the proportion of CD34+ cells is high.

Description

technical field

[0001] The invention belongs to the technical field of stem cell extraction, and in particular relates to a method for extracting hematopoietic stem cells. Background technique

[0002] With the progress of hematopoietic stem cell research, hematopoietic stem cell transplantation has become an effective treatment for patients with malignant hematological diseases and certain solid tumors. Studies have shown that from the 6th week of pregnancy to the end of pregnancy, the placenta contains hematopoietic stem cells (Hematopoietic stem cells, HSC), and the number of cells is large. Obtaining hematopoietic stem cells from the placenta has the advantages of convenient acquisition and no ethical issues during use. A major obstacle to wider clinical application of umbilical cord blood (UCB) transplantation is its limited cell content. Studies have found that the content of CD34+ cells in human placenta is much higher than that in umbilical cord blood, which can pr...

Examples

Embodiment 1

[0026] Implementation time: 2018.04.07;

[0027] Placenta collection location: Fuzhou General Hospital of Nanjing Military Region;

[0028] Location: Experimental Center of Fujian Haixi Cell Bioengineering Co., Ltd.

[0029] A method for extracting hematopoietic stem cells, comprising the following steps:

[0030] The placentas that were tested negative for infectious diseases such as hepatitis, syphilis, and AIDS and had no obstetrical complications were selected, and the placenta was collected from the hospital after obtaining the informed consent of the parturient and signing the informed consent form before the operation. The placenta was flushed externally with saline and injected with 30 mL of anticoagulant / vasodilator solution (sodium heparin 20 U / mL, phentolamine 1 mg / mL) at room temperature. Then, one end of the perfusate needle is inserted into the placental umbilical artery, and the other end is inserted into the perfusion container; one end of the reflux needle i...

Embodiment 2

[0033] Implementation time of this embodiment: 2018.05.01;

[0034] Placenta collection location: Fuzhou General Hospital of Nanjing Military Region

[0035] Location: Experimental Center of Fujian Haixi Cell Bioengineering Co., Ltd.;

[0036] A method for extracting hematopoietic stem cells, comprising the following steps:

[0037]The placentas that were tested negative for infectious diseases such as hepatitis, syphilis, and AIDS and had no obstetrical complications were selected, and the placenta was collected from the hospital after obtaining the informed consent of the parturient and signing the informed consent form before the operation. The placenta was flushed externally with saline and injected with 30 mL of anticoagulant / vasodilator solution (heparin 20 U / mL, phentolamine 1 mg / mL) at room temperature. Then insert one end of the perfusate needle into the umbilical artery of the placenta, and insert the other end into the perfusion container; insert one end of the re...

Embodiment 3

[0040] Implementation time of this embodiment: 2018.05.06;

[0041] Placenta collection location: Fuzhou General Hospital of Nanjing Military Region

[0042] Location: Experimental Center of Fujian Haixi Cell Bioengineering Co., Ltd.

[0043] A method for extracting hematopoietic stem cells, comprising the following steps:

[0044] The placentas that were tested negative for infectious diseases such as hepatitis, syphilis, and AIDS and had no obstetrical complications were selected, and the placenta was collected from the hospital after obtaining the informed consent of the parturient and signing the informed consent form before the operation. The placenta was flushed externally with saline and injected with 30 mL of anticoagulant / vasodilator solution (heparin 20 U / mL, phentolamine, 1 mg / mL) at room temperature. Then insert one end of the perfusate needle into the placental umbilical artery, and insert the other end into the perfusion container; one end of the return fluid n...

technical field

[0001] The invention belongs to the technical field of stem cell extraction, and in particular relates to a method for extracting hematopoietic stem cells. Background technique

[0002] With the progress of hematopoietic stem cell research, hematopoietic stem cell transplantation has become an effective treatment for patients with malignant hematological diseases and certain solid tumors. Studies have shown that from the 6th week of pregnancy to the end of pregnancy, the placenta contains hematopoietic stem cells (Hematopoietic stem cells, HSC), and the number of cells is large. Obtaining hematopoietic stem cells from the placenta has the advantages of convenient acquisition and no ethical issues during use. A major obstacle to wider clinical application of umbilical cord blood (UCB) transplantation is its limited cell content. Studies have found that the content of CD34+ cells in human placenta is much higher than that in umbilical cord blood, which can pr...

Examples

Embodiment 1

[0026] Implementation time: 2018.04.07;

[0027] Placenta collection location: Fuzhou General Hospital of Nanjing Military Region;

[0028] Location: Experimental Center of Fujian Haixi Cell Bioengineering Co., Ltd.

[0029] A method for extracting hematopoietic stem cells, comprising the following steps:

[0030] The placentas that were tested negative for infectious diseases such as hepatitis, syphilis, and AIDS and had no obstetrical complications were selected, and the placenta was collected from the hospital after obtaining the informed consent of the parturient and signing the informed consent form before the operation. The placenta was flushed externally with saline and injected with 30 mL of anticoagulant / vasodilator solution (sodium heparin 20 U / mL, phentolamine 1 mg / mL) at room temperature. Then, one end of the perfusate needle is inserted into the placental umbilical artery, and the other end is inserted into the perfusion container; one end of the reflux needle i...

Embodiment 2

[0033] Implementation time of this embodiment: 2018.05.01;

[0034] Placenta collection location: Fuzhou General Hospital of Nanjing Military Region

[0035] Location: Experimental Center of Fujian Haixi Cell Bioengineering Co., Ltd.;

[0036] A method for extracting hematopoietic stem cells, comprising the following steps:

[0037]The placentas that were tested negative for infectious diseases such as hepatitis, syphilis, and AIDS and had no obstetrical complications were selected, and the placenta was collected from the hospital after obtaining the informed consent of the parturient and signing the informed consent form before the operation. The placenta was flushed externally with saline and injected with 30 mL of anticoagulant / vasodilator solution (heparin 20 U / mL, phentolamine 1 mg / mL) at room temperature. Then insert one end of the perfusate needle into the umbilical artery of the placenta, and insert the other end into the perfusion container; insert one end of the re...

Embodiment 3

[0040] Implementation time of this embodiment: 2018.05.06;

[0041] Placenta collection location: Fuzhou General Hospital of Nanjing Military Region

[0042] Location: Experimental Center of Fujian Haixi Cell Bioengineering Co., Ltd.

[0043] A method for extracting hematopoietic stem cells, comprising the following steps:

[0044] The placentas that were tested negative for infectious diseases such as hepatitis, syphilis, and AIDS and had no obstetrical complications were selected, and the placenta was collected from the hospital after obtaining the informed consent of the parturient and signing the informed consent form before the operation. The placenta was flushed externally with saline and injected with 30 mL of anticoagulant / vasodilator solution (heparin 20 U / mL, phentolamine, 1 mg / mL) at room temperature. Then insert one end of the perfusate needle into the placental umbilical artery, and insert the other end into the perfusion container; one end of the return fluid n...

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