TH64281B - In vitro techniques for stem cells - Google Patents

Abstract

DC60 (01/10/47) A method for extracted blood has been developed consisting of centrifugation in a first gradient suitable for the selection of first-pass cells with a density of less than 1.077 g. /ml Subsequently, the first-pass cells were centrifuged in a second gradient suitable for screening second-pass cells with a density between 1.055 and 1.074 g/ml. Let's increase the number By cultivating second-pass cells for between 3-30 days, a method for the extracted blood has been devised, consisting of centrifuging the blood in a first gradient suitable for the selection of first-pass cells. Subsequently, first-pass cells were centrifuged in a second gradient suitable for selecting second-pass cells with a density between 1.055 and 1.074 g/ml, cells with a density between 1.055. and 1.074 g/ml were proliferated by culturing second-pass cells for between 3-30 days.

Claims (6)

Disclaimer (all) which will not appear on the advertisement page: Revised 25/8/2017 1. Methods for use with extracted blood, including: introducing blood into the gradient. A first gradient, which is suitable for screening first-pass cells with a density of less than 1.077 g / ml; Introduction of first-pass cells into a second gradient suitable for screening second-pass cells with a density between 1.055 and 1.074 g / ml; Increasing the number of cells with a density between 1.055 and 1.074 g / ml by culturing second-pass cells for between 1 and 30 days in a culture compound consisting of one or more substances selected from the group consisting of: Autologasserum, VEGF, b-FGF, IGF and heparin (heparin); In order to use cultured cell populations containing blood cell progenerators. (hematopoietic progenitor cell), and especially in the endothelial Progester cells (EPCs) ------------------------------------------ -------------------------------------------------- --------------------- (Old) 1. Methods used with the extracted blood are as follows: Centrifuge the blood in a first gradient suitable for the selection of first-pass cells with density less than 1.077g / ml; The first-pass cells were centrifuged in a second gradient, which was suitable for the selection of second-pass cells with a density between 1.055 and 1.074 g / ml; The number of cells with a density of 1.055 and 1.074g / ml was increased by culturing second-pass cells for 3-30 days. 2. Method in claim 1: The centrifugation of blood cells in the first gradient includes Centrifugation of blood cells in a phicol-like gradient 3. Method in claim 1 Centrifugation in the second gradient includes centrifugation of blood cells in a percol-like gradient. 4. Method of claim 1: Centrifugation in the second gradient includes centrifugation of blood cells in a gradient like an optipress. Extracted It is as follows: tissue containing stem cells is centrifuged in gradient. First, which is suitable for casting first-passcells. With a density of less than 1.077 g / ml; The first-pass cells were centrifuged in a second gradient, which was suitable for the selection of second-pass cells with a density between 1.055 and 1.074 g / ml; Second-pass cells were centrifuged in a third gradient, which was suitable for screening third-pass cells with densities between 1.055 and 1.068 g / ml; The cell density between 1.055 and 1.074 g / ml was increased by culturing third-pass cells for 3-30 days. 6. Method in claim 5. The third gardent centrifugation was suitable for selection. Cell density between 1.059 and 1.068 g / ml and centrifugation of second-pass cells in the third gradient was defined as the selection of cells with a density between 1.059 and 1.068 g / ml. Extracted Is as follows The blood was centrifuged in a second gradient suitable for the selection of first-pass cells with a density of less than 1.077g / ml. The first-pass cells were centrifuged in a second gradient, which was suitable for the selection of second-pass cells at Has a density between 1.055 and 1.074 g / ml; And bake second-pass cells on the surface area with antibodies. 8. Method of application with blood extracted. It is as follows: Centrifugation in one gradient suitable for screening first-pass cells with density less than 1.077g / ml; The first-pass cells were centrifuged in a second gradient, which was suitable for the selection of second-pass cells with a density between 1.055 and 1.074 g / ml; And bake second-pass cells on the surface area with growth-accelerating molecules in addition to collagen Or Fobronectin 9. Method in claim 8 Cultivation of second-pass cells means laying second-pass cells on the ground consisting of at least one collagen and fibronectin. Accelerated molecules Growth 1 0. Method of application with blood extracted. It is as follows: Centrifugation in one gradient suitable for screening first-pass cells with density less than 1.077g / ml; The first-pass cells were centrifuged in a second gradient, which was suitable for the selection of second-pass cells with a density between 1.055 and 1.074 g / ml; Second-cells were cultured for 1-5 days in a culture media containing less than 5% serum 1 1. Methods applied to the extracted blood. It is as follows: Centrifugation in one gradient which is suitable for screening first-pass cells with density less than 1.077 g / ml; The first-pass cells were centrifuged in a second gradient, suitable for the selection of second-pass cells with a density between 1.055 and 1.074 g / ml; Second-pass cells were cultured for 1-5 days in a culture media containing more than 10% serum 1 2. Method in claim 11 Second-pass cells were cultured for second-pass cells in the culture medium. The serum with less than 20% 1 3. Method of application with blood extracted. It is as follows: Centrifugation in one gradient suitable for screening first-pass cells with density less than 1.077g / ml; The first-pass cells were centrifuged in a second gradient, which was suitable for the selection of second-pass cells with a density between 1.055 and 1.074 g / ml; In low-serum, second-pass cells are cultured in culture media with less than 10% serum and in high serum, second-pass cells are cultured in culture media with serum greater than or equal to 10% 1 4. Method In claim 13, seeding second-pass-cells in the low-serum range means cultivating second-pass cells for a period of 1-5 days. 1 5. Method for claim 13. High-serum means culturing second-pass cells between 1-30 days. 1 6. Method in claim 13 Low-serum second-pass-cells cultivation is performed after second-pass cells are cultivated during the period of time. high-serum 1 7. Method for claim 13 Low-serum second-pass-cells culture is performed after second-pass cells in the high-serum period. Extracted Is as follows The blood was centrifuged in one gradient which was suitable for screening first-pass cells with density less than 1.077g / ml; The first-pass cells were centrifuged in a second gradient, which was suitable for the selection of second-pass cells with a density between 1.055 and 1.074 g / ml; During hypoxia, which has occurred for at least 2 hours, cultivate low-oxygen second-pass cells. And during hypoxia, which occurs for at least 1 day, second-pass cells are cultured under hypoxia. 1 9. Method in claim 18, hypoxia and hypoxia occur during the cultivation phase. Low oxygen second-pass cells culture means low oxygen second-pass cells cultured during the first two days of the 2 0. Hold the right 18 Low and low oxygen conditions occur during the incubation phase. Low oxygen second-pass cells were cultured in less than 30 days and second-pass cells were cultured under hypoxia during the last two days of the cell culture period. Hold the right 18 Low and low oxygen conditions occur during the incubation phase. Low oxygen second-pass cells are cultured for at least 2 hours during the first two days, and low oxygen second-pass cells are cultured for at least 2 hours during the first two days. Two days later of the cell culture period 2 2. Method in claim 18 The cultivation of second-pass cells under hypoxia is to be performed prior to culturing second-pass cells under hypoxia. 18. Cultivation of second-pass cells under hypoxia is performed after culturing second-pass cells in hypoxia. 2 4. Methods applied to the extracted blood. It is as follows: Centrifugation in one gradient suitable for screening first-pass cells with density less than 1.077g / ml; The first-pass cells were centrifuged in a second gradient, which was suitable for the selection of second-pass cells with a density between 1.055 and 1.074 g / ml; The second-pass cells were cultured in a culture medium consisting of at least one of the following substances: erythropoi, itinstadin, molecule, and anti-diabetic. 24 anti-diabetic drugs: rosiglidasone; second-pass cells culture: second-pass cells cultured in a cell culture with rosieglida zone 2. 6. Methods taken Applied to the extracted blood It is as follows: The first gradient of the stem cell tissue was centrifuged in the first gradient, suitable for the selection of first-pass cells with density less than 1.077 g / ml; The first-pass cells were centrifuged in a second gradient, which was suitable for the selection of second-pass cells with a density between 1.055 and 1.074 g / ml; The number of cells with a density of 1.055 and 1.074 g / ml was increased by culturing second-pass cells for 3-30 days. 2 7. Method in claim 26 consisted of an extract of marrow stem cells 2. 8. The method in claim 26 consists of the removal of the stem cells from the fat. Suction to assist in stem cell extraction 2. 9. Method in claim 26 consists of blood stem cell extract 3. 0. Method in claim 26: second-pass cells cultivate second-pass cells. In the first container during cell culture, some of the second-pass cells are removed from the first container at the end of the first cell culture, and the cells removed from the first container. The second in the second part of the Cell culture 3 1. The method in claim 30, removing some of the second-pass cells, means selecting a cell so that the cell adjacent to the surface of the first container comes out. 2. The method in claim 30, removing some of the second-pass cells, means selecting the cells so that the cells that do not stick to the surface of the first container. 3. Method in claim 30 one container has a growth accelerator molecule on the surface and cell culture in one container means culture cells in one container with Growth accelerated molecules 3 4. Methods in claim 33 Growth-accelerating molecules were selected from the list of growth factor collagen pebronetin. And an anti-body for receptor 3 of stem cell surfaces 5. Method in claim 30, the second container contains the surface growth accelerator molecules and the second container cell culture means cell culture in the second container. There are 3 growth accelerating molecules 6. Methods in claim 35 Molecular growth accelerators were selected from the list of growth factor collagen pebronetin. And an anti-body for receptor, stem cell surface