63 results about "Autologous serum" patented technology

The invention relates to lyophilized powder of human mesenchymal stem cell culture supernatant and a preparation method of the lyophilized powder. The preparation method comprises the following steps of: culturing a sub-culturing human mesenchymal stem cell by using a mesenchymal stem cell culture medium containing autologous serums; collecting a cell culture supernatant with low sub-culturing times; and preparing the collected supernatant into the lyophilized powder. The lyophilized powder prepared by the preparation method of the lyophilized powder of the human mesenchymal stem cell culture supernatant effectively preserves various cell factor mixtures with biological activities in the human mesenchymal stem cell culture supernatant, is capable of effectively solving the problem of short storage life of the supernatant and lays a foundation for an industrial application of the human mesenchymal stem cell culture supernatant.

The invention belongs to the technology field of the cellular immunology, and particularly relates to a method for the dominant amplification of in vitro peripheral blood nature killer cells with a large amount. The method comprises the steps as follows: (1) separating single nucleus cell from human or animal peripheral blood; (2) suspending the single nucleus cell in an RPMI 1640 culture solution containing 5%-15% autologous serum and pretreating with methyl-Beta-cyclodextrin with an effective concentration of 1-4mmol / L for 36-60 hours; and (3) adding recombined interleukin 2, and amplification-culturing in an RPMI 1640 culture solution containing 5% of new-born calf serum and 5% of autologous serum for above 10 days. The method has the advantages that the blood sample amount is small; the cost is low; the operation is convenient; and the amplification multiple is high, and the nature killer cells can be amplified by 392-1752 times in a short term.

The invention relates to an in-vitro pure culture method for natural killer cells. The method includes the following steps that peripheral blood is collected, single karyocyte is separated, magnetic beads combined with specific antibodies are adopted to remove the NK cells, and NK cell populations are obtained; the NK cell populations are put into a culture bottle coated with multiple antibodies,and are subjected to activation culture with a complete medium containing IL-2, IL-7, IL-15, anti-CD16, OK432 and inactivated autologous serum, and inactivated NK cell populations are obtained; the inactivated NK cell populations are put into a culture bottle free of antibody coating and continue to carry out multiplication culture, and the natural killer cells are obtained. Any heterologous serumingredients and trophoblast cells are not used in the in-vitro pure culture method, and high amplification efficiency can be wholly obtained; meanwhile, the purity of the amplified NK cells reaches 90% or above, in-vitro efficient pure culture is achieved, and the clinical application requirement is conveniently met.

The invention relates to a preparation method used for high efficiency amplification of natural killer (NK) cells. The preparation method comprises following steps: peripheral blood is collected, andsingle karyocytes are separated; the karyocytes are introduced into a culture bottle treated via coating with CD16 monoclonal antibody and HER2 monoclonal antibody for culturing; IL-2, IL-15, IL-21, and inactivated autologous serum are introduced into a culture medium at the same time; the cells are transformed into a culture bottle free of coating treatment, liquid supply is carried out every 2 to 3 days; and at last the obtained NK cells are collected. According to the preparation method, no heterogenous serum or trophoblastic cell is adopted, operation process is simple, high NK cell yieldis achieved, and the clinical application value is high.

The invention discloses a method for establishing a PDX (Patient Derived Xenograft) model of human blood tumor. The method comprises the steps of extracting a blood tumor cell of a patient, adding rabbit anti-human thymocyte immunoglobulin ATG (Anti-Thymocyte Globulin) and patient autologous serum, mixing, incubating and after completing the incubation, re-suspending the obtained cell and inoculating in a mouse; and feeding the inoculated mouse with CsA (Cyclosporine A) and lasting for 5-10 days starting from 2 days before the inoculation of the blood tumor cell. The method disclosed by the invention has the beneficial effects that a novel highly immunodeficient NCG mouse independently developed in China is firstly adopted to establish a human leukemia PDX model; by orally taking human thymocyte immunoglobulin pretreatment sample combined with the cyclosporine for a long term, donor-derived T cells are removed and inhibited; and by combining the toxic effect of the ATG on lymphocytes with the functional blocking effect of the CsA on T lymphocytes, the immune function of the T lymphocytes can be persistently inhibited and the success rate of PDX modeling of the blood tumor is significantly improved.

The invention relates to a stem cell repairing material as well as a preparation method and application thereof. The stem cell repairing material comprises a stem cell and a cell carrier, wherein the stem cell is an autologous adipose-derived mesenchymal stem cell (ADSC) with a concentration of 105-108 / ml, and the cell carrier is autoserum, normal saline (0.9%) or a glucose injection (5%). In the invention, adopted seed cells are prepared through separating and purifying autologous adipose tissues of patients, thereby avoiding the ethical disputes and the immunological rejection. The material drawing of the adipose tissues can be performed by using an instrument suction method, which is simple in operation and can bring less traumas and pains to the patients. The cell carrier used in the invention is the autoserum, normal saline (0.9%) or glucose injection (5%), therefore, the cell carrier mixed with mesenchymal stem cells and other ingredients can be injected into the patents by virtue of syringes. By using the stem cell repairing material provided by the invention, the operation process is simple, no scar is left, and the wounds and pains for the patients are avoided, therefore, the stem cell repairing material can be easily accepted by the patients.

The invention relates to a method of the induction and differentiation of the neural stem cells and the neural cells from bone marrow stromal cells which are cultured by autologous bone marrow, autologous serum and autologous cerebrospinal fluid. The method pertains to the methods for differentiating the bone marrow stem cells into the neural stem cells. A conventional low-sugar DMEM / F 12 culture medium is adopted, the autologous serum replaces the original fetal bovine serum, the autologous cerebrospinal fluid replaces the original additive and a stimulating factor, a small amount of autologous bone marrow is used for culturing more cells with the number that can meet the clinical needs. The method has the advantages of low cost and easy material selection, avoiding zoonosis and foreign protein exclusive reaction, and so on. The method of inducing the bone marrow stromal cells into the neural stem cells by using cerebrospinal fluid can effectively solve the shortcomings of the traditional methods.

The invention discloses a method for preparing CIK cells with a killing effect on tumor cells. The CIK cells are obtained through induced culture of 17-20 days by using a ficoll mononuclear cell separation medium with a density of 1.084 in the PBS system to separate mononuclear cells in peripheral blood or umbilical cord blood. According to the present invention, mononuclear cells are separated and obtained efficiently by using the ficoll density gradient centrifugation method, and a sufficient amount of CIK is obtained by using cell culture bags and a CIK cell culture system to meet the needs of clinical treatment. The Takara lymphocyte culture medium and the autologous serum and cytokine co-culture technique are used in the method, thereby preventing application of fetal bovine serum, reducing contamination risks of exogenous pyrogen and sensitinogen, and while maintaining the advantage of CIK cell efficient proliferation. The cell culture bag technology is used to reduce risk of cell contamination, and is suitable for clinical therapeutic application.

Certain embodiments here in are directed to a method of treating a tissue associated with a defect in a human including wrinkles, rhytids, depressed scar, cutaneous depressions, stretch marks, hyperplasia of the lip, nasolabial fold, melolabial fold, scarring from acne vulgaris, and post-rhinoplasty irregularity. The tissue defect may be treated by introducing a plurality of in vitro cultured autologous fibroblast cells at or proximal to the defect area of the patient's tissue. The autologous fibroblast cells may have been cultured in vitro to expand the number of fibroblast cells in at least one medium that comprises autologous serum. The autologous fibroblast cell cultures may be derived from connective tissue, dermal, fascial fibroblasts, papillary fibroblasts, and / or reticular fibroblasts.

The present invention provides methods for culturing primary cells and tissues from a subject in the presence of the subject's own serum, ascites or pleural effusion fluid. Methods of treating cancer, and screening for the effectiveness or toxicity of drugs are also provided herein.

The invention discloses a method for freezing DC cells and CIK seed cells in blood, and the prepared cells and their application. The mononuclear cells in peripheral blood or umbilical cord blood are frozen in autologous serum after appropriate treatment, and the cell The cryoprotectant makes the survival rate of the cells reach 80%. The resuscitated cells can be used to prepare DC-cells and CIK cells at the same time, and finally prepare DC-CIK cell preparations that can meet the needs of clinical applications. In this way, the requirement of multiple DC-CIK clinical treatments can be realized in a single blood collection, thus providing support for the large-scale development of immune cell therapy.

A medium for stem cell in regeneration of intervertebral disc comprising an autologous serum obtained by sterilization and immobilization of serum of an individual having the intervertebral disc to be regenerated, a medium for cell culture and at least one antibiotic and a method for regenerating an intervertebral disc comprising suspending, in the above-mentioned medium or embedding in a cell carrier, stem cells collected from the subject individual or a homogenous or heterogenous individual, either directly or after culturing the same using the medium for regenerating the stem cell for regenerating intervertebral disc; and then transplanting the suspension or the embedded carrier of the stem cells into the nucleus pulposus cavity of the intervertebral disc.

The invention relates to the technical field of cell cryopreservation solutions, and in particular relates to a cryopreservation solution for the cultured NKT cells and a preparation method of the cryopreservation solution. The cryopreservation solution for the cultured NKT cells comprises lentinan, algal polysaccharides, autoserum and DMSO. By virtue of adding the autoserum, the pollution of foreign animal viruses and the introduction of foreign proteins are avoided, and the safety is high; by virtue of adding the lentinan and the algal polysaccharides, the activity of the cells can be effectively retained, and no harm to a human body is generated; meanwhile, the state of the cells recovered from cryopreservation is basically close to the state before cryopreservation, and the cryopreservation effect is good.

Certain embodiments here in are directed to a method of treating a tissue associated with a defect in a human including wrinkles, rhytids, depressed scar, cutaneous depressions, stretch marks, hyperplasia of the lip, nasolabial fold, melolabial fold, scarring from acne vulgaris, and post-rhinoplasty irregularity. The tissue defect may be treated by introducing a plurality of in vitro cultured autologous fibroblast cells at or proximal to the defect area of the patient's tissue. The autologous fibroblast cells may have been cultured in vitro to expand the number of fibroblast cells in at least one medium that comprises autologous serum. The autologous fibroblast cell cultures may be derived from connective tissue, dermal, fascial fibroblasts, papillary fibroblasts, and / or reticular fibroblasts.

The invention relates to a clinical applicable culture system for efficient amplification of NK cells. The invention specifically provides a serum-free culture system for efficient amplification of NK cells. The culture system comprises the following components: 1640 medium, sodium bicarbonate, human serum albumin, transferrin, linoleic acid, oleic acid, palmitic acid, sodium pyruvate, human recombinant insulin, non-essential amino acids, 2-mercaptoethanol and interleukin-2. The invention also provides a culture system containing 1-12 vol.% of human autoserum and for efficient amplification of NK cells. The culture system provided by the invention has simple composition, avoids the risks of animal ingredients in animal serum on cell treatment and uncertainty of cell culture caused by the uncertain components in the animal serum, and increases yield of NK cell. The obtained activated NK cells have significant tumor killing effect. Therefore, the system can not only be applied in scientific research but also be used in clinical treatment.

The invention provides a culture and cryopreservation method for a prostatic cancer organoid with a tumor immune microenvironment. The culture and cryopreservation method comprises the following steps: separating mononuclear cells and autoserum from peripheral blood, and stimulating the mononuclear cells by using a CD28 antibody and IL-2; separating tumor cells from a prostatic cancer tissue sample; mixing the tumor cells with a culture medium and a temperature-sensitive hydrogel, and then carrying out culturing and subculturing to obtain a tumor organoid precursor; stimulating the tumor organoid precursor by using IFN-gamma, and then conducting dissociating to form stimulated tumor cells; mixing the stimulated tumor cells with the stimulated mononuclear cells, conducting stimulating witha PD-1 antibody, mixing the stimulated mixture with the temperature-sensitive hydrogel, and carrying out culturing; and cryopreserving the tumor organoid with a cryopreservation solution containing FBS and the temperature-sensitive hydrogel. With the method in the invention, the success rate of culturing of organoids with a tumor microenvironment can be greatly improved at low cost.

The invention discloses a method for preparing an autologous triple skin fibroblast preparation. The autologous triple skin fibroblast preparation comprises platelet-rich plasma, autologous collagen concentrate and fibroblasts, wherein the ratio of platelet-rich plasma to the autologous collagen concentrate is 1 to 4; and the concentration of autologous fibroblasts is (1.0-2.0)*10<7> / mL. The preparation method comprises the following steps: separating and culturing human skin fibroblast; preparing the high-concentration autologous collagen solution; preparing autologous platelet-rich plasma; and preparing an autologous triple skin fibroblast injection preparation. Active ingredients of the preparation disclosed by the invention are all from a customer, and are cultured in an autologous serum medium during the cultivation of fibroblasts, so that the preparation is truly safe, effective and individualized. The preparation can be used for skin wrinkle removal, scar repairing, regeneration and other effects, so as to solve a problem of skin aging.

The invention discloses a method for proliferating NK cells in vitro by virtue of magnetic beads coupled with multiple stimulating proteins. The method comprises the following steps: culturing NK cells by adopting an NK cell culture medium, adding magnetic beads coupled with IL-15, 4-1BBligand and IL-21 (of which the molar ratio is (1-3):(1-3):(1-3)), culturing for 12-15 days, and harvesting and washing the NK cells, wherein the NK cell culture medium is prepared by adopting a method comprising the step of adding 10% of human AB serum or autologous serum, 500IU / mL of IL-2 and 5ng / ml of IL-15 into an X-VIVO 20 basic culture medium. The method disclosed by the invention has the advantages that the magnetic beads are coupled with three specificity stimulating factors, the aim of stimulating the NK cells by multiple factors is achieved, and usage of a trophoblastic cell line is also avoided, so that GMP production requirement and amplification efficiency requirement are considered at the same time, and the harvested NK cells can be used for NK allotransplantation.

The invention discloses a method for preparing adipose-derived stem cells by means of extraction. The method comprises the following steps: acquiring normal abdominal subcutaneous adipose tissue, washing the obtained adipose tissue with a phosphate buffer solution (PBS), and then shearing the adipose tissue into pieces; then, carrying out digestion on the adipose tissue for a period of time by using type I collagenase; after digestion, adding the PBS into the product, evenly mixing and filtering; adding a DMEM culture solution containing human autologous serum, putting into a culture box for culturing, and replacing the culture solution with a fresh culture solution at regular intervals, wherein the cells growing from the tissue are primary cells; when the primary cells reach a certain degree of fusion, washing the cells with the PBS again, and then adding pancreatin and ethylene diamine tetraacetic acid (EDTA) digestive juice; after the cells suspend, adjusting the density of the cells, and adding the DMEM culture solution containing the human autologous serum into the cells again to obtain second-generation cells; and when the second-generation cells reach a certain degree of fusion, adding normal saline into the cells, transferring the cells into a centrifuge tube for carrying out centrifugal filtration, taking supernatant, concentrating and extracting to obtain the adipose-derived stem cells. The adipose-derived stem cells extracted by the method contain multiple life active substances, so that the stem cell properties of the adipose-derived stem cells are further improved.

The invention relates to a composition for promoting hair growth. The composition includes lyophilized platelet-rich plasma, and further includes one or more of autologous serum, immune cells and vascular matrix components, and further includes one or more of a hair growth nutrient solution and normal saline. The composition is a non-surgical safe and effective hair growth composition, and is usedfor long term without obvious side effects, from the perspective of cytology and biotechnology, growth factors, nutritional components, immune cells and the like are provided, multi-angle, multi-level and multi-dimension improvement of scalp microenvironment can be improved, scalp local microcirculation is promoted, hair follicle stem cells and hair matrix cells are activated, the hair abnormal resting period is shortened, the secretion of sebaceous glands is effectively inhibited, the physiological function of hair follicles is improved, and quick-acting, intermediate-acting, and long-actingpromotion of hair continuous regeneration is achieved.

The invention discloses a peripheral blood NKT cell culture solution and a culture method. The NKT cell culture method comprises the following steps of adding peripheral blood mononuclear cells into a lymphocyte serum-free culture medium of autoserum containing cytokines, and culturing for about 14 days; in the culture period, a culture solution containing cell factors needing to be added every 2-3 days; after culture is finished, performing centrifugal treatment and cleaning with a 0.9% sodium chloride solution to obtain peripheral blood NKT cells; adding human serum albumin into the peripheral blood NKT cells, and fixing the volume by using a 0.9% sodium chloride solution to obtain the peripheral blood NKT cells. According to the culture method, a peripheral blood single core has no immunological rejection reaction on autoserum, cells are purer, and the safety is higher; the NKT cells are fast in growth and high in amplification rate; and the NKT cell killing activity is high, and a relatively strong tumor cell killing effect is achieved.

The process in general is based on taking a sample of skin and a sample of blood from the patient and based on these two elements skin is cultured, being placed on a collagen patch to produce a dressing which is subsequently placed on the patient requiring same.

The invention relates to a culture method for rhesus peripheral blood mononuclear macrophages. The method specifically comprises the following steps: separating peripheral blood mononuclear cells, resuspending and washing the collected peripheral blood mononuclear cells by a PBS (Phosphate Buffer Solution) twice, recovering the washed cells through centrifuging, centrifuging at a speed of 1200rpm for 8min each time, finally resuspending by an RPMI1640 culture solution containing 2-4% of rhesus autologous serum, 1% of mycillin and 0.1% of HEPES, controlling the density of the resuspended cells at 3*106 cells / ml, immediately adding into a 96-pore culture plate or a 48-pore culture plate of a CELLBIND Surface, culturing at a density of 0.8*106 cells per pore or 3*106 cells per pore, washing away suspended cells by the RPMI1640 culture solution after 24h, then continuously culturing adherent cells by a full culture medium containing 2-4% of rhesus autologous serum, changing the culture solution every two to three days, observing the cell morphology after seven days, and judging the cell differentiation result. The rhesus mononuclear macrophages obtained through differentiation culture are high in purity and have the typical macrophage morphology and immunology properties. The method is simple, economical and effective.

The present invention provides methods for culturing primary cells and tissues from a subject in the presence of the subject's own serum, ascites or pleural effusion fluid. Methods of treating cancer, and screening for the effectiveness or toxicity of drugs are also provided herein.

Certain embodiments herein are directed to a method of treating a tissue associated with a defect in a human including wrinkles, rhytids, depressed scar, cutaneous depressions, stretch marks, hyperplasia of the lip, nasolabial fold, melolabial fold, scarring from acne vulgaris, and post-rhinoplasty irregularity. The tissue defect may be treated by introducing a plurality of in vitro cultured autologous fibroblast cells at or proximal to the defect area of the patient's tissue. The autologous fibroblast cells may have been cultured in vitro to expand the number of fibroblast cells in at least one medium that comprises autologous serum. The autologous fibroblast cell cultures may be derived from connective tissue, dermal, fascial fibroblasts, papillary fibroblasts, and / or reticular fibroblasts.

The invention relates to application of a clinical applicable culture system for efficient amplification of NK cells. The invention specifically provides application of a serum-free culture system for amplification of NK cells. The culture system comprises the following components: 1640 medium, sodium bicarbonate, human serum albumin, transferrin, linoleic acid, oleic acid, palmitic acid, sodium pyruvate, human recombinant insulin, non-essential amino acids, 2-mercaptoethanol and interleukin-2. The invention also provides application of a culture system containing 1-12 vol.% of human autoserum and for amplification of NK cells. The culture system provided by the invention has simple composition, avoids the risks of animal ingredients in animal serum on cell treatment and uncertainty of cell culture caused by the uncertain components in the animal serum, and increases yield of NK cell. The obtained activated NK cells have significant tumor killing effect. Therefore, the system can not only be applied in scientific research but also be used in clinical treatment.

The invention provides a culture medium for in-vitro amplification of autologous human hair follicle mesenchymal stem cells and a culture method of the culture medium, and belongs to the technical field of cell culture. The culture medium for in-vitro amplification of the autologous human hair follicle mesenchymal stem cells takes DMEM / F-12 as a basic culture medium and is prepared from the following components in percentage by volume: 4 to 6 percent of autologous serum, 9 to 11ng / ml of bFGF and 0.8 to 1.2 percent of penicillin-streptomycin. According to the culture method, human autoserum isused as an additive to prepare an amplification culture medium, and in-vitro large-scale amplification of autologous human hair follicle mesenchymal stem cells is carried out. Compared with the traditional mesenchymal stem cells, the hair follicle mesenchymal stem cells cultured by the method have the advantages that other animal serum or allogeneic blood products are not required to be added, thesafety is high, the allogeneic disease propagation risk is avoided, the cell proliferation rate is high, and the characteristics of the mesenchymal stem cells can still be maintained after multiple passages.

The invention discloses a cryopreservation liquid for autologous umbilical cord mesenchymal stem cells and its preparation and cryopreservation method. The cryopreservation liquid is composed of liquid A and liquid B used together, and the volume ratio of liquid A to liquid B is 6-9: 1-3; the A solution is calculated in parts by volume, and includes the following components: 65-75 parts of basal medium containing pre-supplements; the ratio of pre-supplements to basal medium is 7-12mg: 0.6-0.9mL; Wherein, the pre-additives are calculated in parts by weight, including: 4-6 parts of Morinda officinalis alcohol extract, 0.13-0.16 parts of type I collagen, 0.05-0.15 parts of sodium hyaluronate and 3-5 parts of lipoic acid; Liquid B is calculated in parts by volume and includes the following components: 15‑25 parts of autologous serum. Using the cryopreservation solution of the present invention can simultaneously obtain a sufficient amount of blood for the separation of autologous serum during the delivery of newborns, thereby effectively avoiding the contamination of serum proteins from heterogeneous allogeneic sources, and at the same time simulating the microenvironment of the extracellular matrix to improve the survival rate of cell resuscitation, Therefore, it is beneficial to its application in the standardized operation of biological sample banks and clinical cell therapy.