27 results about "Cryopreserved Cell" patented technology

The invention belongs to the technical field of cell cryopreservation, and relates to a cryopreservation method based on ultrasonic ice planting. In the method, the nucleation temperature of ice planting is less than-9 DEG C, and the cooling rate from nucleation to icing is less than 5 DEG C / min. Compared with the prior art, the method has the advantages that the time required for cryopreservation of the cells is remarkably shortened, meanwhile, damage to the cells caused by ice crystals generated in the cooling process is reduced, and the problem that an existing cell cryopreservation cooling method cannot meet the nucleation and freezing uniformity of a cell solution in the cooling process is solved.

Provided are methods for the creation of ultra-high density cryopreserved cell banks. In certain embodiments, these methods employ altered perfusion culture techniques that allow for production of ultra-high density cell cultures that can be cryopreserved at unexpectedly high cell densities without the need for any cell concentration steps, while retaining excellent cell viability and quality.

The invention discloses a novel antifreeze polypeptide with a clear structure and excellent performance and a protection method of cryopreserved cells. The core structure of the polypeptide is formed by covalently connecting two functional modules, the N end is a unit with ice crystal binding performance, the C end is a super-hydrophilic unit, and the two functional modules are bridged through a flexible linker. Experimental verification shows that the synthetic polypeptide shows excellent anti-freezing activity, can significantly inhibit nucleation, growth and recrystallization processes of ice crystals, and has particularly outstanding ice recrystallization inhibition activity. The novel antifreeze polypeptide shows excellent biocompatibility, and is suitable for low-temperature cryopreservation of cells sensitive to freezing damage and other living microbial preparations. Results show that the polypeptide can greatly improve the freeze-drying survival rate of cells and the activity of the cells after recovery, is non-toxic to the cells, and shows the potential of replacing a traditional cryoprotectant. Therefore, the method has wide application prospects in food industry, biological medicine, agricultural breeding and low-temperature preservation technologies of biological sample libraries.

The invention relates to the technical field of cell biology, in particular to an iNKT cell cryopreservation solution and a method for cryopreserving iNKT cells by using the cryopreservation solution. The invention provides an immune cell cryopreservation solution. The immune cell cryopreservation solution comprises human serum albumin, a basic low-temperature protection solution and an optional human platelet lysate. The immune cell freezing medium does not contain animal-derived components except for human bodies, and has the advantages of simple components and good stability. According to the cryopreservation liquid disclosed by the invention, through the combined action of the basic low-temperature protection liquid (such as CryoStor10 (CS10)) and the human serum albumin, the motility rate after cell recovery is improved, and the damage to immune cells (such as iNKT) in the cryopreservation process is reduced. After a small amount of human platelet lysate is further added, the survival rate of cell resuscitation and the amplification capacity can be further improved.

The invention relates to the technical field of biology, in particular to a cryopreserved stem cell diluent and application thereof.The cryopreserved stem cell diluent further comprises cysteine and polyvinylpyrrolidone besides a compound electrolyte injection, the concentration of cysteine in the cryopreserved stem cell diluent is 1-10 mM, and the concentration of polyvinylpyrrolidone in the cryopreserved stem cell diluent is 1-10 mM. The concentration of the polyvinylpyrrolidone in the cryopreserved stem cell diluent is 0.02 to 0.1 g / mL. The cryopreserved stem cell diluent can improve the cell activity and cell distribution after the cryopreserved cell preparation is recovered, and can monitor the application of cells in rat living body imaging.

The invention provides a method for quantitatively detecting mitochondrial integrity after cell cryopreservation recovery. The method comprises the following steps: dyeing cells to be detected by adopting a dyeing working solution to obtain dyed cells; the dyeing working solution comprises a JC-10 dyeing working solution or a JC-1 dyeing working solution; mixing the dyed cells with a cryopreservation solution for cryopreservation to obtain cryopreserved cells; wherein the cryopreservation liquid does not contain DMSO (Dimethylsulfoxide) and protein; and after the cryopreserved cells are recovered, directly detecting the mitochondrial integrity of the recovered cells through a dual-emission fluorescence detection method without washing. According to the method for quantitatively detecting the mitochondrial integrity after cell cryopreservation recovery, the cryopreservation liquid which does not contain DMSO and protein is adopted, so that the influence of the cryopreservation liquid on the coloring agent is avoided. The cryopreservation is performed after dyeing, and detection can be directly performed after resuscitation, so that interference caused by mitochondria damage in operation steps such as osmotic pressure change and centrifugation is avoided, and the detection result is more accurate.

The disclosure provides for cryopreservation compositions and methods of use thereof. Aspects of the present disclosure provide for cryopreservation compositions useful in the freezing and thawing of cell therapies. Other aspects of the present disclosure provide for systems for use in the cryopreservation cell therapies. Still other aspects of the present disclosure provide for methods of cryopreserving cell therapies.

The invention discloses an infrared cell resuscitation instrument which comprises a shell and an upper cover movably connected with the shell, an interaction module is arranged on the shell, a control module and one or more resuscitation cavities are arranged in the shell, and an infrared heating module and a temperature sensing module are arranged on the inner wall of each resuscitation cavity. The infrared cell resuscitator is based on an infrared heating principle, does not need to take water as a heat-conducting medium, and can realize efficient cryopreserved cell resuscitation which is not easy to pollute. The infrared cell resuscitation instrument can be further provided with a fan, an optical transmitting unit and an optical receiving unit, so that the cell resuscitation efficiency is improved, and the cell resuscitation effect can be effectively improved.

The portable self-adaptive multi-specification cryopreserved cell water bath resuscitation device comprises a folding water bath tank and an intelligent buckle cover covering the folding water bath tank, the bottom of the folding water bath is connected with a lining heating film, and the outer periphery of the folding water bath is connected with a plurality of memory metal brackets for folding the folding water bath; the interior of the intelligent buckle cover is connected with a clamping matrix plate layer through a 3D multi-degree-of-freedom rocker arm module. The clamping matrix plate layer is connected with a plurality of sets of hydraulic clamping jaws used for clamping cryopreservation bottles. A distance measuring and temperature measuring module is further connected to the lower portion of the clamping matrix plate layer. The device is convenient to carry, and the 3D multi-degree-of-freedom rocker arm module is adopted to shake a cryopreservation bottle.

The utility model discloses a programmed cooling device for cryopreserved cells. The programmed cooling device is characterized in that an inner container structure is arranged in a box body; a groove is formed in the inner container; the cryopreservation tube frame is arranged in the groove; a plurality of tube grooves for placing cryopreservation tubes are formed in the cryopreservation tube rack; the flip cover is hinged to one side of the box body. According to the cryopreservation tube, the tube sleeve is arranged in each tube groove, so that an independent temperature protection sleeve device is provided for the cryopreservation tube. When the cryopreservation tube is taken out from a low-temperature environment, the tube sleeve can effectively slow down heat exchange between the cryopreservation tube and an external normal-temperature environment, the phenomenon that cells are repeatedly frozen and thawed due to rapid temperature rise is avoided, and the survival rate and activity of the cells are remarkably improved.

The invention discloses a method for accurately determining the motility rate of MSC cells, and belongs to the technical field of motility rate detection of mesenchymal stem cells (MSC). The method mainly comprises the following steps: S1, taking out cryopreserved cells from liquid nitrogen, placing the cryopreserved cells in dry ice, transferring the cryopreserved cells to a laboratory, and carrying out unfreezing resuscitation in a 37 DEG C water bath; s2, taking recovered cells (3E6Cell), averagely transferring the recovered cells (3E6Cell) into three 1.5 ml EP tubes (1E6Cell / tube), adding 1ml of LDPBS, carrying out centrifugation for 5 min, discarding the supernatant, repeating the step once, and then carrying out resuspension by using 100 [mu] l of DPBS for later use; s3, dye preparation: taking one EP tube, adding 400 [mu] l of Annexin V Loading buffer, 10 [mu] l of Annexin V and 10 [mu] l of 7AAD into the EP tube, and carrying out vortex uniform mixing for later use; s4, dyeing: adding 100 microliters of prepared dye into 100 microliters of prepared sample, lightly and uniformly mixing, and incubating in a dark place for 10 minutes; s5, after the dyeing is finished, adding 100 [mu] l of Annexin V loading buffer into each tube, uniformly mixing, and carrying out on-machine detection on a flow cytometer; and S6, taking the average value of the three times of detection data for reporting. The method for accurately determining the cell viability of the MSC achieves the effect of reliable cell counting.

The present disclosure provides, among other things, a method of cryopreserving and thawing cells that results in the thawed cells having high cellular viability and functionality post-thawing. In some embodiments, a large-scale method of cryopreserving cells is provided, the method comprising: (a) contacting the cells with a cryopreservation medium; (b) cooling the cells to −80° C. at a controlled rate to minimize latent heat of fusion; and (c) storing the cells in liquid nitrogen vapor phase, thereby cryopreserving the immune cells.

The application discloses a Super-long fragment DNA extraction kit and application. The Super-long fragment DNA extraction kit is universal, can solve the DNA super-long fragment extraction of culture cells, cryopreserved cells, fresh animal tissues, cryopreserved animal tissues, common crops, polysaccharide and polyphenol plants and part of silica gel dry plant samples and the like, can guarantee the purity under the condition that the length reaches 500kb-1Mb, and is suitable for long read length sequencing with high requirements for the DNA fragment length and the DNA purity.

The invention relates to the technical field of cell culture, and discloses a serum-free and DMSO-free human umbilical cord mesenchymal stem cell cryopreservation solution. The invention discloses a serum-free and DMSO (dimethyl sulfoxide)-free human umbilical cord mesenchymal stem cell cryopreservation solution. The serum-free and DMSO-free human umbilical cord mesenchymal stem cell cryopreservation solution is prepared from the following components: ethylene glycol with the final concentration of 1000 to 1500 mmol / L, trehalose with the final concentration of 50 to 150 mmol / L, proline with the final concentration of 50 to 150 mmol / L and 5 to 15 percent of human serum albumin injection. The invention further discloses application of the cell cryopreservation liquid in cryopreservation of cells. The serum-free and DMSO-free human umbilical cord mesenchymal stem cell cryopreservation liquid disclosed by the invention does not contain a dimethyl sulfoxide (DMSO) component and a basic culture medium component, the components are clear, the batch is stable, and the serum-free and DMSO-free human umbilical cord mesenchymal stem cell cryopreservation liquid meets the industrial transformation requirements. Besides, the activity and dryness of the cells can be stably maintained for a long time when the cells are cryopreserved, the recovered cells can be directly used for clinical application, and the safety requirement and the effectiveness requirement of the clinical application are met.

Provided is a method for cryopreserving cells, which is characterized in that the cells are cryopreserved for 30 days or more in a state in which the cells are inoculated on a porous membrane without using liquid nitrogen. Also provided is a cell culture method comprising a step for cryopreserving cells inoculated on the porous membrane for 30 days or more without using liquid nitrogen. Further provided is a cryopreservation base material comprising a porous membrane for cryopreservation of inoculated cells for 30 days or more.

The present invention relates to a frozen cell formulation that can significantly improve the cell viability of cryopreserved cells, preferably stem cells. According to the present invention, the cell viability can be improved when thawing cells after freezing, thereby preventing a decrease in therapeutic efficiency that may occur during the storage and transportation of cell therapy agents.

The presently disclosed subject matter relates to compositions of ready-to-use cryopreserved populations of dissociated cells which can be directly used for downstream application without after-thaw expansion and / or passage. The presently disclosed subject matter also provides for methods of preparing such compositions, and in vitro methods of using such compositions.

The present disclosure provides, among other things, a method of cryopreserving and thawing cells that results in the thawed cells having high cellular viability and functionality post-thawing. In some embodiments, a large-scale method of cryopreserving cells is provided, the method comprising: (a) contacting the cells with a cryopreservation medium; (b) cooling the cells to −80° C. at a controlled rate to minimize latent heat of fusion; and (c) storing the cells in liquid nitrogen vapor phase, thereby cryopreserving the immune cells.

The invention belongs to the field of cell culture, and particularly relates to a sorting method of cryopreserved cells. The method comprises the following steps: sequentially adding nuclease and sorting magnetic beads into resuscitated cryopreserved cells, and incubating and centrifuging to obtain sorted target cells; wherein the steps of incubation and centrifugation are not carried out between nuclease addition and magnetic bead separation. According to the method, the nuclease treatment step and the cell sorting step are combined, the whole steps are simple, the sorted cell yield is high, and the state after culture is good.

The invention relates to a cell cryopreservation solution which is suitable for cryopreservation of animal cells. The compound electrolyte comprises a compound electrolyte solution, a human serum albumin solution and dimethyl sulfoxide (DMSO). The cell cryopreservation liquid has good biocompatibility, the survival rate and functionality of cells under the low-temperature condition are remarkably improved, and the cell cryopreservation liquid is particularly suitable for cryopreservation and recovery of immune cells and stem cells (such as natural killer cells, T cells and mesenchymal stem cells). The cryopreservation liquid not only can be used for in-vitro cryopreservation of cells, but also can be directly applied to a clinical tested object together with the cryopreserved cells.

The application discloses a universal DNT cell public library, a DNT cell preparation and a preparation method thereof, 200 mL of peripheral blood of a healthy adult is separated to obtain mononuclear cells, the mononuclear cells are cultured for 12 days by using zoledronate in vitro, and cell surface markers are detected; when the proportion of CD3+CD4-CD8- is greater than 50%, the cells are frozen to obtain DNT public library cells; when needed, the DNT public library cells are co-cultured with zoledronate for 7 days, and a DNT cell preparation with high purity and high quality can be obtained. The application avoids the ethics and operation burden of repeatedly collecting blood of a donor, reduces production cost and risk of dependence on the donor, has a short preparation period, can support multi-batch production, and has stable quality between batches, and can provide sufficient raw material reserves for clinical trials and patients with diseases.

The invention relates to the technical field of stem cell quality detection, in particular to a method for evaluating the quality of an umbilical cord mesenchymal stem cell preparation, which comprises the following steps: S1, screening cord blood infectious disease markers and ALT; s2, detecting the motility rate, the form, the 24-hour dynamic phenotype before cryopreservation / after resuscitation of the P2-generation cryopreserved cells, the multidirectional differentiation potential, double pollution of a blood culture instrument and fluorescent quantitative PCR, and the like; s3, detecting clinical adaptive cytokines (such as BMP-2 and NGF) and inflammatory factors of the issued cells; and S4, performing graded packaging (with built-in temperature sensitive labels) according to clinical requirements, and performing temperature-controlled cryopreservation and transportation. A full-dimensional quality control system is constructed, the functional activity, stability and safety of the preparation are guaranteed, clinical requirements are met, and safe clinical conversion of UC-MSCs is assisted.

The invention discloses a peripheral blood mononuclear cell cryopreservation protective agent and a cryopreservation method.The cryopreservation protective agent is prepared from, by volume, 5% of dimethyl sulfoxide (DMSO), 5% of dextran, 45% of autologous plasma and 45% of a serum-free culture medium. The method comprises the following steps: centrifuging a negative peripheral blood sample for infectious disease detection, counting mononuclear cells, preparing cryopreserved cell sap, carrying out programmed cooling, and transferring to low-temperature storage. The cryopreservation liquid has the advantages that the cryopreservation liquid can protect cells, and the effectiveness and the stability of the motility rate of the cryopreserved cells after recovery each time are ensured. The cell viability after recovery is high and can reach 95% or above, no heterologous animal component exists, and high-quality cryopreserved peripheral blood mononuclear cells are provided for recovery culture of immune cells later.

The invention relates to a non-cryopreserved cell preservation solution and application thereof. The cell preservation solution comprises a basic solution and a core additive, wherein the core additive comprises a ferroptosis inhibitor and a mitochondrial protective agent, based on an intracellular fluid type ionic environment and multi-mechanism synergy, an optimized basic fluid system with high potassium, low sodium and histidine buffering is adopted, the ferroptosis inhibitor and the mitochondrial targeted protective agent are integrated, and by simulating an intracellular environment, stabilizing pH and inhibiting a key death pathway, the activity of the ferroptosis inhibitor is improved; the survival rate and the functional activity of various cells under the low-temperature non-cryopreservation condition are remarkably improved, and the method has important application significance in the field of cell preservation.

The present application provides a clinical level DMSO-free, serum-free and any protein component-free mesenchymal stem cell cryopreservation solution, which can be used for long-term preservation of mesenchymal stem cells in scientific research and clinical practice, such as establishment of mesenchymal stem cell bank, etc. Since the cryopreservation solution does not contain DMSO, serum and any protein and other animal-derived components, it is safer and more suitable for clinical needs. In particular, there is no need for programmed cooling when freezing cells, which simplifies the freezing process and unexpectedly improves the recovery rate and viability of the frozen cells.

This invention provides a cell cryopreservation method that avoids or improves upon the use of DMSO as a cryoprotective agent. [Solution] Cell material containing living cells is preserved during a cryopreservation protocol by combining the cell material with a cryoprotective formulation / medium / solution containing an effective amount of trehalose (in the absence of DMSO and / or any other added cryoprotective agents). That is, the cryopreservation protocol does not contain any cryoprotective agents other than trehalose, and the cryopreservation protocol includes exposing the cell material to a cryoprotective formulation containing an effective amount of trehalose to act as a cryoprotective agent, cooling the cell material to a predetermined temperature below -20°C at a cooling rate in the range of -3°C / min to -50°C / min, and obtaining warmed cryopreserved cell material.

The present disclosure provided methods and compositions for generating a triple cell culture comprising neurons, astrocytes, and microglia. The method includes providing neurons, astrocytes, and microglial cells or precursors, and culturing them under conditions that support survival, maturation, and functional interaction. In some embodiments, the cells are derived from pluripotent stem cells and seeded in defined ratios. The cultures are suitable for modeling central nervous system function, disease processes, and for evaluating candidate therapeutic agents. Also disclosed are variations of the culture system including use of reporter genes, cryopreserved cell components, three-dimensional formats, and high-throughput assay configurations.