The present invention discloses an in-vitro culture, induction, activation and cryopreservation method and cell bank establishment for immune cells. The method includes the follows: using a dedicated amplification medium of immune cells to perform first-stage amplification culture on mononuclear cells to obtain preliminarily amplified immune cells; using a dedicated induction medium of immune cells to perform second-stage induction and amplification culture on the preliminarily amplified immune cells to obtain induced immune cells; using a dedicated activation medium of immune cells to perform third-stage activation and amplification culture on the induced immune cells to obtain a large number of immune cells with activation functions; using a dedicated cryopreserving fluid of immune cells to cryopreserve the immune cells to obtain cryopreserved immune cells; and performing preservation according to ABO/RH typing and HLA typing; and establishing an information file of immune cells for retrieval to construct an immune cell bank.

The present invention discloses an in-vitro culture, induction, activation and cryopreservation method and cell bank establishment for immune cells. The method includes the follows: using a dedicated amplification medium of immune cells to perform first-stage amplification culture on mononuclear cells to obtain preliminarily amplified immune cells; using a dedicated induction medium of immune cells to perform second-stage induction and amplification culture on the preliminarily amplified immune cells to obtain induced immune cells; using a dedicated activation medium of immune cells to perform third-stage activation and amplification culture on the induced immune cells to obtain a large number of immune cells with activation functions; using a dedicated cryopreserving fluid of immune cells to cryopreserve the immune cells to obtain cryopreserved immune cells; and performing preservation according to ABO/RH typing and HLA typing; and establishing an information file of immune cells for retrieval to construct an immune cell bank.

Provided in the present invention is a cell freezing medium for clinical use. In particular, the cell freezing medium of the present invention comprises the following components: (1) human albumin; (2) cryoprotectant: the cryoprotectant comprises a combination of one or more of dimethyl sulfoxide, glycerol, and ethylene glycol; (3) a saline buffer; wherein the salt buffer is a solution containing Na.sup.+, K.sup.+, Mg.sup.+, Cl.sup., and CH.sub.3COO.sup. ions; (4) a vitamin; and (5) an amino acid, wherein the human albumin concentration is 1%-20% (w/v). The cell, after long-term cryopreservation with the freezing medium of the present invention, has a high viability, and the cellular efficiency maintains a high uniformity. The grade of purity of the freezing medium of the present invention is the pharmaceutical grade or USP grade; and the freezing medium is safe and reliable for clinical use, and can be used or conventional adherent and suspension cells.

Provided in the present invention is a cell freezing medium for clinical use. In particular, the cell freezing medium of the present invention comprises the following components: (1) human albumin; (2) cryoprotectant: the cryoprotectant comprises a combination of one or more of dimethyl sulfoxide, glycerol, and ethylene glycol; (3) a saline buffer; wherein the salt buffer is a solution containing Na.sup.+, K.sup.+, Mg.sup.+, Cl.sup., and CH.sub.3COO.sup. ions; (4) a vitamin; and (5) an amino acid, wherein the human albumin concentration is 1%-20% (w/v). The cell, after long-term cryopreservation with the freezing medium of the present invention, has a high viability, and the cellular efficiency maintains a high uniformity. The grade of purity of the freezing medium of the present invention is the pharmaceutical grade or USP grade; and the freezing medium is safe and reliable for clinical use, and can be used or conventional adherent and suspension cells.

Human induced pluripotent stem cells (hiPSCs) possess tremendous potential for tissue regeneration and banking hiPSCs by cryopreservation for their ready availability is crucial to their widespread use. However, contemporary methods for hiPSC cryopreservation are associated with both limited cell survival and high concentration of toxic cryoprotectants and/or serum. The latter may cause spontaneous differentiation and introduce xenogeneic factors, which may compromise the quality of hiPSCs. Here, sand from nature is discovered to be capable of seeding ice above 10 C., which enables cryopreservation of hiPSCs with no serum, minimized cryoprotectant, and high cell survival. Furthermore, the cryopreserved hiPSCs retain high pluripotency and functions judged by the pluripotency marker expression, cell cycle analysis, and capability of differentiation into the three germ layers. This unique sand-mediated cryopreservation method may greatly facilitate the convenient and ready availability of high-quality hiPSCs and probably many other types of cells/tissues for the emerging cell-based translational medicine.

Human induced pluripotent stem cells (hiPSCs) possess tremendous potential for tissue regeneration and banking hiPSCs by cryopreservation for their ready availability is crucial to their widespread use. However, contemporary methods for hiPSC cryopreservation are associated with both limited cell survival and high concentration of toxic cryoprotectants and/or serum. The latter may cause spontaneous differentiation and introduce xenogeneic factors, which may compromise the quality of hiPSCs. Here, sand from nature is discovered to be capable of seeding ice above 10 C., which enables cryopreservation of hiPSCs with no serum, minimized cryoprotectant, and high cell survival. Furthermore, the cryopreserved hiPSCs retain high pluripotency and functions judged by the pluripotency marker expression, cell cycle analysis, and capability of differentiation into the three germ layers. This unique sand-mediated cryopreservation method may greatly facilitate the convenient and ready availability of high-quality hiPSCs and probably many other types of cells/tissues for the emerging cell-based translational medicine.

Disclosed herein are methods of detecting a target nucleic acid sequence, determining the localization of the target nucleic acid sequence, and/or quantifying the number of target nucleic acid sequences in a cell. This method may be used on small target nucleic acid sequences, and may be referred to as Nano-FISH.

Disclosed herein are methods of detecting a target nucleic acid sequence, determining the localization of the target nucleic acid sequence, and/or quantifying the number of target nucleic acid sequences in a cell. This method may be used on small target nucleic acid sequences, and may be referred to as Nano-FISH.

Described are methods for the production and use of cryopreservable double negative T cells (DNTs) for the treatment of cancer as an off-the-shelf cellular therapy. A sample population of DNTs is expanded using DNTs from one or more donors. The expanded population of DNTs from different donors does not exhibit alloreactivity against allogenic cells in the expanded population. The expanded populations of DNTs can be long-term stored as cryopreserved products.

Described are methods for the production and use of cryopreservable double negative T cells (DNTs) for the treatment of cancer as an off-the-shelf cellular therapy. A sample population of DNTs is expanded using DNTs from one or more donors. The expanded population of DNTs from different donors does not exhibit alloreactivity against allogenic cells in the expanded population. The expanded populations of DNTs can be long-term stored as cryopreserved products.