Methods and products obtained from the method for isolating and culturing mixed populations of stem cells, making decellularized tissue matrix, making decellularized tissue matrix infused with said mixed populations of stem cells, and methods of stem cell therapy are provided.

The in vitro organoid model is a major technological breakthrough and an essential tool to study the basic biology of an organ system and for the development of various clinical applications for disease intervention. Organoids can self-renew and exhibit similarities in function as of their tissue of origin. Here, a step-by-step protocol is described to isolate region-specific progenitors from the human lung and generate 3D organoid cultures as an experimental and validation tool.

Provided is an isolated population of human pluripotent stem cells comprising at least 50% human pluripotent stem cells characterized by an OCT4+/TRA1-60−/TRA1-81−/SSEA1+/SSEA4− expression signature, and novel methods of generating and maintaining same in a pluripotent, undifferentiated state a suspension culture devoid of cell clumps. Also provided are novel culture media, cell cultures and methods for culturing pluripotent stem cells in a suspension culture or a two-dimensional culture system while maintaining the cells in a proliferative, pluripotent and undifferentiated state. The novel culture media comprise interleukin 11 (IL11) and Ciliary Neurotrophic Factor (CNTF); bFGF at a concentration of at least 50 ng/ml and an IL6RIL6 chimera; or an animal contaminant-free serum replacement and an IL6RIL6 chimera. Also provided are methods for generating lineage-specific cells from the pluripotent stem cells.

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.

A method of producing the constituents of a therapeutic product from mammalian cells is described herein. Cells are isolated from a mammalian source. The cells are exposed to supercritical carbon dioxide (SCCO.sub.2) for 1 to 30 minutes, where the SCCO.sub.2 is maintained at a pressure of 1071 pounds per square inch (PSI) and a temperature of 31.1 to 45 degrees Celsius during the exposure. The exposure dissociates the cellular membranes of the cells to release intramembrane components therein to produce constituents of the therapeutic product. The mammalian cells may include at least one of platelets, stem cells, germ cells, and somatic cells. The methods described herein are particularly advantageous for releasing and capturing therapeutic intramembrane components from platelets and alpha-granules.

The present invention provides an in vitro method for obtaining and purifying regulatory T cells from thymic tissue (or thyTreg cells), which makes it possible to obtain more than 10 billion cells from a single thymus. These thyTregs obtained in the invention have a purity of more than 95% and very high suppressive capacity, survival and viability, in addition to being safe from a clinical viewpoint. The foregoing would not require the use of massive ex vivo cell expansion protocols. The transfer of these thyTreg cells to patients enables immune tolerance induction. Thus, said cells may be used as cell therapy to induce immune tolerance in the treatment and/or prevention of transplant rejections and in autoimmune diseases.

A method of permeabilizing a cell is disclosed. Kits for permeabilizing cells and analysing RNA contents are also disclosed.

An apparatus for collecting or culturing cells or cell colonies includes: a common substrate formed from a flexible resilient polymeric material and having a plurality of wells formed therein; and a plurality of rigid cell carriers releasably connected to said common substrate, with said carriers arranged in the form of an array, and with each of the carriers resiliently received in one of the wells. A method of collecting or culturing cells or cell colonies with such an apparatus is carried out by depositing a liquid media carrying cells on the apparatus so that said cells settle on or adhere to said the carriers; and then (c) releasing at least one selected carrier having said cells thereon by gradual application of release energy to each carrier from the cavity in which it is received (e.g., by pushing with a probe).

An apparatus for collecting or culturing cells or cell colonies includes: a common substrate formed from a flexible resilient polymeric material and having a plurality of wells formed therein; and a plurality of rigid cell carriers releasably connected to said common substrate, with said carriers arranged in the form of an array, and with each of the carriers resiliently received in one of the wells. A method of collecting or culturing cells or cell colonies with such an apparatus is carried out by depositing a liquid media carrying cells on the apparatus so that said cells settle on or adhere to said the carriers; and then (c) releasing at least one selected carrier having said cells thereon by gradual application of release energy to each carrier from the cavity in which it is received (e.g., by pushing with a probe).

A device for mechanical fragmentation of tissues facilitates preparation of the composition of isolated cells from a tissue sample, the preparation including mechanical fragmentation of the tissue, the device including a motorized support and a container, the motorized support including a motor having a rotor secured to a coupling member, the container internally including a rotary shaft with radial arms sweeping the inside of the container as the shaft rotates, the motorized support and the container including a complementary securing unit that allow the container to be immobilized on the motorized support, the coupling member and the rotary shaft being configured in such a way as to allow axial rotation of the rotary shaft when the rotor of the motor is turning. The radial arms are of two types: rigid vane-type arms, and flexible filamentary-type arms, notably made of nylon or of metal. A corresponding method is also disclosed.