This document relates to bioengineering and involves bioengineered thymus organoids and related humanized animal models. The thymus organoids and animal models have various commercial and clinical uses, including generating humanized antibodies, making antigen-specific human T cells, inducing transplantation tolerance, rejuvenating thymus functions, and modeling human diseases.

Compositions disclosed herein, and methods of use thereof included those for treating or preventing SARS-CoV-2 vims infection in a subject in need, including methods of extending of the survival of a subject suffering from COVID-19, and reduction of organ dysfunction or failure due to COVID-19 or associated symptoms. Methods of treating or preventing COVID-19 in a subject in need includes administering compositions comprising early apoptotic cells or early apoptotic cell supernatants. Compositions and methods of use thereof may reduce the negative proinflammatory effect accompanying COVID-19 and symptoms thereof. Further, anti-inflammatory cytokine release may be increased. In certain instances, compositions may include additional agents.

Herein is reported a method for co-cultivating one or more B-cells comprising the step of incubating the one or more B-cells with EL4-B5 cells, whereby the EL4-B5 cells have been obtained/are from a cultivation of EL4-B5 cells that has a cell density of from 600,000 cells/ml up to 1,500,000 cells/ml.

Embodiments of the disclosure include compositions and methods related to engineered invariant natural killer T (iNKT) cells for off-the-shelf use for clinical therapy. In particular embodiments, the iNKT cells are produced from hematopoietic stem progenitor cells and also are suitable for allogeneic cellular therapy because they are HLA negative. In specific embodiments, the cells are cultured in a particular in vitro three-dimensional artificial thymic organoid system and the cells have imaging and suicide targeting capabilities.

Herein is reported a method for obtaining a B-cell comprising the following steps a) labeling B-cells, b) depositing the labeled B-cells as single cells, c) co-cultivating the single cell deposited B-cells with feeder cells, d) selecting a B-cell proliferating and secreting IgG in step c) and thereby obtaining a B-cell. The labeling can be of IgG.sup.+CD19.sup.+-B-cells, IgG.sup.+CD38.sup.+-B-cells, IgG.sup.+CD268.sup.+-B-cells, IgG.sup.CD138.sup.+-B-cells, CD27.sup.+CD138.sup.+-B-cells or CD3.sup.CD27.sup.+-B-cells. The method can comprise the step of incubating said B-cells at 37 C. for one hour in EL-4 B5 medium prior to the depositing step. The method can also comprise the step of centrifuging said single cell deposited B-cells prior to the co-cultivation. In the co-cultivation a feeder mix comprising interleukin-1beta, and tumor necrosis factor alpha and Staphylococcus aureus strain Cowans cells or BAFF or interleukin-2 and/or interleukin-10 and/or interleukin-6 and/or interleukin-4 can be used.

Herein is reported a method for co-cultivating one or more B-cells comprising the step of incubating the one or more B-cells with EL4-B5 cells, whereby the EL4-B5 cells have been obtained/are from a cultivation of EL4-B5 cells that has a cell density of from 600,000 cells/ml up to 1,500,000 cells/ml.

Herein is reported a method for co-cultivating one or more B-cells comprising the step of incubating the one or more B-cells with EL4-B5 cells, whereby the EL4-B5 cells have been obtained/are from a cultivation of EL4-B5 cells that has a cell density of from 600,000 cells/ml up to 1,500,000 cells/ml.

Herein is reported a method for the co-cultivation of single deposited B-cells, which can be of any source, with EL-4 B5 feeder cells in a suitable co-cultivation medium. In the herein reported methods the EL-4 B5 cells have been irradiated with a dose of less than 40 Gy, preferably 9.5 Gy or less. Thereby the EL-4 B5 cells have a higher viability and maintain the ability to divide in cultivation at doses less than 6 Gy.

The present disclosure provides engineered thymic epithelial cells and cell lines, as well as extracellular vesicles derived therefrom. The disclosure also provides exosomes that display specific surface proteins, and provides methods for using said materials for treating subjects and for identifying cells.

Herein is reported a method for producing a thymocyte supernatant comprising the steps of co-cultivating thymocytes and mononuclear cells at a cell ratio of at least 0.5:1.2 in the presence of phorbol-12-myristate-13-acetate and Phytohemagglutinin M for up to 60 hours, and separating the co-cultivation medium from the cells and thereby producing the thymocyte supernatant.