Method and kit for characterizing dopaminergic progenitor cells obtained by differentiating pluripotent stem cells
AssigneeMiltenyi Biotec GmbH
InventorsMelanie Jungblut, Andreas Bosio, Sebastian Knobel, Agnete Kirkeby, Malin Parmar
Life of this patent
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Abstract
This disclosure provides a method and kit for analyzing compositions of human floorplate mesencephalic dopaminergic (mesDA) progenitor cells that are obtained, for example, by differentiating embryonic stem cells. The mesDA progenitors are characterized using the positive markers FOXA2. OTX2, and optionally NKX2. 1. in combination with the negative markers OCT3/4, and optionally PAX6, and SOXI. Cells with an appropriate protein expression profile have therapeutic potential for restoring the function of dopaminergic neurons in conditions such as Parkinson's Disease.
Claims18
A method for obtaining and characterizing a composition of human dopaminergic progenitor cells, the method comprising:
- a) differentiating a population of human pluripotent stem cells or progeny thereof into a preparation of neural progenitor cells;
- b) maturing the preparation of neural progenitor cells into a cell composition that contains dopaminergic progenitor cells;
- c) determining what percentage of cells in the composition expresses each of a panel of markers consisting of FOXA2, OTX2, and PAX6, thereby obtaining a protein expression profile;
- whereby the cell composition is characterized as having a desirable phenotype if the cell expression profile is as follows:
- 80 to 100% of said cells are positive for FOXA2,
- 80 to 100% of said cells are positive for OTX2, and
- less than 10% of said cells are positive for PAX6;
- wherein the percentage of cells in the composition that express each of said markers is determined by flow cytometry or immunocytochemistry of single cells in the cell composition using antigen binding molecules that are specific for each of the markers.
A kit for analyzing the protein expression profile of a preparation of dopaminergic progenitor cells to determine if cells in the preparation have a desirable phenotype,
- wherein the kit comprises a panel of antigen binding molecules that consists of antibody binding molecules specific for each of FOXA2, OTX2, and PAX6, optionally in combination with other reagents,
- wherein the kit is configured and the antigen binding molecules are formulated whereby a user of the kit may determine on a single cell basis the percentage of cells in said preparation of neural progenitor cells that express each of said markers, thereby obtaining said protein expression profile.
A kit for analyzing the protein expression profile of a preparation of dopaminergic progenitor cells to determine if cells in the preparation have a desirable phenotype,
- wherein the kit comprises a panel of antigen binding molecules that consists of antibody binding molecules specific for the three positives markers FOXA2, OTX2, and NKX2. 1 and for the three negative markers OCT3/4, PAX6, and SOX1, optionally in combination with other reagents,
- wherein the kit is configured and the antigen binding molecules are formulated whereby a user of the kit may determine on a single cell basis the percentage of cells in said preparation of neural progenitor cells that express each of said markers, thereby obtaining said protein expression profile.
Dependencies are inferred from the claim text.