CPC C12N 5/0657 (2013.01) [A61K 35/34 (2013.01); G01N 1/286 (2013.01); G01N 33/56966 (2013.01); C12N 2500/30 (2013.01); C12N 2501/115 (2013.01); C12N 2501/14 (2013.01); G01N 2001/2873 (2013.01)] | 17 Claims |
1. A method for the isolation of human cardiac progenitor cells subpopulations, comprising the following steps:
a) mincing a heart tissue sample containing a population of cardiac cells;
b) 4-times progressive digestion of the cardiac tissue with an enzymatic mixture comprising collagenase and excluding trypsin in a concentration of between 0.1 and 3 mg/ml until a cell suspension is achieved and filtration is achieved through filters of size comprised between 30 μm and 100 μm;
c) further digestion of residual cardiac tissue obtained in step b) for 16 hours with an enzymatic mixture comprising collagenase and excluding trypsin in a concentration of between 0.1 and 3 mg/ml until a cell suspension is achieved and filtration through filters of size comprised between 30 μm and 100 μm;
d) culturing the cell suspension obtained in step b) and c) in a culture medium suitable to propagate the cardiac cells;
e) preliminary expansion of the cell suspension obtained in step d) in a culture medium suitable to expand the cardiac cells in the presence of an enzymatic or non-enzymatic solution;
f) secondary expansion of the cardiac cells obtained in step e) in a culture medium suitable to further expand the cardiac cells in the presence of an enzymatic or non enzymatic non-enzymatic solution;
g) isolating one or more subpopulations of cardiac cells by positive and/or negative selection by the use of monoclonal antibodies directed against one or more surface antigens expressed in the starting population of cardiac progenitor cells.
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