Method of profiling a cell population
View Patent ↗The present invention relates to a method of profiling a cell population comprising a step of detecting the presence or absence of at least two biological markers in said cell population, wherein at least one of said markers is a cell surface marker, which is a sialylated N-glycan marker with structure NeuNAcα3Gal, and at least one of said markers is a mRNA marker related to glycoproteins and/or glycosynthase proteins. The invention also relates to method for purification of cord blood cell population and to a complete cell population from cord blood purified by said method.
1. A method for purification of a cell population from a sample containing multiple cell populations, comprising the following sequential steps:
a) contacting a sample with a cell-type-specific binding reagent linked to a polyvalent matrix by mixing said reagent with the sample so that a complex of said binding reagent and a cell population of interest forms;
b) immobilizing the complex obtained from step a);
c) removing the material that is not immobilized;
d) releasing the immobilized complex;
e) contacting the sample obtained from step d) with an additional amount of the specific binding reagent linked to a polyvalent matrix by mixing the additional amount of binding reagent with said sample;
f) immobilizing the complex obtained from step e);
g) removing the material that is not immobilized;
h) releasing the immobilized complex; and
i) recovering a cell population bound to the binding reagent.
2. The method according to claim 1 , wherein said polyvalent matrix is selected from the group consisting of: a solid phase, a solid phase which comprises magnetic material, and a solid phase which comprises a magnetic bead, and said magnetic bead is applied to a magnetic field.
3. The method according to claim 1 , wherein the cell population of step i) is recovered as a complex of the binding reagent and the cell population, and said cell population is released from the binding reagent.
4. The method according to claim 1 , wherein the binding reagent is a monoclonal antibody.
5. The method according to claim 1 , wherein the cells in said sample are labeled with a marker and the binding reagent is a labeled antibody applicable for an immunoassay or FACS analysis.
6. The method according to claim 1 , wherein the washing in steps c) and g) involve up to 1-5 washing steps.
7. The method according to claim 1 , wherein the cell population is a stem cell population.
8. The method according to claim 1 , wherein said cord blood cell population contains CD34 or CD133 antigen.
9. The method according to claim 8 further comprising the step of profiling the obtained cell population by detecting the presence or absence of at least one mRNA marker listed in Table 1, 2, 4-15, 17, 18, 20, 22, or 26 in said cell population.
10. The method according to claim 8 further comprising the step of detecting expression of sialyltransferase ST3Ga1VI and amount of α3-linked sialic acids on N-glycans in said cell population.
11. A purified cord blood cell population containing CD34 antigen obtained by the method according to claim 8 , wherein said cells are CD34+ cells with a purity of at least 90% as assessed by flow cytometry and wherein said sample of cells comprises above 5% CD34+ dim cells as observable by flow cytometry.
12. The purified cord blood cell population according to claim 11 , wherein the cells show expression of at least one mRNA marker listed in Table 17 or 19.
13. The purified cord blood cell population according to claim 11 , wherein total CFU count for the CD34+ cells is about 80/1000 cells.
14. The method of claim 1 , wherein the cell population is a cord blood cell population.