System and method for extraction and cryopreservation of bone marrow

1. A method for high-yield recovery of stem cells from cadaver bone or cadaver bone fragments, the method comprising:

(a) obtaining cadaver bone or cadaver bone fragments;

(b) processing the bone or bone fragments to extract bone marrow cells;

(c) obtaining a reduced density solution mixture of a hydrophilic copolymer of sucrose and epichlorohydrin at density 1.077 g/mL, mixed with an electrolyte medium, wherein the reduced density solution mixture provides a density gradient of 1.063-1.052 g/m;

(d) combining the extracted bone marrow cells with the reduced density solution mixture;

(e) centrifuging the extracted bone marrow cells, thereby separating out buffy coat cells comprising stem cells; and

(f) harvesting the buffy coat cells, thereby recovering stem cells comprising hematopoietic stem cells and/or mesenchymal stem cells.

2. The method of claim 1 , wherein after centrifuging in step (e), the buffy coat cells are located in the density gradient at a density of about 1.063 g/ml.

3. The method of claim 1 wherein the centrifuging occurs for about 30 minutes at 400 g.

4. The method of claim 1 , wherein the centrifuging occurs for about 5 minutes at 400 g.

5. The method of claim 1 , wherein the cadaver bone or cadaver bone fragment has not been cryopreserved prior to processing in step (b).

6. The method of claim 1 , wherein the purity of recovered stem cells is about 90% or a higher purity.

7. The method of claim 1 , wherein the viability of recovered stem cells is about 90% or a higher viability.

8. The method of claim 1 , wherein the processing in step (b) comprises grinding the cadaver bone fragments to obtain ground cadaver bone, wherein grinding uses a bone grinder in the presence of a grinding medium, and, optionally, further comprises dividing cadaver bone into cadaver bone fragments.

9. The method of claim 8 , wherein the grinding medium comprises two or more of enzymes that cleave DNA and RNA, human serum albumin (HSA) heparin, and an electrolyte medium.

10. The method of claim 8 , wherein the grinding medium promotes extraction of bone marrow cells from the ground cadaver bone.

11. The method of claim 10 , wherein the ground cadaver bone is captured on a filter or sieve and a filtrate that passes thru the filter or sieve comprises the extracted bone marrow cells.

12. The method of claim 1 , further comprising after step (f), a step of washing the harvested buffy coat cells for subsequent use or further processing.

13. The method of claim 12 , wherein the washing includes contacting the cells with phosphate-buffered saline (PBS) containing human serum albumin (HSA) and ethylenediaminetetraacetic acid (EDTA) or with PBS alone.

14. The method of claim 1 , further comprising a step of performing a separation method that isolates CD34+ cells from the recovered stem cells.

15. The method of claim 1 , wherein the cadaver bone or the cadaver bone fragments are derived from a vertebral body, an ileum, or a combination thereof.

16. The method of claim 1 , wherein the cadaver bone or cadaver bone fragments are derived from a plurality of cadaver bones from the same donor.

17. The method of claim 1 , wherein the hematopoietic stem cells are CD34+ cells.

18. The method of claim 1 , wherein the density gradient provides a recovered stem cell population having higher purity CD34+ cells with higher viability than a recovered stem cell population obtained from a conventional hydrophilic copolymer of sucrose and epichlorohydrin gradient.

19. The method of claim 1 , wherein the cadaver bone or cadaver bone fragments have been exposed to a hypothermic temperature.