| CPC A61L 27/3608 (2013.01) [A61K 35/28 (2013.01); A61K 35/32 (2013.01); A61L 27/3604 (2013.01); A61L 27/365 (2013.01); A61L 27/3687 (2013.01); A61L 27/3691 (2013.01); A61L 2430/02 (2013.01)] | 12 Claims |
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1. A method of making a biological composition, comprising:
(a) grinding a cut vertebral body from a human cadaver donor to produce a mixture comprising crushed vertebral bone and bone marrow;
(b) mechanically separating cellular and non-cellular components of the bone marrow from the vertebral bone in the mixture by tumbling and sieving the vertebral bone from a decanted fluid of the bone marrow comprising cellular components comprising whole cells, and non-whole cellular components comprising non-whole cell components;
(c) concentrating the decanted fluid by centrifugation;
(d) filtering the decanted fluid through a blood filter to form a mixture including the cellular and non-whole cellular components and remove any remaining vertebral bone;
(e) separating the cellular components and non-whole cellular components of the bone marrow directly from the mixture of step (d) by density gradient centrifugation, wherein the separation by centrifugation establishes a cell density gradient;
(f) collecting non-cellular fractions, non-cellular components, or combinations thereof of predetermined density from the gradient;
(g) washing the non-cellular fractions, non-cellular components, or combinations thereof to create a mixture;
(h) suspending the mixture of step (g) to a predetermined concentration in a polyampholyte cryoprotectant to form a coated biological composition;
(i) freezing the coated biological composition at a predetermined controlled rate; and
(j) packaging a bone blend having particles of 100 to 300 μm of demineralized cortical bone, mineralized cortical bone, and mineralized cancellous bone with the coated biological composition.
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