Compositions comprising adjustable concentrations of growth factors derived from blood serum and clot hypoxia-conditioned medium and methods of their production

A method for preparing a kit-of parts, said method comprising the steps:

• (i) providing a blood sample from a subject;
• (ii) allowing the blood sample of step (i) to clot or inducing clotting in the blood sample of step (i) thereby obtaining a composition comprising a blood clot and serum;
• (iii) removing at least a part of the serum from the composition obtained in step (ii) and determining the volume of the serum removed;
• (iv) adding medium to the blood clot, wherein the volume of the added medium is equal to or less than the volume of the serum removed in step (iii);
• (v) incubating the clot in said medium at a temperature between 10° C. and 40° C. under hypoxic conditions for 1 to 7 days, thereby obtaining a hypoxia-conditioned medium;
• (vi) collecting at least a part of the hypoxia-conditioned medium obtained in step (v);
• and
• (vii) preparing a kit-of-parts comprising:
• (a) the hypoxia-conditioned medium collected in step (vi) and
• (b) at least a part of the serum removed in step (iii).

A method for producing a solution of growth factors, said method comprising the steps:

• (i) providing a blood sample from a subject;
• (ii) allowing the blood sample of step (i) to clot or inducing clotting in the blood sample of step (i), thereby obtaining a composition comprising a blood clot and serum;
• (iii) removing at least a part of the serum from the composition obtained in step (ii) and determining the volume of the serum removed;
• (iv) adding medium to the blood clot, wherein the volume of the added medium is equal to or less than the volume of the serum removed in step (iii);
• (v) incubating the clot in said medium at a temperature between 10° C. and 40° C. under hypoxic conditions for 1 to 7 days, thereby obtaining a hypoxia-conditioned medium;
• (vi) collecting at least a part of the hypoxia-conditioned medium obtained in step (v);
• (vii) mixing the hypoxia-conditioned medium collected in step (vi) with at least a part of the serum removed in step (iii), thereby obtaining the solution of growth factors.

A method for preparing a kit-of parts, said method comprising the steps:

• (i) providing a first blood sample from a subject;
• (ii) allowing the blood sample of step (i) to clot or inducing clotting in the blood sample of step (i), thereby obtaining a first composition comprising a blood clot and serum;
• (iii) removing at least a part of the serum from the first composition obtained in step (ii) and determining the volume of the serum removed;
• (iv) adding medium to the blood clot, wherein the volume of the added medium is equal to or less than the volume of the serum removed in step (iii);
• (v) incubating the clot in said medium at a temperature between 10° C. and 40° C. under hypoxic conditions for 1 to 7 days, thereby obtaining a hypoxia-conditioned medium;
• (vi) collecting at least a part of the hypoxia-conditioned medium obtained in step (v);
• (vii) providing a second blood sample from a subject;
• (viii) allowing the blood sample of step (vii) to clot or inducing clotting in the blood sample of step (vii), thereby obtaining a second composition comprising a blood clot and serum;
• (ix) recovering at least a part of the serum from the second composition obtained in step (viii);
• and
• (x) preparing a kit-of-parts comprising:
• (a) the hypoxia-conditioned medium collected in step (vi) and
• (b) the serum recovered in step (ix).

A method for producing a solution of growth factors, said method comprising the steps:

• (i) providing a first blood sample from a subject;
• (ii) allowing the blood sample of step (i) to clot or inducing clotting in the blood sample of step (i), thereby obtaining a first composition comprising a blood clot and serum;
• (iii) removing at least a part of the serum from the first composition obtained in step (ii) and determining the volume of the serum removed;
• (iv) adding medium to the blood clot, wherein the volume of the added medium is equal to or less than the volume of the serum removed in step (iii);
• (v) incubating the clot in said medium at a temperature between 10° C. and 40° C. under hypoxic conditions for 1 to 7 days, thereby obtaining a hypoxia-conditioned medium;
• (vi) collecting at least a part of the hypoxia-conditioned medium obtained in step (v);
• (vii) providing a second blood sample from a subject;
• (viii) allowing the blood sample of step (vii) to clot or inducing clotting in the blood sample of step (vii), thereby obtaining a second composition comprising a blood clot and serum;
• (ix) recovering at least a part of the serum from the second composition obtained in step (viii), thereby obtaining a serum;
• and
• (x) mixing the hypoxia-conditioned medium collected in step (vi) with at least a part of the serum recovered in step (ix), thereby obtaining the solution of growth factors.

A kit for preparing serum and/or hypoxia-conditioned medium derived from blood, said kit comprising:

• (a) a first syringe,
• (b) a second syringe,
• (c) optionally a third syringe,
• (d) optionally a fourth syringe,
• (e) a three-way valve,
• (f) a filter,
• (g) a cap,
• (h) optionally a container containing sterile medium,
• (i) optionally a catheter or butterfly cannula,
• (j) optionally a sticking plaster,
• (k) optionally a rack into which the first syringe can be placed and maintained in upright position, and
• (l) optionally a spatula.