BLOOD FACTOR MONITORING ASSAY AND USES THEREOF
The present disclosure provides methods and compositions for diagnosing and treating subject having a bleeding disorder. The disclosed methods comprise contacting a sample, e.g., a blood or plasma sample obtained from the patient, with an activation mixture comprising an activated coagulation factor and a phospholipid mixture, wherein the activation mixture is dried onto a solid substrate. Also provided is a global hemostasis test based on the integration of clotting time (Ct) and pharmacokinetics data. The methods and compositions presented can be applied to point-of-care diagnostic systems.
1 . A composition for the measurement of coagulation factor activity in a sample comprising an activated coagulation factor and a phospholipid mixture, wherein the composition is dried onto a solid substrate.
2 - 4 . (canceled)
5 . The composition of any one of claim 1 , wherein the coagulation factor is FVII, FVIII, FIX, or a fragment, variant, or derivative thereof.
6 - 13 . (canceled)
14 . The composition of claim 1 , wherein the phospholipid mixture comprises phosphatidylcholine, phosphatidylserine, phosphatidylglycerol, or combinations thereof.
15 . (canceled)
16 . The composition of claim 1 , wherein the phospholipid mixture comprises 70 mole-% of phosphatidylcholine and 30 mole-% of phosphatidylserine.
17 . The composition of claim 1 , wherein the phospholipid mixture comprises 80 mole-% of phosphatidylcholine, 10 mole-% of phosphatidylserine, and 10 mole-% of phosphatidylglycerol.
18 . The composition of claim 1 , wherein the phospholipid mixture comprises 75 mole-% of phosphatidylcholine, 20 mole-% of phosphatidylserine, and 5 mole-% of phosphatidylglycerol.
19 . The composition of claim 1 , wherein the phospholipid mixture further comprises cholesterol.
20 . The composition of claim 19 , wherein the cholesterol content in the phospholipid mixture is from about 1 to about 20 mole-% of cholesterol.
21 - 28 . (canceled)
29 . The composition of claim 1 , wherein the composition comprises 80% of 0.1 mg/mL Factor IXa and 20% of a phospholipid mixture comprising 75 mole-% of phosphatidylcholine, 20 mole-% of phosphatidylserine, and 5 mole-% of phosphatidylglycerol.
30 . The composition for the of claim 1 , wherein the composition comprises 80% of a Factor XIa suspension at about 0.1-0.5 mg/mL and 20% of a phospholipid mixture comprising 75 mole-% of phosphatidylcholine, 20 mole-% of phosphatidylserine, and 5 mole-% of phosphatidylglycerol, wherein said composition is dried onto a solid substrate.
31 . A kit for performing a measurement of coagulation factor activity or coagulation time in a sample comprising the composition of claim 1 .
32 - 43 . (canceled)
44 . A method for determining a coagulation factor level in a bleeding disorder patient, comprising:
(a) contacting a sample obtained from the patient with an activation mixture comprising an activated coagulation factor and a phospholipid mixture, wherein the activation mixture is dried onto a solid substrate;
(b) measuring the time between the contacting of the activation mixture with the sample and the onset of clotting, thereby calculating the clotting time (Ct); and,
(c) correlating the Ct value with the level of coagulation factor in the sample.
45 . The method of claim 44 , wherein the correlation between Ct and coagulation factor level (% Factor) is calculated according to the formula:
Ct= A ×Ln(% Factor)+ B [Formula I]
wherein, for each coagulation factor, A is a constant value corresponding to the slope of a Ct versus coagulation factor concentration dose-response, and B is patient-specific off-set value.
46 - 83 . (canceled)
84 . The method of claim 45 , wherein the phospholipid mixture comprises 70 mole-% of phosphatidylcholine and 30 mole-% of phosphatidylserine.
85 . The method of claim 45 , wherein the phospholipid mixture comprises 80 mole-% of phosphatidylcholine, 10 mole-% of phosphatidylserine, and 10 mole-% of phosphatidylglycerol.
86 . The method of claim 45 , wherein the phospholipid mixture comprises 75 mole-% of phosphatidylcholine, 20 mole-% of phosphatidylserine, and 5 mole-% of phosphatidylglycerol.
87 . The method of claim 45 , wherein the phospholipid mixture further comprises cholesterol.
88 . The method of claim 87 , wherein the cholesterol content in the phospholipid mixture is from about 1 to about 20 mole-% of cholesterol.
89 - 108 . (canceled)