Claims for Patent: 6,472,162

Summary for Patent: 6,472,162

Title:	Method for preparing thrombin for use in a biological glue
Abstract:	A sterile method for preparing stable thrombin component from a single donor\'s plasma in which the thrombin component and the clotting and adhesive proteins component are harvested simultaneously from the same donor plasma in less than one hour. The combined components provide an improved biological hemostatic agent and tissue sealant by virtue of its freedom from the risk of contaminating viruses or bacteria from allogenic human or bovine blood sources. The thrombin provides polymerization of the clotting and adhesive proteins in less than five seconds, and is sufficiently stable to provide that fast clotting over a six hour period. Further, the clotting times can be predictably lengthened by diluting the thrombin with saline.
Inventor(s):	Coelho; Philip Henry (El Dorado Hills, CA), Kingsley; Phil (Sacramento, CA), Brausch; Jim (Sacramento, CA), Godsey; James H. (Folsom, CA), Rock; Gail (Ottawa, CA), Madsen; Trista K. (Elk Grove, CA), Frausto; Sona B. (Sacramento, CA)
Assignee:	ThermoGenesis Corp. (Rancho Cordovo, CA)
Application Number:	09/328,350
Patent Claims:	1. A method for extracting autologous thrombin from a patient, the steps consisting of: obtaining a blood product from the patient; sequestering plasma from the blood product; adding ethanol to the plasma to prepare a solution containing prothrombin, wherein ethanol is present in the solution at a concentration between about 8% and about 20% volume per unit volume; converting the prothrombin in the solution to thrombin; filtering the thrombin to remove particulate matter; and applying the thrombin to the patient. 2. The method of claim 1 further including the step of mixing the thrombin with clotting proteins to form a fibrin clot, but first altering time required for the thrombin to convert fibrinogen to a fibrin clot. 3. The method of claim 2 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes diluting the thrombin with saline. 4. The method of claim 3 including filtering the plasma to separate included particles by weight, size and protein binding. 5. The method of claim 2 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding a source of calcium ions to the solution. 6. The method of claim 2 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding CaCl.sub.2 to the solution. 7. The method of claim 2 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding saline to the solution. 8. The method of claim 2 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding sterile water to the solution. 9. The method of claim 2 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes changing concentration of the ethanol. 10. The method of claim 1 wherein the converting step includes adding CaCl.sub.2 to the solution. 11. The method of claim 10 including centrifuging the blood product for obtaining plasma. 12. The method of claim 1 wherein ethanol is present in the solution at a concentration of about 18.9% volume per unit volume. 13. The method of claim 1 wherein the time required to extract the thrombin is between about 30 minutes and about 75 minutes. 14. The method of claim 10 wherein CaCl.sub.2 is present in the solution at a concentration between about 4.5 mM and about 23.0 mM by volume. 15. The method of claim 14 wherein CaCl.sub.2 is present in the solution at a concentration of about 5.7 mM by volume. 16. The method of claim 10 wherein the converting step further includes agitating the solution after adding CaCl.sub.2 to the solution. 17. A method for extracting and then dispensing thrombin, the steps consisting of: taking whole blood from a person, sequestering prothrombin from the whole blood by addition of ethanol, wherein ethanol is present at a concentration between about 8% and about 20% volume per unit volume, converting the prothrombin to thrombin, loading the thrombin into a syringe, and using the syringe to dispense the thrombin to stem blood flow. 18. The method of claim 17 including loading clotting proteins into another syringe and dispensing the clotting proteins concurrently with the thrombin. 19. A method for extracting thrombin from one person, the steps consisting of: adding ethanol to sequester prothrombin from plasma taken from one person, wherein ethanol is present at a concentration between about 8% and about 20% volume per unit volume, converting the prothrombin to thrombin, and removing particulate material from the thrombin. 20. The method of claim 19 further including diluting the thrombin to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 21. The method of claim 20 including adding a source of calcium ions to the thrombin to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 22. The method of claim 21 including adding CaCl.sub.2 to the thrombin to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 23. The method of claim 20 including adding saline to the thrombin to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 24. The method of claim 20 including adding sterile water to the thrombin to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 25. The method of claim 19 wherein ethanol is present at a concentration of about 18.9% volume per unit volume. 26. The method of claim 19 wherein the time required to extract the thrombin is between about 30 minutes and about 75 minutes. 27. The method of claim 19 wherein the time required to extract the thrombin is less than about one hour and greater than zero minutes. 28. The method of claim 19 wherein the converting step includes adding CaCl.sub.2. 29. A method for extracting thrombin from one person, the steps consisting of: taking whole blood from one person, obtaining plasma from the whole blood, adding ethanol to the plasma to prepare a solution containing prothrombin, wherein ethanol is present in the solution at a concentration between about 8% and about 20% volume per unit volume, converting the prothrombin to thrombin, and sequestering the thrombin. 30. The method of claim 29 further including the step of mixing the thrombin with clotting proteins to form a fibrin clot, but first altering time required for the thrombin to convert fibrinogen to a fibrin clot to a time of between about two seconds and about five seconds. 31. The method of claim 30 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding a source of calcium ions to the solution. 32. The method of claim 31 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding CaCl.sub.2 to the solution. 33. The method of claim 30 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding saline to the solution. 34. The method of claim 30 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding sterile water to the solution. 35. The method of claim 29 including making the thrombin stable for a period of time between about 15 minutes and about 360 minutes. 36. The method of claim 35 including adding a source of calcium ions to the solution to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 37. The method of claim 36 including adding CaCl.sub.2 to the solution to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 38. The method of claim 35 including adding saline to the solution to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 39. The method of claim 35 including adding sterile water to the solution to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 40. The method of claim 29 wherein ethanol is present in the solution at a concentration of about 18.9% volume per unit volume. 41. The method of claim 29 wherein the time required to extract the thrombin is between about 30 minutes and about 75 minutes. 42. The method of claim 29 wherein the time required to extract the thrombin is less than about one hour and greater than zero minutes. 43. The method of claim 29 wherein the converting step includes adding CaCl.sub.2 to the solution. 44. A method for extracting autologous thrombin from a patient, the steps consisting essentially of: obtaining a blood product from the patient; sequestering plasma from the blood product; adding ethanol to the plasma to prepare a solution containing prothrombin, wherein ethanol is present in the solution at a concentration between about 8% and about 20% volume per unit volume; converting the prothrombin in the solution to thrombin; filtering the thrombin to remove particulate matter; and applying the thrombin to the patient. 45. The method of claim 44 wherein ethanol is present in the solution at a concentration of about 18.9% volume per unit volume. 46. The method of claim 44 wherein the time required to extract the autologous thrombin is between about 30 minutes and about 75 minutes. 47. The method of claim 44 further including the step of mixing the thrombin with clotting proteins to form a fibrin clot, but first altering time required for the thrombin to convert fibrinogen to a fibrin clot. 48. The method of claim 47 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes changing the concentration of the ethanol. 49. The method of claim 47 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes diluting the thrombin with saline. 50. The method of claim 44 wherein the converting step includes adding CaCl.sub.2 to the solution. 51. The method of claim 50 wherein CaCl.sub.2 is present in the solution at a concentration between about 4.5 mM and about 23.0 mM by volume. 52. The method of claim 51 wherein CaCl.sub.2 is present in the solution at a concentration of about 5.7 mM by volume. 53. The method of claim 50 wherein the converting step further includes agitating the solution after the addition of the CaCl.sub.2 to the solution. 54. The method of claim 50 including centrifuging the blood product for obtaining plasma. 55. A method for extracting and then dispensing thrombin, the steps consisting essentially of: taking whole blood from a person, sequestering prothrombin from the whole blood by addition of ethanol, wherein ethanol is present at a concentration between about 8% and about 20% volume per unit volume, converting the prothrombin to thrombin, loading the thrombin into a syringe, and using the syringe to dispense the thrombin to stem blood flow. 56. The method of claim 55 including loading clotting proteins into another syringe and dispensing the clotting proteins concurrently with the thrombin. 57. A method for extracting thrombin from one person, the steps consisting essentially of: adding ethanol to sequester prothrombin from plasma taken from one person, wherein ethanol is present at a concentration between about 8% and about 20% volume per unit volume, converting the prothrombin to thrombin, and removing particulate material from the thrombin. 58. The method of claim 57 wherein ethanol is present at a concentration of about 18.9% volume per unit volume. 59. The method of claim 57 wherein the time required to extract the thrombin is between about 30 minutes and about 75 minutes. 60. The method of claim 57 wherein the time required to extract the thrombin is less than about one hour and greater than zero minutes. 61. The method of claim 57 wherein the converting step includes adding CaCl.sub.2. 62. The method of claim 57 further including diluting the thrombin to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 63. The method of claim 62 including adding a source of calcium ions to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 64. The method of claim 63 including adding CaCl.sub.2 to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 65. The method of claim 62 including adding saline to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 66. The method of claim 62 including adding sterile water to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 67. A method for extracting thrombin from one person, the steps consisting essentially of: taking whole blood from one person, obtaining plasma from the whole blood, adding ethanol to the plasma to prepare a solution containing prothrombin, wherein ethanol is present in the solution at a concentration between about 8% and about 20% per volume per unit volume, converting the prothrombin to thrombin, and sequestering the thrombin. 68. The method of claim 67 wherein ethanol is present in the solution at a concentration of about 18.9% volume per unit volume. 69. The method of claim 67 wherein the time required to extract the thrombin is between about 30 minutes and about 75 minutes. 70. The method of claim 67 wherein the time required to extract the thrombin is less than about one hour and greater than zero minutes. 71. The method of claim 67 wherein the converting step includes adding CaCl.sub.2 to the solution. 72. The method of claim 67 further including the step of mixing the thrombin with clotting proteins to form a fibrin clot, but first altering time required for the thrombin to convert fibrinogen to a fibrin clot to a time of between about two seconds and about five seconds. 73. The method of claim 72 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding a source of calcium ions to the solution. 74. The method of claim 73 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding CaCl.sub.2 to the solution. 75. The method of claim 72 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding saline to the solution. 76. The method of claim 72 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding sterile water to the solution. 77. The method of claim 67 including making the thrombin stable for a period of time between about 15 minutes and about 360 minutes. 78. The method of claim 77 including adding a source of calcium ions to the solution to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 79. The method of claim 78 including adding CaCl.sub.2 to the solution to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 80. The method of claim 77 including adding saline to the solution to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 81. The method of claim 77 including adding sterile water to the solution to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 82. A method for extracting thrombin, the steps consisting essentially of: taking whole blood from a person, obtaining plasma from the whole blood, adding ethanol to the plasma to prepare a solution containing prothrombin, wherein ethanol is present in the solution at a concentration between about 8% and about 20% per volume per unit volume, converting the prothrombin to thrombin, and sequestering the thrombin. 83. The method of claim 82 wherein ethanol is present in the solution at a concentration of about 18.9% volume per unit volume. 84. The method of claim 82 wherein the time required to extract the thrombin is between about 30 minutes and about 75 minutes. 85. The method of claim 82 wherein the time required to extract thrombin is less than about one hour and greater than zero minutes. 86. The method of claim 82 wherein the converting step includes adding CaCl.sub.2 to the solution. 87. The method of claim 82 further including the step of mixing the thrombin with clotting proteins to form a fibrin clot but first altering time required for the thrombin to convert fibrinogen to a fibrin clot to a time of between about two seconds and about five seconds. 88. The method of claim 87 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding a source of calcium ions to the solution. 89. The method of claim 88 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding CaCl.sub.2 to the solution. 90. The method of claim 87 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding saline to the solution. 91. The method of claim 87 wherein altering time required for the thrombin to convert fibrinogen to a fibrin clot includes adding sterile water to the solution. 92. The method of claim 82 including making the thrombin stable for a period of time between about 15 minutes and about 360 minutes. 93. The method of claim 92 including adding a source of calcium ions to the solution to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 94. The method of claim 93 including adding CaCl.sub.2 to the solution to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 95. The method of claim 92 including adding saline to the solution to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 96. The method of claim 92 including adding sterile water to the solution to alter time required for the thrombin to convert fibrinogen to a fibrin clot. 97. A method for extracting and then dispensing thrombin, the steps consisting of: taking whole blood from a person, sequestering prothrombin from the whole blood by addition of ethanol, converting the prothrombin to thrombin, making the thrombin stable for a period of time between about 15 minutes and about 360 minutes, loading the thrombin into a syringe, and the syringe to dispense the thrombin to stem blood flow. 98. A method for extracting thrombin from one person, the steps consisting of: taking whole blood from one person, obtaining plasma from the whole blood, adding ethanol to the plasma to prepare a solution containing prothrombin, converting the prothrombin to thrombin, making the thrombin stable for a period of time between about 15 minutes and about 360 minutes, and sequestering the thrombin. 99. A method for extracting and then dispensing thrombin, the steps consisting essentially of: taking whole blood from a person, sequestering prothrombin from the whole blood by addition of ethanol, converting the prothrombin to thrombin, making the thrombin stable for a period of time between about 15 minutes and about 360 minutes, loading the thrombin into a syringe, and using the syringe to dispense the thrombin to stem blood flow. 100. A method for generating thrombin from one person, the steps consisting essentially of: taking whole blood from one person, obtaining plasma from the whole blood, adding ethanol to the plasma to prepare a solution containing prothrombin, converting the prothrombin to thrombin, making the thrombin stable for a period of time between about 15 minutes and about 360 minutes, and sequestering the thrombin. 101. A method for extracting thrombin, the steps consisting essentially of: taking whole blood from a person, obtaining plasma from the whole blood, adding ethanol to the plasma to prepare a solution containing prothrombin, converting the prothrombin to thrombin, making the thrombin stable for a period of time between about 15 minutes and about 360 minutes, and sequestering the thrombin. 102. A method for extracting autologous thrombin from a patient, the steps consisting of: obtaining a blood product from the patient; sequestering plasma from the blood product; adding ethanol to the plasma to prepare a solution containing prothrombin; converting the prothrombin in the solution to thrombin; filtering the thrombin to remove particulate matter; and applying the thrombin to the patient, wherein the time required to extract the autologous thrombin is between about 30 minutes and about 75 minutes. 103. A method for extracting and then dispensing thrombin, the steps consisting of: taking whole blood from a person, sequestering prothrombin from the whole blood by addition of ethanol, converting the prothrombin to thrombin, loading the thrombin into a syringe, and using the syringe to dispense the thrombin to stem blood flow, wherein the time required to extract the thrombin is between about 30 minutes and about 75 minutes. 104. A method for extracting thrombin from one person, the steps consisting of: adding ethanol to sequester prothrombin from plasma taken from one person, converting the prothrombin to thrombin, and removing particulate material from the thrombin, wherein the time required to extract the thrombin is between about 30 minutes and about 75 minutes. 105. A method for extracting thrombin from one person, the steps consisting of: taking whole blood from one person, obtaining plasma from the whole blood, adding ethanol to the plasma to prepare a solution containing prothrombin, converting the prothrombin to thrombin; and sequestering the thrombin, wherein the time required to extract the thrombin is between about 30 minutes and about 75 minutes. 106. A method for extracting autologous thrombin from a patient, the steps consisting essentially of: obtaining a blood product from the patient; sequestering plasma from the blood product; adding ethanol to the plasma to prepare a solution containing prothrombin; converting the prothrombin in the solution to thrombin; filtering the thrombin to remove particulate matter; and applying the thrombin to the patient, wherein the time required to extract the autologous thrombin is between about 30 minutes and about 75 minutes. 107. A method for extracting and then dispensing thrombin, the steps consisting essentially of: taking whole blood from a person, sequestering prothrombin from the whole blood by addition of ethanol, converting the prothrombin to thrombin, loading the thrombin into a syringe, and using the syringe to dispense the thrombin to stem blood flow, wherein the time required to extract the thrombin is between about 30 minutes and about 5 minutes. 108. A method for extracting thrombin from one person, the steps consisting essentially of: adding ethanol to sequester prothrombin from plasma taken from one person, converting the prothrombin to thrombin, and removing particulate material from the thrombin, wherein the time required to extract the thrombin is between about 30 minutes and about 75 minutes. 109. A method for extracting thrombin from one person, the steps consisting essentially of: taking whole blood from one person, obtaining plasma from the whole blood, adding ethanol to the plasma to prepare a solution containing prothrombin, converting the prothrombin to thrombin, and sequestering the thrombin, wherein the time required to extract the thrombin is between about 30 minutes and about 75 minutes. 110. A method for extracting thrombin, the steps consisting essentially of: taking whole blood from a person, obtaining plasma from the whole blood, adding ethanol to the plasma to prepare a solution containing prothrombin, converting the prothrombin to thrombin, and sequestering the thrombin. 111. A method for extracting autologous thrombin from a patient, the steps consisting of: obtaining a blood product from the patient; sequestering plasma from the blood product; adding ethanol to the plasma to prepare a solution containing prothrombin, converting the prothrombin in the solution to thrombin; filtering the thrombin to remove particulate matter; applying the thrombin to the patient; and mixing the thrombin with clotting proteins to form a fibrin clot, but first altering time required for the thrombin to convert fibrinogen to a fibrin clot to a time of between about two seconds and about five seconds by adding an altering agent selected from the group including calcium ion sources, CaCl.sub.2, saline, and sterile water. 112. A method for extracting and then dispensing thrombin, the steps consisting of: taking whole blood from a person, sequestering prothrombin from the whole blood by addition of ethanol, converting the prothrombin to thrombin, loading the thrombin into a syringe, using the syringe to dispense the thrombin to stem blood flow, and mixing the thrombin with clotting proteins to form a fibrin clot, but first altering time required for the thrombin to convert fibrinogen to a fibrin clot to a time of between about two seconds and about five seconds by adding an altering agent selected from the group including calcium ion sources, CaCl.sub.2, saline, and sterile water. 113. A method for extracting thrombin from one person, the steps consisting of: adding ethanol to sequester prothrombin from plasma taken from one person, converting the prothrombin to thrombin, removing particulate material from the thrombin, and mixing the thrombin with clotting proteins to form a fibrin clot, but first altering time required for the thrombin to convert fibrinogen to a fibrin clot to a time of between about two seconds and about five seconds by adding an altering agent selected from the group including calcium ion sources, CaCl.sub.2, saline, and sterile water. 114. A method for extracting thrombin from one person, the steps consisting of: taking whole blood from one person, obtaining plasma from the whole blood, adding ethanol to the plasma to prepare a solution containing prothrombin, converting the prothrombin to thrombin, sequestering the thrombin, and mixing the thrombin with clotting proteins to form a fibrin clot, but first altering time required for the thrombin to convert fibrinogen to a fibrin clot to a time of between about two seconds and about five seconds by adding an altering agent selected from the group including calcium ion sources, CaCl.sub.2, saline, and sterile water. 115. A method for extracting autologous thrombin from a patient, the steps consisting essentially of: obtaining a blood product from the patient; sequestering plasma from the blood product; adding ethanol to the plasma to prepare a solution containing prothrombin; converting the prothrombin in the solution to thrombin; filtering the thrombin to remove particulate matter; applying the thrombin to the patient; and mixing the thrombin with clotting proteins to form a fibrin clot, but first altering time required for the thrombin to convert fibrinogen to a fibrin clot to a time of between about two seconds and about five seconds by adding an altering agent selected from the group including calcium ion sources, CaCl.sub.2, saline, and sterile water. 116. A method for extracting and then dispensing thrombin, the steps consisting essentially of: taking whole blood from a person, sequestering prothrombin from the whole blood by addition of ethanol, converting the prothrombin to thrombin, loading the thrombin into a syringe, using the syringe to dispense the thrombin to stem blood flow, and mixing the thrombin with clotting proteins to form a fibrin clot, but first altering time required for the thrombin to convert fibrinogen to a fibrin clot to a time of between about two seconds and about five seconds by adding an altering agent selected from the group including calcium ion sources, CaCl.sub.2, saline, and sterile water. 117. A method for extracting thrombin from one person, the steps consisting essentially of: adding ethanol to sequester prothrombin from plasma taken from one person, converting the prothrombin to thrombin, removing particulate material from the thrombin, and mixing the thrombin with clotting proteins to form a fibrin clot, but first altering time required for the thrombin to convert fibrinogen to a fibrin clot to a time of between about two seconds and about five seconds by adding an altering agent selected from the group including calcium ion sources, CaCl.sub.2, saline, and sterile water. 118. A method for extracting thrombin from one person, the steps consisting essentially of: taking whole blood from one person, obtaining plasma from the whole blood, adding ethanol to the plasma to prepare a solution containing prothrombin, converting the prothrombin to thrombin, sequestering the thrombin, and mixing the thrombin with clotting proteins to form a fibrin clot, but first altering time required for the thrombin to convert fibrinogen to a fibrin clot to a time of between about two seconds and about five seconds by adding an altering agent selected from the group including calcium ion sources, CaCl.sub.2, saline, and sterile water. 119. A method for extracting thrombin, the steps consisting essentially of: taking whole blood from a person, obtaining plasma from the whole blood, adding ethanol to the plasma to prepare a solution containing prothrombin, converting the prothrombin to thrombin, sequestering the thrombin, and mixing the thrombin with clotting proteins to form a fibrin clot, but first altering time required for the thrombin to convert fibrinogen to a fibrin clot to a time of between about two seconds and about five seconds by adding an altering agent selected from the group including calcium ion sources, CaCl.sub.2, saline, and sterile water.

Details for Patent 6,472,162

Applicant	Tradename	Biologic Ingredient	Dosage Form	BLA	Approval Date	Patent No.	Expiredate
Merck Sharp & Dohme Corp.	ZOSTAVAX	zoster vaccine live	For Injection	125123	05/25/2006		2039-09-23