Non-pyrogenic, endotoxin-frei stroma-free tetrameric hemoglobin

Abstract

A non-pyrogenic, endotoxin-free, stroma-free, blood substitute capable of being used in humans or other mammals to provide oxygen to tissue and to deliver carbon dioxide to the lung and a process for its preparation are described.

1 - 18 . (canceled).  
     
     
         19 . A preparation of non-pyrogenic, endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin produced by the process comprising the steps of: 
 (A) removing endotoxin from a preparation containing red blood cells;    (B) removing oxygen from said preparation containing red blood cells, wherein said oxygen is removed by centrifuging the red blood cells under a vacuum sufficient to remove oxygen from the preparation;    (C) lysing the red blood cells.    
     
     
         20 . (canceled)  
     
     
         21 . The preparation of non-pyrogenic, endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin of  claim 19 , wherein said process additionally comprises the steps of: 
 (D) separating hemoglobin from the stroma of said lysed red blood cells; and    (E) cross-linking said separated hemoglobin.    
     
     
         22 . The preparation of non-pyrogenic, endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin of  claim 21 , wherein said process step (A) additionally comprises washing surfaces and equipment that will come into contract with the cross-linked hemoglobin with a dilute solution of a hemoglobin.  
     
     
         23 . The preparation of endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin of  claim 19 , wherein said process step (B) comprises centrifuging a solution of said cells under vacuum at a speed sufficient to produce a force greater than the surface tension of the solution.  
     
     
         24 . The preparation of endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin of  claim 21 , wherein said red blood cells are human red blood cells.  
     
     
         25 . The preparation of endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin of  claim 21 , wherein said red blood cells are bovine or porcine red blood cells.  
     
     
         26 . The preparation of endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin of  claim 21 , wherein said preparation additionally contains a pharmaceutically acceptable carrier.  
     
     
         27 . A preparation of non-pyrogenic, endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin produced by the process comprising the steps of: 
 (A) removing endotoxin from a preparation containing red blood cells;    (B) lysing red blood cells; and    (C) removing oxygen from hemoglobin of said lysed red blood cells, wherein said oxygen is removed by centrifuging the red blood cells under a vacuum sufficient to remove oxygen from the preparation.    
     
     
         28 . (canceled).  
     
     
         29 . The preparation of non-pyrogenic, endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin of  claim 27 , wherein said process additionally comprises the steps of: 
 (D) separating hemoglobin from the stroma of said lysed red blood cells; and    (E) cross-linking said separated hemoglobin.    
     
     
         30 . The preparation of non-pyrogenic, endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin of  claim 27 , wherein said process step (A) additionally comprises washing surfaces and equipment that will come into contact with the cross-linked hemoglobin with a dilute solution of a hemoglobin.  
     
     
         31 . The preparation of endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin of  claim 27 , wherein said process step (C) comprises centrifuging a solution of said cells under vacuum at a speed sufficient to produce a force greater than the surface tension of the solution.  
     
     
         32 . The preparation of endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin of  claim 27 , wherein said red blood cells are human red blood cells.  
     
     
         33 . The preparation of endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin of  claim 27 , wherein said red blood cells are bovine or porcine red blood cells.  
     
     
         34 . The preparation of endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin of  claim 27 , wherein said preparation additionally contains a pharmaceutically acceptable carrier.  
     
     
         35 - 48 . (canceled).  
     
     
         49 . A method of increasing the oxygen carrying capacity of an individual which comprises administering to said individual a non-pyrogenic, endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin administered by transfusion or injection, wherein said non-pyrogenic, endotoxin-free, stroma-free, cross-linked tetrameric hemoglobin is produced by a process comprising the steps: 
 (A) removing endotoxin from a preparation containing red blood cells;    (B) removing oxygen from said preparation containing red blood cells, wherein said oxygen is removed by centrifuging the red blood cells under a vacuum sufficient to remove oxygen from the preparation; and    (C) lysing red blood cells.    
     
     
         50 - 52 . (canceled).  
     
     
         53 . The method of  claim 49 , wherein said process additionally comprises the steps of: 
 (D) separating hemoglobin from the stroma of said lysed red blood cells; and    (E) cross-linking said separated hemoglobin.    
     
     
         54 . The method of  claim 49 , wherein said process step (A) additionally comprises washing surfaces and equipment that will come into contact with the cross-linked tetrameric hemoglobin with a dilute solution of hemoglobin.  
     
     
         55 . The method of  claim 49 , wherein said process step (B) additionally comprises centrifuging a solution of said cells under vacuum at a speed sufficient to produce a force greater than the surface tension of the solution.  
     
     
         56 . The method of  claim 53 , wherein said hemoglobin is human hemoglobin.  
     
     
         57 . The method of  claim 53 , wherein said hemoglobin is bovine or porcine hemoglobin.  
     
     
         58 - 60 . (canceled).  
     
     
         61 . The method of  claim 56 , wherein said process step (A) comprises washing surfaces and equipment that will come into contact with the cross-linked tetrameric hemoglobin with a dilute solution of hemoglobin.  
     
     
         62 . The method of  claim 56 , wherein said process step (C) additionally comprises centrifuging a solution of said cells under vacuum at a speed sufficient to produce a force greater than the surface tension of the solution.  
     
     
         63 . The method of  claim 53 , wherein said hemoglobin is human hemoglobin.  
     
     
         64 . The method of  claim 53 , wherein said hemoglobin is bovine or porcine hemoglobin.  
     
     
         65 - 71 . (canceled).