Apparatus and method for separating and concentrating fluids containing multiple components

Abstract

An apparatus that allows for separating and collecting a fraction of a sample. The apparatus, when used with a centrifuge, allows for the creation of at least three fractions in the apparatus. It also provides for a new method of extracting the buffy coat phase from a whole blood sample. A buoy system that may include a first buoy portion and a second buoy member operably interconnected may be used to form at least three fractions from a sample during a substantially single centrifugation process. Therefore, the separation of various fractions may be substantially quick and efficient.

1 . A method for concentrating bone marrow aspirate, the method comprising: 
 obtaining a volume of bone marrow aspirate from a mammal;    loading the volume of bone marrow aspirate into a separator comprising two buoys, the separator operable to separate the aspirate into three or more fractions;    centrifuging the separator creating at least one fraction comprising a concentrated bone marrow aspirate; and    extracting the at least one fraction comprising a concentrated bone marrow aspirate.    
     
     
         2 . The method according to  claim 1  wherein the at least one fraction comprises at least one of the group consisting essentially of hematopoietic stem cells, stromal stem cells, mesenchymal stem cells, endothelial progenitor cells, red blood cells, white blood cells, fibroblasts, reticulacytes, adipose cells, and endothelial cells.  
     
     
         3 . The method according to  claim 1  wherein an amount of nucleated cells in the at least one fraction is greater than or equal to 4 times an amount of nucleated cells found in the volume of bone marrow aspirate.  
     
     
         4 . The method according to  claim 1  further comprising applying the at least one fraction into a patient.  
     
     
         5 . A method for concentrating bone marrow aspirate and peripheral blood, the method comprising: 
 collecting a volume of bone marrow aspirate and a volume of peripheral    blood from a patient;    loading the volume of bone marrow aspirate and the volume of peripheral blood into a separator comprising two buoys, separator being operable to separate a combination of the volume of aspirate and the volume of blood into three or more fractions;    centrifuging the at least one separator operably creating at least one fraction comprising a concentration of at least one of bone marrow aspirate and peripheral blood; and    withdrawing the at least one fraction comprising the concentration.    
     
     
         6 . The method according to  claim 5  wherein the concentration comprises at least one of the group consisting essentially of hematopoietic stem cells, stromal stem cells, mesenchymal stem cells, endothelial progenitor cells, red blood cells, white blood cells, fibroblasts, reticulacytes, adipose cells, and endothelial cells.  
     
     
         7 . The method according to  claim 5  wherein an amount of nucleated cells in the concentration is greater than or equal to 4 times an amount of nucleated cells in the combination of the volume of aspirate and the volume of blood.  
     
     
         8 . A method for treating a defect in a mammal using a concentrated bone marrow aspirate, the method comprising: 
 drawing a volume of bone marrow aspirate from the mammal;    loading the volume of bone marrow aspirate into a separator comprising two buoys, the separator being operable to separate the aspirate into three or more fractions;    centrifuging the separator;    separating the volume of bone marrow aspirate into a plurality of fractions including a fraction comprising a concentrated bone marrow aspirate;    extracting the fraction comprising a concentrated bone marrow aspirate; and    applying the fraction comprising a concentrated bone marrow aspirate to site of the defect.    
     
     
         9 . The method according to  claim 8  wherein the fraction comprising a concentrated bone marrow aspirate is essentially buffy coat including a plurality of undifferentiated cells.  
     
     
         10 . The method according to  claim 8  wherein the fraction comprises at least one of the group consisting of hematopoietic stem cells, stromal stem cells, mesenchymal stem cells, endothelial progenitor cells, red blood cells, white blood cells, fibroblasts, reticulacytes, adipose cells, and endothelial cells.  
     
     
         11 . The method according to  claim 8  further comprising combining the fraction with a carrier.  
     
     
         12 . The method according to  claim 11  wherein the carrier is selected from the group consisting essentially of collagen, hydrogel, a bioabsorbable polymer, a biopolymer, water, buffered solution, fibrin, concentrated fibrin, demineralized bone matrix, gelatin, porous calcium based ceramics, porous metal, synthetic fiber matrices, resorbable matrices, autogeneic tissue, allogenic tissue, xenogeneic tissue, and combinations thereof.  
     
     
         13 . The method according to  claim 8  further comprising combining an activating agent with the fraction.  
     
     
         14 . The method according to  claim 8  further comprising combining a pharmaceutical agent with the fraction.  
     
     
         15 . A method for treating a defect in an animal, the method comprising: 
 obtaining a volume of bone marrow aspirate and a volume of whole blood from the animal;    loading the volume of bone marrow aspirate into a first separator comprising two buoys, the separator being operable to separate the aspirate into three or more fractions;    loading the volume of whole blood into a second separator comprising two buoys, the separator being operable to separate the aspirate into three or more fractions;    centrifuging the first separator and the second separators, thereby separating the volume of bone marrow aspirate into a plurality of fractions and separating the volume of whole blood into a plurality of fractions.    collecting a fraction of the bone marrow aspirate;    collecting a fraction of the whole blood; and    applying at least one of the fraction of bone marrow aspirate and the fraction of whole blood to site of the defect.    
     
     
         16 . The method according to  claim 15  wherein the at least one of the fraction of bone marrow aspirate and the fraction of whole blood comprises at least one of the group consisting of hematopoietic stem cells, stromal stem cells, mesenchymal stem cells, endothelial progenitor cells, red blood cells, white blood cells, fibroblasts, reticulacytes, adipose cells, and endothelial cells.  
     
     
         17 . The method according to  claim 15  further comprising combining the at least one of the fraction of bone marrow aspirate and the fraction of whole blood with a carrier.  
     
     
         18 . The method according to  claim 17  wherein the carrier is selected from the group consisting essentially of collagen, hydrogel, a bioabsorbable polymer, a biopolymer, water, buffered solution, fibrin, concentrated fibrin, demineralized bone matrix, gelatin, porous calcium based ceramics, porous metal, synthetic fiber matrices, resorbable matrices, autogeneic tissue, allogenic tissue, xenogeneic tissue, and combinations thereof.  
     
     
         19 . The method according to  claim 17  further comprising adding an activating agent to the at least one of the fraction of bone marrow aspirate and the fraction of whole blood  
     
     
         20 . The method according to  claim 17  further comprising adding a pharmaceutical agent to the at least one of the fraction of bone marrow aspirate and the fraction of whole blood.  
     
     
         21 . A method for treating a defect in a patient, the method comprising: 
 drawing a volume of bone marrow aspirate and a volume of whole blood from the patient;    adding a first anticoagulant to the volume of bone marrow aspirate;    adding a second anticoagulant to the volume of whole blood;    loading the volume of bone marrow aspirate and the volume of whole blood into a separator comprising two buoys, the separator being operable to separate the volume of bone marrow aspirate and the volume of whole blood into three or more fractions;    centrifuging the separator, separating the volume of bone marrow aspirate and the volume of whole blood into a plurality of fractions.    withdrawing a fraction comprising at least one of the group consisting of hematopoietic stem cells, stromal stem cells, mesenchymal stem cells, endothelial progenitor cells, red blood cells, white blood cells, fibroblasts, reticulacytes, adipose cells, and endothelial cells.; and    applying the fraction to site of the defect.    
     
     
         22 . The method according to  claim 21  wherein the fraction is essentially buffy coat including a plurality of undifferentiated cells.  
     
     
         23 . The method according to  claim 21  further comprising combining the fraction with a carrier.  
     
     
         24 . The method according to  claim 32  wherein the carrier is selected from the group consisting essentially of collagen, hydrogel, a bioabsorbable polymer, a biopolymer, water, buffered solution, fibrin, concentrated fibrin, demineralized bone matrix, gelatin, porous calcium based ceramics, porous metal, synthetic fiber matrices, resorbable matrices, autogeneic tissue, allogenic tissue, xenogeneic tissue, and combinations thereof.  
     
     
         25 . The method according to  claim 21  further comprising combining an activating agent with the fraction.  
     
     
         26 . The method according to  claim 21  further comprising combining a pharmaceutical agent with the fraction.  
     
     
         27 . A method of treating a patient with combination of a concentrated bone marrow aspirate and buffy coat, the method comprising: 
 obtaining a volume of a whole blood from the patient;    obtaining a volume of a bone marrow aspirate from the patient;    forming a buffy coat fraction of the whole blood;    forming a concentrated bone marrow aspirate fraction of the bone marrow aspirate; and    applying at least one of the buffy coat fraction or the concentrated bone marrow aspirate fraction to the patient.    
     
     
         28 . The method according to  claim 27 , wherein the forming a first fraction of the first whole material and the forming a second fraction of the second whole material includes: 
 loading the volume of whole blood and the volume of bone marrow aspirate in a container; and    applying a force to the container to form at least three fractions.    
     
     
         29 . The method according to  claim 27 , wherein the forming a first fraction of the first whole material and the forming a second fraction of the second whole material includes: 
 loading the volume of whole blood and the volume of bone marrow aspirate in the container having a separating member, the separating member having a specific gravity substantially dependent upon at least one of the at least three fractions; and    centrifuging the container to move the separating member to a selected position relative to the at least one of the volume of whole blood and the volume of bone marrow aspirate to substantially physically separate the at least three fractions.    
     
     
         30 . The method according to  claim 27  wherein the at least one of the first fraction or the second fraction comprises at least one of the group consisting of hematopoietic stem cells, stromal stem cells, mesenchymal stem cells, endothelial progenitor cells, red blood cells, white blood cells, fibroblasts, reticulacytes, adipose cells, and endothelial cells.  
     
     
         31 . The method according to  claim 27  further comprising combining the at least one of the first fraction or the second fraction with a carrier.  
     
     
         32 . The method according to  claim 31  wherein the carrier is selected from the group consisting essentially of collagen, hydrogel, a bioabsorbable polymer, a biopolymer, water, buffered solution, fibrin, concentrated fibrin, demineralized bone matrix, gelatin, porous calcium based ceramics, porous metal, synthetic fiber matrices, resorbable matrices, autogeneic tissue, allogenic tissue, xenogeneic tissue, and combinations thereof.