Cell Suspension Preparation Technique and Device

Abstract

The present invention provides for methods and devices suitable for producing a transplantable cellular suspension of living tissue suitable for grafting to a patient. In applying the method and/or in using the device, donor tissue is harvested, subjected to a cell dissociation treatment, cells suitable for grafting back to a patient are collected and dispersed in a solution that is suitable for immediate dispersion over the recipient graft site.

1 . A method for preparing a cell suspension suitable for application to a patient, comprising the steps of:
 (a) harvesting cells suitable for grafting to a patient from a tissue sample from a first tissue region of the patient, wherein a cellular stratum in the tissue sample is physically and/or chemically dissociated, wherein the cells are harvested in a nutrient solution that is (i) free of serum xenogenic to the patient, (ii) capable of maintaining viability of the harvested cells, and (iii) suitable for direct application to a second tissue region of the patient; and   (b) filtering the harvested cells to remove cellular conglomerates greater than 200 μM,   thereby producing a cell suspension comprising viable cells autologous to the patient, wherein the cell suspension is free of serum xenogenic to the patient and free of cellular conglomerates greater than 200 μM.   
     
     
         2 . The method according to  claim 1  wherein the first tissue region is the same as the second tissue region. 
     
     
         3 . The method according to  claim 1  wherein the first tissue region is a different region than the second tissue region and is selected to match the second tissue region. 
     
     
         4 . The method according to  claim 1  further comprising dissociating the harvested cells in a solution comprising an enzyme. 
     
     
         5 . The method according to  claim 4  wherein the enzyme is selected from the group consisting of trypsin, trypsin-EDTA, dispase, collagenase, thermolysin, pronase, hyaluronidase, pancreatin, elastase and papain. 
     
     
         6 . The method according to  claim 1  wherein the nutrient solution is Hartmann's solution. 
     
     
         7 . The method according to  claim 1  further comprising multiplicating cells in situ. 
     
     
         8 . A method for delivering a cell suspension to a patient in need of graft surgery, said method comprising the steps of:
 (a) harvesting cells suitable for grafting to the patient from a tissue sample of the patient in a nutrient solution that is (i) free of serum xenogenic to the patient, (ii) capable of maintaining viability of the harvested cells, and (iii) suitable for direct application to a second tissue region of the patient;   (b) filtering the harvested cells to remove cellular conglomerates greater than 200 μM to produce a cell suspension; and   (c) administering the cell suspension to a tissue region of the patient in need of a cell graft.   
     
     
         9 . The method of  claim 8 , wherein said administering comprises spreading the cell suspension over the tissue region in a substantially even distribution. 
     
     
         10 . The method of  claim 9 , wherein said spreading comprises spraying or dripping the cell suspension through a nozzle. 
     
     
         11 . The method of  claim 8  further comprising multiplicating cells in situ. 
     
     
         12 . An apparatus for delivering cells to a patient in need of graft surgery, said apparatus comprising an aerosol bottle adapted to deliver a medium comprising a nutrient solution that is (i) free of serum xenogenic to the patient, (ii) capable of maintaining viability of a cell, and (iii) suitable for direct application to a tissue region of the patient; and wherein the medium further comprises cells that are harvested from a tissue sample of the patient and free of cellular conglomerates greater than 200 μM. 
     
     
         13 . The apparatus of  claim 12 , wherein the aerosol bottle comprises a syringe for collecting the medium and a nozzle attached to the syringe for spraying or dripping the medium to the tissue region of the patient. 
     
     
         14 . The apparatus of  claim 12  further comprising a filter adapted to remove cellular conglomerates greater than 200 μM. 
     
     
         15 . An apparatus for developing a solution for tissue regeneration, said apparatus comprising:
 (a) a heating means suitable for heating an enzyme solution to a predetermined temperature and capable of maintaining the enzyme solution at the predetermined temperature for a suitable amount of time; and   (b) a filter recess comprising a filter for removing cellular conglomerates greater than 200 μM.   
     
     
         16 . The apparatus according to  claim 15  further comprising a reservoir adapted to hold a tissue sample and a nutrient solution. 
     
     
         17 . The apparatus according to  claim 15  further comprising one or more fluid containment wells for storage of fluids. 
     
     
         18 . An apparatus for developing a solution for tissue regeneration, said comprising a first and second member, wherein:
 (i) the first member comprises:   (a) a heating means suitable for heating an enzyme solution to a predetermined temperature and capable of maintaining the enzyme solution at the predetermined temperature for a suitable amount of time;   (b) a filter recess comprising a filter for removing cellular conglomerates greater than 200 μM from a cellular suspension; and   (c) at least one fluid containment well for storage of a nutrient solution;   (ii) the second member comprises a reservoir capable of holding a tissue sample and a nutrient solution in one or more fluid containment wells;   and wherein the first member provides a seat upon which the second member is placed during manipulation of the tissue sample.   
     
     
         19 . A method for expanding a population of cells in situ, the method comprising:
 (a) harvesting cells suitable for grafting to a patient from a tissue sample of the patient, wherein a cellular stratum in the tissue sample is physically and/or chemically dissociated, wherein the cells are harvested in a nutrient solution that is (i) free of serum xenogenic to the patient, (ii) capable of maintaining viability of the harvested cells, and (iii) suitable for direct application to a second tissue region of the patient;   (b) filtering the harvested cells to remove cellular conglomerates greater than 200 μM to produce a cell suspension;   (c) applying the cell suspension to a tissue region of the patient in need of a cell graft; and   (d) permitting cells to expand in situ under conditions suitable for cellular multiplication.