Method for preparing cell cultures from biological specimens for chemotherapeutic and other assays

Abstract

An improved method for preparing a cell culture is disclosed. The method includes culturing a multicellular tissue explant in the presence of growth medium that is substantially free of enzymes capable of digesting the explant and, subsequently, removing the explant at a predetermined time.

What is claimed is:  
     
         1 . A method for preparing a cell culture, the method comprising: 
 culturing a multicellular tissue explant in the presence of growth medium that is substantially free of enzymes capable of digesting said explant; and    removing said multicellular tissue explant from said growth medium at a predetermined time,    thereby to produce a cell culture monolayer.    
     
     
         2 . The method of  claim 1 , further comprising preparing a cell suspension from said cell culture monolayer.  
     
     
         3 . The method of  claim 1 , further comprising inoculating cells from said cell culture monolayer into at least one segregated site.  
     
     
         4 . The method of  claim 1 , wherein said tissue explant comprises tumor tissue.  
     
     
         5 . The method of  claim 1 , further comprising the step of mechanically fragmenting the multicellular tissue explant in a medium that is free of enzymes that are capable of digesting said explant.  
     
     
         6 . The method of  claim 1 , comprising the step of removing said multicellular tissue explant from said growth medium prior to an emergence of a substantial number of stromal cells from said explant, thereby to produce a cell culture monolayer that is predominantly composed of non-stromal cells.  
     
     
         7 . The method of  claim 6  wherein said non-stromal cells comprise tumor cells.  
     
     
         8 . The method of  claim 1 , wherein the cell culture monolayer exhibits a percent confluency, and wherein the step of removing said multicellular tissue explant comprises removing said multicellular tissue explant at about 10 to about 50 percent confluency.  
     
     
         9 . The method of  claim 8 , wherein the step of removing said multicellular tissue explant comprises removing said multicellular tissue explant at about 15 to about 25 percent confluency.  
     
     
         10 . The method of  claim 8 , wherein the step of removing said multicellular tissue explant comprises removing said multicellular tissue explant at about 20 percent confluency.  
     
     
         11 . A cell culture preparation resulting from the method of  claim 1  or  10 .  
     
     
         12 . A method for determining the chemosensitivity of a tissue in a patient to an agent, the method comprising the steps of: 
 (a) culturing a multicellular tissue explant from a patient in the presence of growth medium that is substantially free of enzymes capable of digesting said explant so as to produce a cell culture monolayer;    (b) removing said multicellular tissue explant from said growth medium at a predetermined time;    (d) inoculating cells from said cell culture monolayer into at least one segregated site;    (e) exposing said segregated site to at least one agent; and    (f) assessing the chemosensitivity of said cells in said segregated site, wherein said chemosensitivity of said cells is indicative of the chemosensitivity of the tissue in the patient.    
     
     
         13 . The method of  claim 12 , wherein said multicellular tissue explant comprises tumor tissue.  
     
     
         14 . The method of  claim 13 , wherein the cell culture monolayer exhibits a percent confluency, and wherein the step of removing said multicellular tissue explant comprises removing said multicellular tissue explant at about 10 to about 50 percent confluency.  
     
     
         15 . The method of  claim 14 , wherein the step of removing said multicellular tissue explant comprises removing said multicellular tissue explant at about 15 to about 25 percent confluency.  
     
     
         16 . The method of  claim 14 , wherein the step of removing said multicellular tissue explant comprises removing said multicellular tissue explant at about 20 percent confluency.  
     
     
         17 . A method for determining the efficacy of an agent, the method comprising the steps of: 
 (a) preparing a cell culture in accordance with  claim 3;     (b) exposing said segregated site to at least one agent; and    (c) assessing the chemosensitivity of said cells in said segregated site, thereby to determine the efficacy of said agent.    
     
     
         18 . The method of  claim 17 , wherein the cell culture monolayer exhibits a percent confluency, and wherein the step of removing said multicellular tissue explant comprises removing said multicellular tissue explant at about 15 to about 25 percent confluency.  
     
     
         19 . The method of  claim 18 , wherein the step of removing said multicellular tissue explant comprises removing said multicellular tissue explant at about 20 percent confluency.  
     
     
         20 . A method for identifying an agent having anti-tumorogenic effect, the method comprising the steps of: 
 (a) preparing a cell culture in accordance with  claim 4;     (b) inoculating cells from said cell culture monolayer into a plurality of segregated sites    (c) exposing each of said segregated sites to an agent; and    (d) assessing the chemosensitivity of said cells in each of said segregated sites, thereby to identify an agent having an anti-tumorogenic effect.