Compositions for the rehydration of an electrophoresis support in order to improve zone electrophoresis

Abstract

The invention concerns a composition for use in a process for separating the constituents of a sample by electrophoresis on an electrophoresis support, comprising one or more ionic compounds which, on applying an electric field to an electrophoresis support having negative surface charges, causes hydration of the zone for loading the sample to be separated when said zone carries a compression mark resulting from loading the sample.

1 . A composition for use in electrophoresis, comprising: at least one ionic compound in an amount which, when applied to an electrophoresis support and upon application of an electric field, causes reswelling of a compression mark in a surface of said electrophoresis support resulting from loading of a sample thereon.  
     
     
         2 . The composition of  claim 1 , wherein said ionic compound is capable of reswelling a compression mark in a surface of an electrophoresis support having negative surface charges.  
     
     
         3 . The composition according to  claim 2 , wherein said ionic compound is a zwitterionic compound which will not undergo substantial displacement in an electric field.  
     
     
         4 . The composition according to  claim 3 , wherein said zwitterionic compound is an amino acid.  
     
     
         5 . The composition according to  claim 4 , wherein the formula of said amino acid is NH 2 —R—COOH, and wherein R comprises an uncharged polar side chain.  
     
     
         6 . The composition according to  claim 4 , wherein the formula of said amino acid is NH 2 —R—COOH, and wherein R comprises a basic side chain.  
     
     
         7 . The composition according to  claim 4 , wherein said amino acid is glycine, present in a concentration which is compatible with its solubility in water at 25° C., or phenylalanine in a concentration of about 0.28 M.  
     
     
         8 . The composition according to  claim 6 , wherein said amino acid is selected from lysine in a concentration of at least about 1 M, arginine in a concentration in the range of about 0.45 M to about 0.86 M, and histidine in a concentration of at most about 0.27 M.  
     
     
         9 . The composition according to  claim 3 , wherein said zwitterionic compound is a zwitterionic buffer.  
     
     
         10 . The composition according to  claim 9 , wherein said zwitterionic buffer is: 
 TRICINE (N-tris(hydroxymethyl)methyl-glycine);    HEPES (N-(2-hydroxyethyl)piperazine-N′-(2-ethane sulphonic acid));    PIPES (piperazine-N-N′-bis(2-ethane sulphonic acid);    MOPS (3-(N-morpholino)propane sulphonic acid);    TAPS (N-tris(hydroxymethyl)methyl-3-aminopropane sulphonic acid);    AMPSO (3-((1,1-dimethyl-2-hydroxyethyl)amino)-2-hydroxypropane sulphonic acid)    TES (N-tris(hydroxymethyl)methyl-2-aminoethane sulphonic acid);    BES (N,N′-bis(2-hydroxyethyl)-2-aminoethane sulphonic acid); 
 BICINE (5N,N-bis(2-hydroxyethyl)glycine);  
 CHES (2-(N-cyclohexylamino)ethane sulphonic acid);  
   DIPSO (3-(N,N-bis(2-hydroxyethyl)amino)-2-hydroxy-propane sulphonic acid;    EPPS (N-(2-hydroxyethyl)piperazine-N′-(3-propane sulphonic acid));    MOPSO (3-(N-morpholino)propane sulphonic acid); POPSO (piperazine-N,N′-bis(2′hydroxypropane sulphonic acid));    MES (2-morpholinoethane sulphonic acid);    or a salt of said buffers.    
     
     
         11 . The composition according to  claim 9 , in which the concentration of the zwitterionic buffer is more than about 0.3 M.  
     
     
         12 . The composition according to  claim 2 , wherein said ionic compound will migrate under the influence of an electrical field.  
     
     
         13 . The composition according to  claim 12 , wherein said ionic compound is a salt.  
     
     
         14 . The composition according to claims  12  or  13 , wherein said ionic compound is selected from NaCl, KCl or NaHCO 3  in a concentration of at least about 1 M, a boric acid salt in a concentration of at least about 0.15 M, a phosphoric acid salt or an acetic acid salt each in a concentration of at least about 0.5 M.  
     
     
         15 . A formulation comprising: a sample to be separated by electrophoresis mixed with a composition for use in electrophoresis, said composition including at least one ionic compound in an amount which, when applied to an electrophoresis support, causes reswelling of a compression mark in a surface of said electrophoresis support resulting from loading of a sample thereon upon application of an electric field.  
     
     
         16 . The formulation of  claim 15 , wherein said ionic compound is capable of reswelling a compression mark in a surface of an electrophoresis support having negative surface charges.  
     
     
         17 . The formulation according to  claim 16 , wherein said ionic compound is a zwitterionic compound which will not undergo substantial displacement in an electric field.  
     
     
         18 . The formulation according to  claim 17 , wherein said zwitterionic compound is an amino acid.  
     
     
         19 . The formulation according to  claim 18 , wherein the formula of said amino acid is NH 2 —R—COOH, and wherein R comprises an uncharged polar side chain.  
     
     
         20 . The formulation according to  claim 18 , wherein the formula of said amino acid is NH 2 —R—COOH, and wherein R comprises a basic side chain.  
     
     
         21 . The formulation according to  claim 18 , wherein said amino acid is glycine, present in a concentration which is compatible with its solubility in water at 25° C., or phenylalanine in a concentration of about 0.28 M.  
     
     
         22 . The formulation according to  claim 18 , wherein said amino acid is selected from lysine in a concentration of at least about 1 M, arginine in a concentration in the range of about 0.45 M to about 0.86 M, and histidine in a concentration of at most about 0.27 M.  
     
     
         23 . The formulation according to  claim 16 , wherein said ionic compound is a zwitterionic buffer.  
     
     
         24 . The formulation according to  claim 23 , wherein said zwitterionic buffer is: 
 TRICINE (N-tris(hydroxymethyl)methyl-glycine);    HEPES (N-(2-hydroxyethyl)piperazine-N′-(2-ethane sulphonic acid));    PIPES (piperazine-N-N′-bis(2-ethane sulphonic acid);    MOPS (3-(N-morpholino)propane sulphonic acid);    TAPS (N-tris(hydroxymethyl)methyl-3-aminopropane sulphonic acid);    AMPSO (3-((1,1-dimethyl-2-hydroxyethyl)amino)-2-hydroxypropane sulphonic acid)    TES (N-tris(hydroxymethyl)methyl-2-aminoethane sulphonic acid);    BES (N,N′-bis(2-hydroxyethyl)-2-aminoethane sulphonic acid);    BICINE (5N,N-bis(2-hydroxyethyl)glycine);    CHES (2-(N-cyclohexylamino)ethane sulphonic acid);    DIPSO (3-(N,N-bis(2-hydroxyethyl)amino)-2-hydroxy-propane sulphonic acid;    EPPS (N-(2-hydroxyethyl)piperazine-N′-(3-propane sulphonic acid));    MOPSO (3-(N-morpholino)propane sulphonic acid); POPSO (piperazine-N,N′-bis(2′hydroxypropane sulphonic acid));    MES (2-morpholinoethane sulphonic acid);    or a salt of said buffers.    
     
     
         25 . The formulation according to  claim 23 , in which the concentration of the zwitterionic buffer is more than about 0.3 M.  
     
     
         26 . The formulation of  claim 16 , wherein said ionic compound will migrate under the influence of an electrical field.  
     
     
         27 . The formulation according to  claim 16 , wherein said ionic compound is a salt.  
     
     
         28 . The formulation according to claims  26  or  27 , wherein said salt is selected from NaCl, KCl or NaHCO 3  in a concentration of at least about 1 M, a boric acid salt in a concentration of at least about 0.15 M, a phosphoric acid salt or an acetic acid salt each in a concentration of at least about 0.5 M.  
     
     
         29 . A kit for carrying out a process for separating sample constituents by electrophoresis, which comprises: 
 a composition for use in electrophoresis, which includes at least one ionic compound in an amount which, when applied to an electrophoresis support and upon application of an electric field, causes reswelling of a compression mark in a surface of said electrophoresis support resulting from loading of a sample thereto; and    an electrophoresis support gel with a non zero electroosmotic flow.    
     
     
         30 . The kit according to  claim 29 , wherein said electrophoresis support is an agarose gel.  
     
     
         31 . The kit according to  claim 29 , wherein said electrophoresis support has a negative surface charge.  
     
     
         32 . The kit according to  claim 31 , wherein said ionic compound is a zwitterionic compound which will not undergo substantial displacement in an electric field.  
     
     
         33 . The composition according to  claim 32 , wherein said zwitterionic compound is an amino acid.  
     
     
         34 . The composition according to  claim 33 , wherein the formula of said amino acid is NH 2 —R—COOH, and wherein R comprises an uncharged polar side chain.  
     
     
         35 . The composition according to  claim 33 , wherein the formula of said amino acid is NH 2 —R—COOH, and wherein R comprises a basic side chain.  
     
     
         36 . The composition according to  claim 33 , wherein said amino acid is glycine, present in a concentration which is compatible with its solubility in water at 25° C., or phenylalanine in a concentration of about 0.28 M.  
     
     
         37 . The composition according to  claim 33 , wherein said amino acid is selected from lysine in a concentration of at least about 1 M, arginine in a concentration in the range of about 0.45 M to about 0.86 M, and histidine in a concentration of at most about 0.27 M.  
     
     
         38 . The composition according to  claim 32 , wherein said zwitterionic compound is a zwitterionic buffer.  
     
     
         39 . The composition according to  claim 38 , wherein said zwitterionic buffer is: 
 TRICINE (N-tris(hydroxymethyl)methyl-glycine);    HEPES (N-(2-hydroxyethyl)piperazine-N′-(2-ethane sulphonic acid));    PIPES (piperazine-N-N′-bis(2-ethane sulphonic acid);    MOPS (3-(N-morpholino)propane sulphonic acid);    TAPS (N-tris(hydroxymethyl)methyl-3-aminopropane sulphonic acid);    AMPSO (3-((1,1-dimethyl-2-hydroxyethyl)amino)-2-hydroxypropane sulphonic acid)    TES (N-tris(hydroxymethyl)methyl-2-aminoethane sulphonic acid);    BES (N,N′-bis(2-hydroxyethyl)-2-aminoethane sulphonic acid);    BICINE (5N,N-bis(2-hydroxyethyl)glycine);    CHES (2-(N-cyclohexylamino)ethane sulphonic acid);    DIPSO (3-(N,N-bis(2-hydroxyethyl)amino)-2-hydroxy-propane sulphonic acid;    EPPS (N-(2-hydroxyethyl)piperazine-N′-(3-propane sulphonic acid));    MOPSO (3-(N-morpholino)propane sulphonic acid); POPSO (piperazine-N,N′-bis(2′hydroxypropane sulphonic acid));    MES (2-morpholinoethane sulphonic acid);    or a salt of said buffers.    
     
     
         40 . The composition according to  claim 38 , in which the concentration of the zwitterionic buffer is more than about 0.3 M.  
     
     
         41 . The composition according to  claim 31 , wherein said ionic compound will migrate under the influence of an electrical field.  
     
     
         42 . The composition according to  claim 41 , wherein said ionic compound is a salt.  
     
     
         43 . The composition according to claims  41  or  42 , wherein said salt is selected from NaCl, KCl or NaHCO 3  in a concentration of at least about 1 M, a boric acid salt in a concentration of at least about 0.15 M, a phosphoric acid salt or an acetic acid salt each in a concentration of at least about 0.5 M.  
     
     
         44 . The composition according to  claim 1 , wherein said at least one ionic compound includes both a zwitterionic compound and a salt.  
     
     
         45 . The composition according to  claim 3 , further comprising salt.  
     
     
         46 . The composition according to  claim 13 , further comprising a zwitterionic compound.  
     
     
         47 . The composition according to  claim 2 , further comprising an ionic compound which will migrate under the influence of an electrical field.