Simple fixation and stabilisation

Abstract

It has surprisingly been discovered that it is possible to stabilize biomolecules such as RNA, DNA and proteins in biological samples such as cells, tissues, biopsies and blood using deep eutectic solvents (DES). It has also been discovered that DES mixtures can be used to fix and preserve cell morphology in biological samples such as tissue blocks, cancer biopsies and whole blood. This invention describes methods to stabilize and preserve biomolecules, whole cells, tissues, blood and biological samples using DES mixtures.

1 - 20 . (canceled) 
     
     
         21 . A method of inhibiting the degradation of a biomolecule selected from an RNA and a DNA, which method comprises mixing the biomolecule with, or immersing the biomolecule in, a eutectic solvent comprising a first component and a second component,
 wherein the first component comprises a compound of formula:   
       
         
           
           
               
               
           
         
         wherein:
 R 6  is H or OH; 
 R 7  is selected from H, CH 3 , Cl, Br, a carbonyl oxygen, and 
 
       
       
         
           
           
               
               
           
         
         and
 Z is selected from —CH 2 —, O and S; 
 R 8  is selected from OH, an alkyl group having one to three carbon atoms, 
 a monochloroalkyl group having one to three carbon atoms, and a mono-, di- or tri-fluoroalkyl group having one to three carbon atoms; 
 
         wherein the second component is a sugar or a sugar alcohol having at least 3 carbon atoms; and 
         wherein the method prevents degradation of the biomolecule. 
       
     
     
         22 . The method according to  claim 21 , wherein the first component is N,N,N-trimethyl glycine. 
     
     
         23 . The method according to  claim 21 , wherein the second component is a sugar alcohol. 
     
     
         24 . The method according to  claim 21 , wherein the sugar alcohol is selected from sorbitol, and xylitol. 
     
     
         25 . The method according to  claim 21 , wherein the first component and the second component are present at a molar ratio in the range 1:2 to 2:1. 
     
     
         26 . The method according to  claim 25 , wherein the first component and the second component are present at a molar ratio in the range 1:0.8 to 1:1.2. 
     
     
         27 . The method according to  claim 21 , wherein the deep eutectic solvent is free of metal salts. 
     
     
         28 . The method according to  claim 21 , wherein the eutectic solvent has a pH in the range 5 to 7.5. 
     
     
         29 . The method according to  claim 21 , wherein the eutectic solvent further comprises at least one additive selected from a colourant, a dye, a detergent, a quaternary ammonium salt, a saponin, an anti-microbial, a dessicant, a probe, an internal control, an antioxidant, a ribonuclease inhibitor, a buffer, a chelator, a dissolved gas, an alcohol, and a protein precipitant,
 wherein the at least one additive is present in the eutectic solvent in an amount in the range 0.01% to 1.5% by weight   
     
     
         30 . The method according to  claim 21 , wherein the first component is N,N,N-trimethylglycine and the second component is sorbitol. 
     
     
         31 . The method according to  claim 21 , wherein the first component is N,N,N-trimethylglycine and the second component is xylitol. 
     
     
         32 . The method according to  claim 21 , wherein the eutectic solvent further comprises water in an amount of up to 50% by weight of the eutectic solvent. 
     
     
         33 . A method of inhibiting the degradation of a biomolecule selected from an RNA and a DNA, which method comprises mixing the biomolecule with, or immersing the biomolecule in, a eutectic solvent comprising a first component and a second component,
 wherein the first component is proline;   wherein the second component is a sugar or a sugar alcohol having at least 3 carbon atoms; and   wherein the method prevents degradation of the biomolecule.   
     
     
         34 . The method according to  claim 33 , wherein the second component is selected from adonitol, ribitol, rhamnose, trehalose, sorbitol, sorbose, xylitol, glucose, sucrose, lactose, fructose, maltose, mannose, mannitol, arabinose, galactose, raffinose, inositol, erythritol and xylose. 
     
     
         35 . The method according to  claim 34 , wherein the second component is sorbitol. 
     
     
         36 . The method according to  claim 34 , wherein the second component is xylitol. 
     
     
         37 . The method according to  claim 34 , wherein the second component is arabitol. 
     
     
         38 . The method according to  claim 33 , wherein the deep eutectic solvent is free of metal salts. 
     
     
         39 . The method according to  claim 33 , wherein the first component and the second component are present at a molar ratio in the range 1:2 to 2:1.