Method of preparing a standard diagnostic gene transcript pattern

Abstract

A method for preparing a gene transcript pattern probe kit characteristic of a disease or condition or a stage thereof in a prokaryotic or eukaryotic organism using mRNA which is differentially expressed in the disease or condition or stage as probes, methods of diagnosis using the method and kits for performing the same are disclosed.

1 - 16 . (canceled)  
     
     
         17 . A method of obtaining isolated selected mRNA species or isolated selected cDNA species useful for diagnosing or identifying a disease or condition or a stage thereof in a human comprising the steps of: 
 (a) isolating mRNA from cells from blood of one or more humans which are known to have the disease or condition or a stage thereof (disease sample), wherein said cells have not contacted the area of said disease or condition and said blood is obtained from a part of said human distant to the area of said disease or condition, wherein the resulting isolated mRNA is optionally subjected to reverse transcription to obtain isolated cDNA;    (b) isolating mRNA from corresponding cells from blood of one or more corresponding normal humans (normal sample), wherein the resulting isolated mRNA is optionally subjected to reverse transcription to obtain isolated cDNA;    (c) separating mRNA species or cDNA species present within each of the resulting isolated mRNA or isolated cDNA of step (a) and step (b), wherein the resulting separated mRNA species are optionally subject to reverse transcription to obtain separated cDNA species;    (d) selecting 10 or more mRNA species or 10 or more cDNA species from the resulting separated mRNA species or resulting separated cDNA species obtained in step (c), respectively, which are present at a different level in the normal sample than in the disease sample by identifying a signal corresponding to each mRNA species or cDNA species, wherein the resulting selected 10 or more mRNA species are optionally subjected to reverse transcription to obtain 10 or more selected cDNA species; and    (e) isolating the resulting 10 or more selected mRNA species or resulting 10 or more selected cDNA species obtained in step (d) to obtain isolated selected mRNA species or isolated selected cDNA species, wherein the resulting isolated selected mRNA species are optionally subjected to reverse transcription to obtain isolated selected cDNA species.    
     
     
         18 . A method of preparing a gene transcript pattern probe kit comprising the steps of: 
 (a) isolating mRNA from cells from blood of one or more humans which are known to have a disease or condition or a stage thereof (disease sample), wherein said cells have not contacted the area of said disease or condition and said blood is obtained from a part of said human distant to the area of said disease or condition, wherein the resulting isolated mRNA is optionally subjected to reverse transcription to obtain isolated cDNA;    (b) isolating mRNA from corresponding cells from blood of one or more corresponding normal humans (normal sample), wherein the resulting isolated mRNA is optionally subjected to reverse transcription to obtain isolated cDNA;    (c) separating mRNA species or cDNA species present within each of the resulting isolated mRNA or isolated cDNA of step (a) and step (b), wherein the resulting separated mRNA species are optionally subject to reverse transcription to obtain separated cDNA species;    (d) selecting 10 or more mRNA species or 10 or more cDNA species from the resulting separated mRNA species or resulting separated cDNA species obtained in step (c), respectively, which are present at a different level in the normal sample than in the disease sample by identifying a signal corresponding to each mRNA species or cDNA species, wherein the resulting selected 10 or more mRNA species are optionally subjected to reverse transcription to obtain 10 or more selected cDNA species;    (e) isolating the resulting 10 or more selected mRNA species or resulting 10 or more selected cDNA species obtained in step (d) to obtain isolated selected mRNA species or isolated selected cDNA species, wherein the resulting isolated selected mRNA species are optionally subjected to reverse transcription to obtain isolated selected cDNA species; and    (f) immobilizing the resulting isolated selected mRNA species or isolated selected cDNA species of step (e) on at least one solid support so as to form a gene transcript pattern probe kit.    
     
     
         19 . The method as claimed in  claim 17  or  18 , wherein said separation in step (c) is performed by a non-sequence based separation technique.  
     
     
         20 . The method as claimed in  claim 17  or  18 , wherein in steps (a) and (b), the resulting isolated mRNA is subjected to reverse transcription to obtain isolated cDNA.  
     
     
         21 . The method as claimed in  claim 20 , wherein said isolated cDNA is amplified.  
     
     
         22 . The method as claimed in  claim 17  or  18 , wherein in step (e), between 50 and 100 mRNA species or cDNA species are isolated and selected.  
     
     
         23 . The method as claimed in  claim 17  or  18 , wherein, in step (e), between 10 and 500 mRNA species or cDNA species are isolated and selected.  
     
     
         24 . The method as claimed in  claim 17  or  18 , wherein, in step (c), said separation technique is gel electrophoresis.  
     
     
         25 . The method as claimed in  claim 18 , wherein, prior to immobilizing in step (f), the resulting isolated selected mRNA species or isolated selected cDNA species of step (e) are amplified.  
     
     
         26 . A method of preparing a standard gene transcript pattern characteristic of a disease or condition or a stage thereof of a human comprising the steps of: 
 (a) isolating mRNA from cells from blood of one or more humans known to have said disease or condition or a stage thereof (disease sample), wherein said blood is obtained from a part of said human distant to the area of said disease or condition, wherein the resulting isolated mRNA is optionally subjected to reverse transcription to obtain isolated cDNA;    (b) hybridizing the resulting isolated mRNA or isolated cDNA of step (a) to 10 or more mRNA species which are present at a different level in cells in a blood sample from one or more normal humans than in corresponding cells in a blood sample from one or more humans known to have said disease or condition or a stage thereof, or to 10 or more cDNA species transcribed from said mRNA species, wherein the 10 or more mRNA species or cDNA species are specific for said disease or condition or stage thereof and wherein said cells have not contacted the area of said disease or condition and said blood is obtained from a part of said human distant to the area of said disease or condition, wherein said mRNA or cDNA species are immobilized on a solid support; and    (c) assessing the amount of hybridization so as to obtain said standard gene transcript pattern.    
     
     
         27 . A method of preparing a human test gene transcript pattern for a disease or condition or a stage thereof comprising the steps of: 
 (a) isolating mRNA from cells from blood of a test human, wherein said blood is obtained from a part of said human distant to the area of said disease or condition, wherein the resulting isolated mRNA is optionally subjected to reverse transcription to obtain isolated cDNA;    (b) hybridizing the resulting isolated mRNA or isolated cDNA of step (a) to 10 or more mRNA species which are present at a different level in cells in a blood sample from one or more normal humans than in corresponding cells in a blood sample from one or more humans known to have said disease or condition or a stage thereof, or to 10 or more cDNA species transcribed from said mRNA species, wherein the 10 or more mRNA species or cDNA species are specific for said disease or condition or stage thereof and wherein said cells have not contacted the area of said disease or condition and said blood is obtained from a part of said human distant to the area of said disease or condition, wherein said mRNA or cDNA species are immobilized on a solid support; and    (c) assessing the amount of hybridization so as to obtain said test gene transcript pattern.    
     
     
         28 . A method of diagnosing or identifying a disease or condition or a stage thereof in a test human comprising the steps of: 
 (a) isolating mRNA from cells from blood of a test human, wherein said blood is obtained from a part of said human distant to the area of said disease or condition, wherein the resulting isolated mRNA is optionally subjected to reverse transcription to obtain isolated cDNA;    (b) hybridizing the resulting isolated mRNA or isolated cDNA of step (a) to 10 or more mRNA species which are present at a different level in cells in a blood sample from one or more normal humans than in corresponding cells in a blood sample from one or more humans known to have said disease or condition or a stage thereof, or to 10 or more cDNA species transcribed from said mRNA species, wherein the 10 or more mRNA species or cDNA species are specific for said disease or condition or stage thereof and wherein said cells have not contacted the area of said disease or condition and said blood is obtained from a part of said human distant to the area of said disease or condition, wherein said mRNA or cDNA species are immobilized on a solid support;    (c) assessing the amount of hybridization so as to obtain a hybridization pattern; and    (d) comparing the resulting hybridization pattern obtained in step (c) with a hybridization pattern obtained by hybridizing isolated mRNA or isolated cDNA prepared from corresponding cells from blood of one or more corresponding humans known to have said disease or condition or stage thereof to the 10 or more mRNA species or 10 or more cDNA species transcribed from said mRNA species, wherein said mRNA species or cDNA species are immobilized on a solid support, so as to determine the degree of correlation indicative of the presence of said disease or condition or stage thereof, and so as to diagnose or identify said disease or condition or a stage thereof in said test human.    
     
     
         29 . The method as claimed in  claim 26 ,  27  or  28 , wherein in step (a), the resulting isolated mRNA is subjected to reverse transcription to obtain isolated cDNA.  
     
     
         30 . The method as claimed in  claim 29 , wherein said isolated cDNA is amplified.  
     
     
         31 . The method as claimed in  claim 17 ,  18 ,  26 ,  27  or  28 , wherein said isolated cDNA is labelled.  
     
     
         32 . The method as claimed in  claim 26 ,  27  or  28 , wherein, in step (b), between 50 and 100 mRNA species or cDNA species are used.  
     
     
         33 . The method as claimed in  claim 26 ,  27  or  28 , wherein, in step (b), between 10 and 500 mRNA species or cDNA species are used.  
     
     
         34 . The method as claimed in  claim 18 ,  26 ,  27  or  28 , wherein said solid support is a filter.  
     
     
         35 . The method as claimed in  claim 17 ,  18 ,  26 ,  27  or  28 , wherein said disease or condition is selected from the group consisting of: cancer, diseases or conditions caused by viruses, parasites, toxins, drugs or allergens, psychiatric or psychosomatic diseases and diseases of unknown cause.  
     
     
         36 . The method as claimed in  claim 17 ,  18 ,  26 ,  27  or  28 , wherein said disease or condition is selected from the group consisting of: multiple sclerosis, schizophrenia, Alzheimer's disease, stomach, lung, breast, prostate and bowel cancer.  
     
     
         37 . The method as claimed in  claim 28 , wherein said method is used to detect a human's reaction to drug treatment.