Method of detection by enhancement of silver staining

Abstract

The present invention relates to a method for amplifying a detection signal by enhancing or promoting the deposition of additional silver in assay detection systems where the formation of a silver spot serves as a reporter for the presence of a target molecule, including biological polymers (e.g., proteins and nucleic acids) and small molecules.

What we claim:  
     
         1 . A method for amplifying a signal comprising: 
 (a) providing a substrate having deposited silver;    (b) contacting the substrate having deposited silver with a solution comprising nanoparticles having oligonucleotides bound thereto so as to produce a substrate having a nanoparticles-silver sandwich;    (c) washing the substrate having said sandwich; and    (d) contacting the substrate having said sandwich with silver ions and a reducing agent to promote silver deposition onto the nanoparticles of said sandwich and produce silver-nanoparticles-silver sandwich.    (e) washing the substrate having the silver-nanoparticles-silver sandwich.    
     
     
         2 . The method according to  claim 1 , wherein the nanoparticles comprise gold, silver, platinum or mixtures thereof.  
     
     
         3 . The method according to  claim 3 , wherein the nanoparticles having oligonucleotides bound thereto comprise gold nanoparticle-oligonucleotide conjugates or complexes.  
     
     
         4 . The method according to  claim 2 , wherein the silver ion is derived from a silver salt comprising silver acetate, silver lactate, or silver nitrate.  
     
     
         5 . The method according to  claim 1  wherein the reducing agent comprises hydroquinone, n-propyl galate, p-phenylenediamine, or formaldehyde.  
     
     
         6 . The method according to  claim 1  wherein step (b) contacting is performed for a period ranging from about 5 to 30 minutes.  
     
     
         7 . The method according to  claim 6  wherein step (b) contacting is performed for about 10 minutes.  
     
     
         8 . The method according to  claim 1  wherein step (d) contacting is performed for a period ranging from about 5 to 10 minutes.  
     
     
         9 . The method according to  claim 8  wherein step (d) contacting is performed for about 5 minutes.  
     
     
         10 . The method according to  claim 1  wherein the deposited silver bound to the substrate is arranged in the form of an array.  
     
     
         11 . The method according to  claim 1 , wherein step (c) washing is performed with water.  
     
     
         12 . The method according to  claim 1 , wherein the substrate is glass.  
     
     
         13 . A method for promoting silver deposition onto a surface comprising silver, said method comprising the steps of: 
 (a) providing a surface having silver bound thereto;    (b) contacting the surface with a solution comprising nanoparticles having oligonucleotides bound thereto so as to produce a surface having a nanoparticles-silver sandwich;    (c) washing the surface having said nanoparticles-silver sandwich;    (d) contacting the surface having said nanoparticles-silver sandwich with a solution including silver ions under reducing conditions to promote silver deposition onto said nanoparticles of said nanoparticles-silver sandwich; and    (e) washing the surface having deposited silver.    
     
     
         14 . The method according to  claim 13  wherein said surface comprises cells or tissue.  
     
     
         15 . The method according to  claim 13 , wherein the nanoparticles comprise gold, silver, platinum or combinations thereof.  
     
     
         16 . The method according to  claim 13 , wherein the nanoparticles having oligonucleotides bound thereto comprise gold nanoparticles having oligonucleotides bound thereto.  
     
     
         17 . The method according to  claim 16 , wherein the gold nanoparticles having oligonucleotides bound thereto comprise gold nanoparticle-oligonucleotide conjugates or complexes.  
     
     
         18 . The method according to  claim 13 , wherein the silver ion is derived from a silver salt comprising silver acetate, silver lactate, or silver nitrate.  
     
     
         19 . The method according to  claim 13  wherein the reducing agent comprising hydroquinone, n-propyl galate, p-phenylenediamine, or formaldehyde.  
     
     
         20 . The method according to  claim 13  wherein step (b) contacting is performed for a period ranging from about 5 to 30 minutes.  
     
     
         21 . The method according to  claim 20  wherein step (b) contacting is performed for a period of about 10 minutes.  
     
     
         22 . The method according to  claim 13  wherein step (d) contacting is performed for a period ranging from about 5 to 10 minutes.  
     
     
         23 . The method according to  claim 22  wherein step (d) contacting is performed for about 5 minutes.  
     
     
         24 . The method according to  claim 13  wherein the silver bound to the substrate are arranged in the form of an array.  
     
     
         25 . A kit for signal amplification comprising: 
 (b) container including nanoparticles having oligonucleotides bound thereto;    (b) container including a silver salt; and    (c) container including a reducing agent.    
     
     
         26 . The kit according to  claim 25 , wherein the nanoparticles comprise gold, silver, platinum or combinations thereof.  
     
     
         27 . The kit according to  claim 26 , wherein the nanoparticles comprising gold nanoparticles having oligonucleotides bound thereto.  
     
     
         28 . The kit according to  claim 27 , wherein the gold nanoparticles having oligonucleotides bound thereto comprise gold nanoparticle-oligonucleotide conjugates or complexes.  
     
     
         29 . The kit according to  claim 25 , wherein the silver ion is derived from a silver salt comprising silver acetate, silver lactate, or silver nitrate.  
     
     
         30 . The kit according to  claim 25 , wherein the reducing agent comprises hydroquinone, n-propyl galate, p-phenylenediamine, or formaldehyde.