Retentate chromatography and protein chip arrays with applications in biology and medicine

Abstract

This invention provides methods of retentate chromatography for resolving analytes in a sample. The methods involve adsorbing the analytes to a substrate under a plurality of different selectivity conditions, and detecting the analytes retained on the substrate by desorption spectrometry. The methods are useful in biology and medicine, including clinical diagnostics and drug discovery.

1 - 26 . (canceled)  
     
     
         27 . A method for detecting translation of a polynucleotide comprising the steps of: 
 a) providing a substrate for use in desorption spectrometry, wherein the substrate comprises a surface and an adsorbent attached to the surface;    b) providing an mRNA encoding a polypeptide and reagents for in vitro translation of the mRNA;    c) translating the mRNA in situ on the substrate, whereby the polypeptide is produced and is bound through the adsorbent to the substrate;    d) exposing the substrate to an eluant to wash off unbound material and to allow retention of the polypeptide by the adsorbent; and    e) detecting retained polypeptide by desorption spectrometry;    whereby detection of the polypeptide provides detection of translation of the mRNA.    
     
     
         28 - 35 . (canceled)  
     
     
         36 . The method of  claim 27  wherein the adsorbent specifically binds the polypeptide.  
     
     
         37 . The method of  claim 36  wherein the adsorbent comprises an antibody.  
     
     
         38 . The method of  claim 27  wherein the adsorbent is a hydrophilic interaction adsorbent, a hydrophobic interaction adsorbent, a metal chelate adsorbent, an anionic adsorbent or a cationic adsorbent.  
     
     
         39 . The method of  claim 27  wherein step (b) further comprises providing reagents for in vitro transcription of the mRNA.  
     
     
         40 . The method of  claim 27  wherein the polynucleotide is comprised in a genetic package.  
     
     
         41 . The method of  claim 27  wherein the genetic package is a bacteriophage.  
     
     
         42 . The method of  claim 27  wherein step (c) comprises creating a well over the substrate with the adsorbent at a bottom of the well and placing the reagents and the polynucleotide in the well.  
     
     
         43 . The method of any of claims  27  and  36 - 43  wherein the substrate is a mass spectrometry probe and desorption spectrometry comprises laser desorption mass spectrometry.