2'-fluoropyrimidine anti-calf intestinal phosphatase nucleic acid ligands

Abstract

Methods are described for the identification and preparation of nucleic acid ligands to calf intestinal phosphatase. Included in the invention are specific RNA ligands to calf intestinal phosphatase identified by the SELEX method.

What is claimed is:  
     
         1 . A reagent for the detection of a target molecule, the reagent comprising: 
 a) a nucleic acid ligand to said target; and    b) a nucleic acid ligand to calf intestinal phosphatase (CIP);    wherein the sequence of said nucleic acid ligand to said target is contiguous with the sequence of said nucleic acid ligand to CIP.    
     
     
         2 . The reagent of  claim 1  wherein said nucleic acid ligand to CIP is a non-inhibitory ligand.  
     
     
         3 . The reagent of  claim 1  wherein said nucleic acid ligand to CIP is an inhibitory ligand.  
     
     
         4 . The reagent of  claim 3  wherein the binding of said target to said nucleic acid ligand to said target releases any CIP that is bound to said nucleic acid ligand to CIP.  
     
     
         5 . A method for determining the quantity of a target molecule in a sample suspected of containing such target molecule, the method comprising: 
 a) providing a reagent for the detection of said target molecule, the reagent comprising: 
 i) a nucleic acid ligand to said target; and  
 ii) an inhibitory nucleic acid ligand to calf intestinal phosphatase (CIP); wherein the sequence of said nucleic acid ligand to said target is contiguous with the sequence of said nucleic acid ligand to CIP, and wherein the binding of said target to said nucleic acid ligand to said target releases any CIP that is bound to said inhibitory nucleic acid to CIP;  
   b) contacting said reagent with a predetermined quantity of CIP, wherein substantially all of the CIP is bound and inhibited by said inhibitory nucleic acid ligand to CIP;    c) contacting said reagent and said predetermined quantity of CIP with said sample to form a test mixture; and    d) measuring the CIP activity in said test mixture;    whereby binding of said target in said sample releases CIP that is bound to the inhibitory nucleic acid ligand to CIP of said reagent.    
     
     
         6 . The method of  claim 5  wherein step d) is accomplished using a detectable substrate of calf intestinal phosphatase.  
     
     
         7 . The method of  claim 6  wherein said detectable substrate is selecting from the group consisting of 1,2 dioxetane and p-nitrophenylphosphate.  
     
     
         8 . An antibody conjugated to a non-inhibitory nucleic acid ligand to calf intestinal phosphatase (CIP).  
     
     
         9 . A method of determining the quantity of a target molecule in a sample suspected of containing said target molecule, the method comprising: 
 a) immobilizing said sample on a solid support;    b) contacting said immobilized sample with an antibody to said target, wherein said antibody is conjugated to a non-inhibitory nucleic acid ligand to calf intestinal phosphatase (CIP);    c) contacting said immobilized sample with CIP; and    d) determining the level of CIP activity.    
     
     
         10 . The method of  claim 9  wherein step d) is accomplished using a detectable substrate of calf intestinal phosphatase.  
     
     
         11 . The method of  claim 10  wherein said detectable substrate is selecting from the group consisting of 1,2 dioxetane and p-nitrophenylphosphate.