Low Density Microarrays for Vaccine Related Protein Quantification, Potency Determination, and Efficacy Evaluation

Abstract

Methods for the quantification of influenza HA proteins and anti-influenza antibodies for the fields of vaccine-related protein quantification, potency determination, and efficacy evaluation are provided. According to the technology, quantification is achieved by providing capture agents attached to an array in a series of decreasing concentrations. Serial dilutions of a reference material also may be introduced. The reference material within each solution binds to the capture agents on the array and is labeled with a label agent capable of producing a detectable signal used to construct a calibration curve. A target material of unknown concentration is introduced to a separate identical array, and the target material binds to the capture agents and also is labeled by a label agent to produce a detectable signal. The calibration curve based on the reference material is then utilized to determine the concentration of the target material without the need to perform replicate experiments.

1 - 15 . (canceled) 
     
     
         16 . A system for quantification of an amount of influenza hemagglutinin (HA) target protein present in a sample, comprising:
 a solid support;   a plurality of capture agents affixed to said solid support, wherein:
 said plurality of capture agents bind influenza HA, 
 said plurality of capture agents are affixed to the solid support in a plurality of discrete areas in an array, said discrete areas each containing only a single unique capture agent; 
 said plurality of capture agents comprise:
 a first anti-influenza A H1 antibody capture agent; 
 a second anti-influenza A H3 antibody capture agent; and 
 a third anti-influenza B antibody capture agent; or 
 sialic acid glycopeptides or proteins; 
 
   at least one label agent to produce a detectable signal indicative of binding between said plurality of capture agents and the influenza HA target.   
     
     
         17 . (canceled) 
     
     
         18 . The system of  claim 16 , wherein the at least one label agent binds target influenza hemagglutinin A H1N1 and A H3 and influenza B. 
     
     
         19 . The system of  claim 16 , wherein at least one of said capture agents is a universal anti-influenza antibody and the at least one label agent binds target influenza hemagglutinin. 
     
     
         20 . (canceled) 
     
     
         21 . (canceled) 
     
     
         22 . The system of  claim 16 , wherein said plurality of discrete areas are arranged in a configuration of decreasing concentration of said capture agents. 
     
     
         23 . The system of  claim 16 , wherein the label agent is a fluorescent, colorimetric, electrical or magnetic label. 
     
     
         24 . The system of  claim 16 , further comprising a calibration curve to quantify influenza HA in the sample from the detected signal. 
     
     
         25 . The system of  claim 24 , wherein the calibration curve is based on a magnitude of detectable signal created by binding between a standard influenza antigen and the capture agents. 
     
     
         26 . The system of  claim 16 , wherein said anti-influenza A H1 antibody capture agent is a monoclonal antibody specific to influenza A H1. 
     
     
         27 . The system of  claim 26 , wherein the influenza A H1 is influenza A H1N1. 
     
     
         28 . The system of  claim 16 , wherein said anti-influenza A H3 antibody capture agent is a monoclonal antibody specific to influenza A H3. 
     
     
         29 . The system of  claim 28 , wherein the influenza A H3 is influenza A H3N2. 
     
     
         30 . The system of  claim 16 , configured for use with a sample comprised of material from an influenza vaccine production process. 
     
     
         31 . The system of  claim 16 , further comprising at least one positive control and at least one negative control affixed to the solid support. 
     
     
         32 . The system of  claim 16 , wherein the capture agents comprise α2,6 and α2,3 sialic acid glycopeptides or proteins. 
     
     
         33 . The system of  claim 16 , wherein the capture agents comprise:
 the first anti-influenza A H1 antibody capture agent;   the second anti-influenza A H3 antibody capture agent; and   the third anti-influenza B antibody capture agent.   
     
     
         34 . The system of  claim 16 , comprising a plurality of label agent species, with each species specific to a unique target protein.