Methods for reducing the range in concentrations of analyte species in a sample

Abstract

The present invention relates to the fields of molecular biology, combinatorial chemistry and biochemistry. Particularly, the present invention describes methods and kits for dynamically reducing the variance between analyte taken from complex mixtures.

1 . A method comprising the steps of: 
 (a) providing a first sample comprising a plurality of different analyte species present in the first sample in a first range of concentrations;    (b) contacting the first sample with an amount of a library comprising at least 100 different binding moieties;    (c) capturing amounts of the different analyte species from the first sample with the different binding moieties and removing unbound analyte species; and    (d) isolating the captured analyte species from the binding moieties to produce a second sample comprising a plurality of different analyte species present in the second sample in a second range of concentrations;    wherein the amount of the library is selected to capture amounts of the different analyte species so that the second range of concentrations is less than the first range of concentrations.    
     
     
         2 . The method of  claim 1 , wherein the first sample comprises at least 100, at least 1,000, at least 10,000, at least 100,000, at least 1,000,000 or at least 10,000,000 different analyte species.  
     
     
         3 . The method of  claim 1 , wherein the library comprises at least 1,000, at least 10,000, at least 100,000, at least 1,000,000 or at least 10,000,000 different binding moieties.  
     
     
         4 . The method of  claim 1 , wherein the binding moieties comprise bio-organic polymers.  
     
     
         5 . The method of  claim 1 , wherein the binding moieties are bound to a solid support or supports.  
     
     
         6 . The method of  claim 1 , wherein the library is a non-selective library.  
     
     
         7 . The method of  claim 1 , wherein the different binding moieties are comprised in a complete or incomplete combinatorial library.  
     
     
         8 . The method of  claim 1 , wherein the second sample has a diversity of analyte species that is substantially the same as the first sample.  
     
     
         9 . The method of  claim 1 , wherein the sample is selected from the group consisting of amniotic fluid, blood, cerebrospinal fluid, intraarticular fluid, intraocular fluid, lymphatic fluid, milk, perspiration plasma, saliva semen, seminal plasma, serum, sputum, synovial fluid, tears, umbilical cord fluid, urine, biopsy homogenate, cell culture fluid, cell extracts, cell homogenate, conditioned medium, fermentation broth, tissue homogenate and derivatives of these.  
     
     
         10 . The method of  claim 1 , further comprising detecting analyte species in the second sample.  
     
     
         11 . The method of  claim 1 , wherein removing unbound analytes comprises washing the captured analytes with a wash buffer.  
     
     
         12 . The method of  claim 1 , wherein the analytes are selected from the group consisting of polypeptides, nucleic acids, complex carbohydrates, complex lipids, synthetic inorganic compounds and synthetic organic compounds.  
     
     
         13 . The method of  claim 1 , further comprising fractionating the analytes in the second sample based on a physical or chemical property.  
     
     
         14 . The method of  claim 1 , further comprising identifying at least one of the isolated analytes.  
     
     
         15 . The method of  claim 1 , further comprising contacting a biospecific binding moiety with the second sample and determining whether the biospecific binding moiety has captured an analyte species from the second sample.  
     
     
         16 . The method of  claim 4 , wherein the bio-organic polymers are selected from the group consisting of peptides, oligonucleotides and oligosaccharides.  
     
     
         17 . The method of  claim 1 , wherein the binding moieties are selected from antibodies and aptamers.  
     
     
         18 . The method of  claim 5 , wherein the solid support or supports is a collection of beads or particles.  
     
     
         19 . The method of  claim 5 , wherein the solid support or supports is selected from the group consisting of fibers, monoliths, membranes and plastic strips.  
     
     
         20 . The method of  claim 6 , wherein the non-selective library is selected from the group consisting of a germ line antibody library, a phage display library of recombinant binding proteins, a dye library or a non-combinatorial library in which the binding specificity of the members is not pre-selected, a combinatorial library and portions thereof.  
     
     
         21 . The method of  claim 7 , wherein the combinatorial library is a hexapeptide library.  
     
     
         22 . The method of  claim 10 , wherein the analytes are detected using a method selected from the group consisting of calorimetric, spectrophotometric, magnetic resonance, ellipsometric, mass spectroscopic, electrophoretic, chromatographic, enzymatic, and sequence analysis.  
     
     
         23 . The method of  claim 13 , wherein fractionating comprises segregating the analytes using a technique selected from the group consisting of chromatography, electrophoresis, capillary electrophoresis, filtration and precipitation.  
     
     
         24 . The method of  claim 18 , wherein each bead or particle is attached to a substantially different binding moiety.  
     
     
         25 . The method of  claim 18 , wherein a plurality of different binding moieties are attached to the same bead or particle.  
     
     
         26 . The method of  claim 18 , wherein the beads or particles have a diameter of less than 1 μm.  
     
     
         27 . The method of  claim 18 , wherein the beads or particles are formed milling microparticulate beads using a method selected from the group consisting of crushing, grinding, and sonicating.  
     
     
         28 . The method of  claim 27 , wherein the microparticulate beads are a polymeric matrix formed from a natural or synthetic polymer.  
     
     
         29 . The method of  claim 27 , wherein the particles are coupled to a second solid support to form an array or dipstick.  
     
     
         30 . A kit for detecting a plurality of analytes in a sample, comprising: 
 (i) a container comprising a library of at least 100 different binding moieties; and    (ii) instructions for using the library to perform the method of  claim 1 .    
     
     
         31 . The kit of  claim 30 , wherein the binding moieties are coupled to the solid support or supports.  
     
     
         32 . The kit of  claim 30 , wherein the library comprises a hexapeptide combinatorial library or a portion thereof wherein the hexapeptides are attached to particles.  
     
     
         33 . The kit of  claim 30 , further comprising a binding buffer for capturing analytes with the binding moieties.  
     
     
         34 . The kit of  claim 30 , further comprising an elution buffer for eluting captured analytes from the binding moieties.  
     
     
         35 . A library comprising at least 100 different binding moieties, wherein a plurality of different binding moieties are attached to the same solid support or supports.  
     
     
         36 . The library of  claim 35 , wherein the binding moieties comprise a combinatorial hexapeptide library or portion thereof.  
     
     
         37 . A method for identifying a diagnostic biomarker, the method comprising the steps of: 
 (a) providing a first set of biosamples from a first set of organisms having a first phenotype;    (b) providing a second set of biosamples from a second set of organisms having a second phenotype;    (c) performing the method of  claim 1  on each of the biosamples, thereby creating a third and fourth set of biosamples, respectively;    (d) detecting analyte species in each of the third and fourth set of biosamples;    (e) identifying at least one analyte species that is differentially present in the third and fourth set of biosamples, whereby the at least one analyte species is a biomarker for distinguishing the first phenotype from the second phenotype.    
     
     
         38 . The method of  claim 37 , wherein step (e) comprises identifying a biomarker profile that provides better predictive power than any one of the biomarkers in the profile, alone.  
     
     
         39 . A method for reducing the relative amounts of analytes in a sample, the method comprising the steps of: 
 (a) providing a first sample comprising a first plurality of different analytes having a first variance in amounts;    (b) contacting the first sample with a plurality of different binding moieties, each binding moiety present in a determined amount;    (c) capturing a portion of the first different analytes from the first sample with the different binding moieties and removing uncaptured analytes; and    (d) isolating the captured analytes from the binding moieties to produce a second sample comprising a second plurality of different analytes having a second variance in amounts;    wherein the determined amount of each of the plurality of different binding moieties is selected to capture amounts of the different analytes whereby the second variance in amounts is less than the first variance in amounts.