Rapid isolation of monoclonal antibodies from animals

Abstract

Methods and compositions for identification of candicate antigen-specific variable regions as well as generation of antibodies or antigen-binding fragments that could have desired antigen specificity are provided. For example, in certain aspects methods for determining amino acid sequences of serum antibody CDR and abundancy level are described. In some aspects, methods for determining nucleic acid sequences of antibody variable region sequences and frequency are provided. Furthermore, the invention provides methods for identification and generation of antibody or antigen-binding fragments that comprise highly-represented CDR.

What is claimed is: 
     
         1 . A method for preparing a report of the sequence information for plurality of antigen-specific antibodies comprising:
 (i) inputting protein mass spectra data obtained from a sample of a subject, having been exposed an antigen, into a computer;   (ii) inputting nucleic acid sequence data, obtained from the sample of the subject, into the computer;   (iii) running an alignment program to correlate the protein mass spectra data and the nucleic sequence data, thereby obtaining the polypeptide sequences for a plurality of antigen-specific antibodies expressed in the serum of the subject; and   (iv) preparing a report of the obtained the polypeptide sequences for a plurality of antigen-specific antibodies.   
     
     
         2 . The method of  claim 1 , wherein the alignment program performs a clustering analysis. 
     
     
         3 . The method of  claim 1 , wherein the alignment program compares multiple sequences by log expectation. 
     
     
         4 . The method of  claim 1 , wherein the alignment program further provides the abundancy level of a plurality of antibodies. 
     
     
         5 . The method of  claim 1 , wherein the sample is a serum sample. 
     
     
         6 . The method of  claim 1 , wherein the sample is from a lymphoid organ. 
     
     
         7 . The method of  claim 1 , wherein the wherein the protein mass spectra data is obtained by high performance liquid chromatography (HPLC)-mass spectroscopy. 
     
     
         8 . The method of  claim 1 , wherein the wherein the protein mass spectra data is obtained from a sample that has been enriched for antibodies that bind to a predetermined antigen. 
     
     
         9 . The method of  claim 1 , wherein the wherein the protein mass spectra data is obtained from a sample that has been enriched for CDR3-containing fragments of antibodies. 
     
     
         10 . The method of  claim 1 , wherein the nucleic acid sequence data is obtained by high throughput DNA sequencing. 
     
     
         11 . The method of  claim 10 , wherein the high-throughput sequencing comprises sequencing-by-synthesis, sequencing-by-ligation, sequencing-by-hybridization, single molecule DNA sequencing, multiplex polony sequencing, nanopore sequencing, or a combination thereof. 
     
     
         12 . The method of  claim 1 , wherein the subject has a tumor, an infectious disease, an autoimmune disease or has been immunized. 
     
     
         13 . The method of  claim 1 , further comprising producing one or more of the plurality of antibodies. 
     
     
         14 . The method of  claim 1 , further comprising evaluating the binding affinity of one or more of the plurality of antibodies. 
     
     
         15 . The method of  claim 1 , wherein the plurality antigen-specific antibodies bind to an infectious agent, a tumor antigen, a tumor cell or a self-antigen. 
     
     
         16 . The method of  claim 1 , wherein the subject is a human subject. 
     
     
         17 . The method of  claim 1 , wherein the plurality antigen-specific antibodies comprise IgG, IgM, or IgA antibodies. 
     
     
         18 . The method of  claim 1 , wherein the plurality antigen-specific antibodies are IgG antibodies.