A novel strategy for proteome-wide ligand screening using cross-linked phage matrices.
To find a suitable ligand from a complex antigen system is still a mission to be accomplished. Here we have explored a novel "library against proteome" panning strategy for ligand screening and antigen purification from a complex system using phage-displayed antibody technology. Human plasma proteome was targeted for phage library panning. During the process, the panning was carried out in solution, using a biotin/streptavidin beads separation system, for three rounds. Nine monoclonal phages, bound tightly to a number of unknown plasma proteins, were selected from the last round, six of which were directly employed as cross-linked matrices to purify their corresponding antigens from the plasma. The proteins isolated by G5 and E1 matrices were identified as amyloid protein and apolipoprotein A-I precursor, respectively. The results demonstrated that it was feasible to simultaneously obtain a number of ligand phages for various antigens, including low abundant proteins in a non-comparative proteome-wide system.
Qian
Chen
C
Institute of Molecular Medicine and State Key Laboratory of Pharmaceutical Biotechnology, Nanjing University Nanjing 210093.
Liu
Jian-Ning
JN
Institute of Molecular Medicine and State Key Laboratory of Pharmaceutical Biotechnology, Nanjing University Nanjing 210093; Institute of Molecular & Experimental Therapeutics, East China Normal University, Shanghai 200062.
Tang
Fengyuan
F
Institute of Molecular Medicine and State Key Laboratory of Pharmaceutical Biotechnology, Nanjing University Nanjing 210093.
Yuan
Dawen
D
Institute of Molecular Medicine and State Key Laboratory of Pharmaceutical Biotechnology, Nanjing University Nanjing 210093.
Guo
Zhigang
Z
Institute of Molecular & Experimental Therapeutics, East China Normal University, Shanghai 200062.
Zhang
Jing
J
Institute of Molecular Medicine and State Key Laboratory of Pharmaceutical Biotechnology, Nanjing University Nanjing 210093; Institute of Molecular & Experimental Therapeutics, East China Normal University, Shanghai 200062. Electronic address: jingzhang2001@gmail.com.
D000941:Antigens / Q000737:chemistry; D001435:Bacteriophages / Q000378:metabolism*; D001710:Biotin / Q000737:chemistry; D019921:Biotinylation; D001798:Blood Proteins / Q000737:chemistry; D003432:Cross-Linking Reagents / Q000737:chemistry; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007074:Immunoglobulin G / Q000737:chemistry; D008024:Ligands; D019151:Peptide Library; D020543:Proteome; D040901:Proteomics / Q000379:methods*; D012709:Serum Albumin / Q000737:chemistry; D019809:Streptavidin / Q000737:chemistry
D000941:Antigens; D001798:Blood Proteins; D003432:Cross-Linking Reagents; D007074:Immunoglobulin G; D008024:Ligands; D019151:Peptide Library; D020543:Proteome; D012709:Serum Albumin; D001710:Biotin; D019809:Streptavidin