title: Immunomagnetic separation of human myeloperoxidase using an antibody-mimicking peptide identified by phage display.
abstract: Phage display biopanning is a powerful in vitro selection process for screening and identifying peptides that bind to a target protein of interest. With the aim of replacing antibodies in immuno-diagnostic applications, we identified peptides whose binding characteristics mimicked those of anti-human myeloperoxidase (hMPO), a biomarker for acute cardiac diseases. Based on ELISA results from four phage clones, we selected and chemically synthesized a 12-mer peptide (SYIEPPERHRHR). Quartz crystal microbalance and surface plasmon resonance analyses revealed that the molar binding equilibrium ratio of the synthesized peptide was 0.023, approximately 43-fold lower than that of the anti-hMPO antibody. The dissociation constant (Kd) was 57nM, which was comparable to that of the native antibody (83nM). Next, we biotinylated the peptide at its N-terminus and attached the biotinylated peptide to the surface of streptavidin-coated magnetic particles to assess its ability to selectively capture hMPO. The binding equilibrium data were similar to the previous analyses; specifically, around 0.021mol peptide bound to 1mol of hMPO. Antigen capture was found to be selective and to be relatively little influenced by the presence of human serum albumin (HSA), an abundant constituent of serum. Our work demonstrates the potential of immunomagnetic isolation to achieve selective capture of a low-concentration antigen from complex solutions such as serum.
authors.affiliation: Department of Bionanotechnology, Graduate School, Hanyang University-ERICA, Ansan 15588, Republic of Korea.
authors.forename: Soi
authors.initials: S
authors.lastname: Yun
authors.affiliation: Department of Bionanotechnology, Graduate School, Hanyang University-ERICA, Ansan 15588, Republic of Korea.
authors.forename: Hyunmin
authors.initials: H
authors.lastname: Ryu
authors.affiliation: Department of Bionanotechnology, Graduate School, Hanyang University-ERICA, Ansan 15588, Republic of Korea. Electronic address: eklee@hanyang.ac.kr.
authors.forename: E K
authors.initials: EK
authors.lastname: Lee
mesh_terms: D000595:Amino Acid Sequence; D000906:Antibodies / Q000737:chemistry*; D000915:Antibody Affinity; D000941:Antigens / Q000276:immunology / Q000302:isolation & purification; D001435:Bacteriophages; D006801:Humans; D007159:Immunologic Tests; D018189:Immunomagnetic Separation / Q000379:methods*; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification; D057786:Peptidomimetics / Q000737:chemistry*; D009195:Peroxidase / Q000737:chemistry / Q000276:immunology*; D011485:Protein Binding; D058525:Quartz Crystal Microbalance Techniques; D000075462:Serum Albumin, Human / Q000737:chemistry; D020349:Surface Plasmon Resonance
chemical_list: D000906:Antibodies; D000941:Antigens; D019151:Peptide Library; D010455:Peptides; D057786:Peptidomimetics; D009195:Peroxidase; D000075462:Serum Albumin, Human
keywords: Antibody-mimicking peptide; Binding affinity; Human myeloperoxidase; Immuno-binding; Magnetic particle; Phage display