title: A novel excision selection method for isolation of antibodies binding antigens expressed specifically by rare cells in tissue sections.
abstract: There is a growing appreciation of single cell technologies to provide increased biological insight and allow development of improved therapeutics. The central dogma explains why single cell technologies is further advanced in studies targeting nucleic acids compared to proteins, as nucleic acid amplification makes experimental detection possible. Here we describe a novel method for single round phage display selection of antibody fragments from genetic libraries targeting antigens expressed by rare cells in tissue sections. We present and discuss the results of two selections of antibodies recognizing antigens expressed by perivascular cells surrounding capillaries located in a human brain section; with the aim of identifying biomarkers expressed by pericytes. The area targeted for selection was identified by a known biomarker and morphological appearance, however in situ hybridizations to nucleic acids can also be used for the identification of target cells. The antibody selections were performed directly on the tissue sections followed by excision of the target cells using a glass capillary attached to micromanipulation equipment. Antibodies bound to the target cells were characterized using ELISA, immunocytochemistry and immunohistochemistry. The described method will provide a valuable tool for the discovery of novel biomarkers on rare cells in all types of tissues.
authors.affiliation: Department of Clinical Medicine, Aarhus University, Nørrebrogade 44, 8000 Aarhus C, Denmark.|Sino-Danish Centre for Education and Research (SDC), Niels Jensens Vej 2, 8000 Aarhus C, Denmark.
authors.forename: Simon
authors.initials: S
authors.lastname: Lykkemark
authors.affiliation: Department of Engineering, Aarhus University, Gustav Wieds Vej 10, 8000 Aarhus C, Denmark.
authors.forename: Ole Aalund
authors.initials: OA
authors.lastname: Mandrup
authors.affiliation: Department of Engineering, Aarhus University, Gustav Wieds Vej 10, 8000 Aarhus C, Denmark.
authors.forename: Mads Bjørnkjær
authors.initials: MB
authors.lastname: Jensen
authors.affiliation: Department of Molecular Biology and Genetics, Gustav Wieds Vej 10, 8000 Aarhus C, Denmark.
authors.forename: Jesper
authors.initials: J
authors.lastname: Just
authors.affiliation: Department of Engineering, Aarhus University, Gustav Wieds Vej 10, 8000 Aarhus C, Denmark.
authors.forename: Peter
authors.initials: P
authors.lastname: Kristensen
mesh_terms: D000918:Antibody Specificity; D000941:Antigens / Q000096:biosynthesis* / Q000235:genetics; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000737:chemistry / Q000166:cytology; D015870:Gene Expression; D006801:Humans; D007128:Immunoglobulin Fragments / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*; D019151:Peptide Library; D059010:Single-Cell Analysis
chemical_list: D000941:Antigens; D007128:Immunoglobulin Fragments; D019151:Peptide Library