title: Real-time femtomolar detection of cancer biomarkers from photoconjugated antibody-phage constructs.
abstract: Here we describe novel covalent conjugates of antibody-phage for the detection of multiple cancer biomarkers using real time immuno-polymerase chain reaction (immuno-PCR). While the conventional process of immuno-PCR utilizes DNA-conjugated antibodies, chemical modification of antibodies not only reduces antibody affinity but also creates a heterogeneous population of products. However, phage naturally encapsulate genomic DNA, which can be used as a PCR template. To produce covalently conjugated antibody-phage constructs without recombinant antibody expression or chemical modification of antibodies, we incorporated a photocrosslinkable non-canonical amino acid within an antibody-binding domain displayed on one of the phage coat proteins. To correlate antigen presence to a specific DNA sequence, the phage genomes were modified with domains that recognized specific sets of primers. The crosslinked antibody-phage conjugates were then tested in a sandwich-type immunoassay using real-time PCR where low pg ml-1concentrations of antigen could be detected and identified from a single solution containing a mixture of three different types of cancer biomarkers.
authors.affiliation: Materials Science and Engineering Program, University of Colorado, Boulder, USA. jennifer.cha@colorado.edu.
authors.forename: M
authors.initials: M
authors.lastname: Brasino
authors.affiliation: Materials Science and Engineering Program, University of Colorado, Boulder, USA. jennifer.cha@colorado.edu and Department of Chemical and Biological Engineering, University of Colorado, Boulder, USA.
authors.forename: J N
authors.initials: JN
authors.lastname: Cha
mesh_terms: D000906:Antibodies / Q000378:metabolism*; D017104:Bacteriophage M13 / Q000235:genetics / Q000378:metabolism*; D014408:Biomarkers, Tumor / Q000032:analysis*; D007118:Immunoassay / Q000379:methods*; D057230:Limit of Detection; D055668:Photochemical Processes; D016133:Polymerase Chain Reaction; D013997:Time Factors
chemical_list: D000906:Antibodies; D014408:Biomarkers, Tumor