Isolation of soluble scFv antibody fragments specific for small biomarker molecule, L-Carnitine, using phage display.

Rabab M Abou El-Magd, Nicolas F Vozza, Jack A Tuszynski, David S Wishart

Journal: Journal of immunological methods 2016;428():9-19

PMID: 26608419

Full Text on Elsevier Science View on PubMed

Abstract

Isolation of single chain antibody fragment (scFv) clones from naïve Tomlinson I+J phage display libraries that specifically bind a small biomarker molecule, L-Carnitine, was performed using iterative affinity selection procedures. L-Carnitine has been described as a conditionally essential nutrient for humans. Abnormally high concentrations of L-Carnitine in urine are related to many health disorders including diabetes mellitus type 2 and lung cancer. ELISA-based affinity characterization results indicate that selectants preferentially bind to L-Carnitine in the presence of key bioselecting component materials and closely related L-Carnitine derivatives. In addition, the affinity results were confirmed using biophysical fluorescence quenching for tyrosine residues in the V segment. Small-scale production of the soluble fragment yielded 1.3mg/L using immunopure-immobilized protein A affinity column. Circular Dichroism data revealed that the antibody fragment (Ab) represents a folded protein that mainly consists of β-sheets. These novel antibody fragments may find utility as molecular affinity interface receptors in various electrochemical biosensor platforms to provide specific L-Carnitine binding capability with potential applications in metabolomic devices for companion diagnostics and personalized medicine applications. It may also be used in any other biomedical application where detection of the L-Carnitine level is important.

Address: Department of Biological Sciences, University of Alberta, CCIS 5-012, Edmonton, AB T6G 2E1, Canada; Genetic Engineering and Biotechnology Research Institute, City of Scientific Research and Technological Applications, 21934 New-Burgelarab City, Alexandria, Egypt. Electronic address: [email protected].; Department of Biological Sciences, University of Alberta, CCIS 5-012, Edmonton, AB T6G 2E1, Canada. Electronic address: [email protected].; Experimental Oncology, Department of Oncology, University of Alberta, Canada; Cross Cancer Institute, 11560 University Avenue, Edmonton, AB T6G 1Z2, Canada; Department of Physics, University of Alberta, Edmonton, AB, Canada. Electronic address: [email protected].; Department of Biological Sciences, University of Alberta, CCIS 5-012, Edmonton, AB T6G 2E1, Canada; Department of Computing Sciences, University of Alberta, Edmonton, AB T6G 2E8, Canada. Electronic address: [email protected].

DOI: 10.1016/j.jim.2015.11.006

Link outs

Free resources

Other Literature Sources:

https://scite.ai/reports/26608419

Subscription / membership required

Full Text Sources:

https://linkinghub.elsevier.com/retrieve/pii/S0022...

MeSH Terms: Antigen-Antibody Reactions, Biomarkers, Carnitine, Enzyme-Linked Immunosorbent Assay, Humans, Peptide Library, Single-Chain Antibodies, Solubility