title: Detection of pathogen-specific antibodies by loop-mediated isothermal amplification.
abstract: Loop-mediated isothermal amplification (LAMP) is a method for enzymatically replicating DNA that has great utility for clinical diagnosis at the point of care (POC), given its high sensitivity, specificity, speed, and technical requirements (isothermal conditions). Here, we adapted LAMP for measuring protein analytes by creating a protein-DNA fusion (referred to here as a "LAMPole") that attaches oligonucleotides (LAMP templates) to IgG antibodies. This fusion consists of a DNA element covalently bonded to an IgG-binding polypeptide (protein L/G domain). In our platform, LAMP is expected to provide the most suitable means for amplifying LAMPoles for clinical diagnosis at the POC, while quantitative PCR is more suitable for laboratory-based quantification of antigen-specific IgG abundance. As proof of concept, we measured serological responses to a protozoan parasite by quantifying changes in solution turbidity in real time. We observed a >6-log fold difference in signal between sera from vaccinated versus control mice and in a clinical patient sample versus a control. We assert that LAMPoles will be useful for increasing the sensitivity of measuring proteins, whether it be in a clinical laboratory or in a field setting, thereby improving acute diagnosis of a variety of infections.
authors: VTT/MSI Molecular Sciences Institute, Berkeley, California, USA.
authors: Ian E
authors: IE
authors: Burbulis
authors: VTT/MSI Molecular Sciences Institute, Berkeley, California, USA.
authors: Kumiko
authors: K
authors: Yamaguchi
authors: Department of Pathology, The Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
authors: Olga V
authors: OV
authors: Nikolskaia
authors: Department of Molecular Microbiology and Immunology, The Johns Hopkins School of Public Health, Baltimore, Maryland, USA.
authors: Sean T
authors: ST
authors: Prigge
authors: Unit of Cellular and Molecular Immunology, Vrije Universiteit Brussel, Brussels, Belgium Department of Structural Biology, Flanders Institute for Biotechnology, Brussels, Belgium.
authors: Stefan
authors: S
authors: Magez
authors: INSERM U1094, Neuroépidémiologie Tropicale, Universite de Limoges, Limoges, France.
authors: Sylvie
authors: S
authors: Bisser
authors: Department of Pathology, The Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
authors: Megan E
authors: ME
authors: Reller
authors: Department of Pathology, The Johns Hopkins University School of Medicine, Baltimore, Maryland, USA dgrab1@jhmi.edu.
authors: Dennis J
authors: DJ
authors: Grab
mesh_terms: D000818:Animals; D000913:Antibodies, Protozoan / Q000097:blood*; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood*; D051379:Mice; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*
chemical_list: D000913:Antibodies, Protozoan; D007074:Immunoglobulin G
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