Switchable reporter enzymes based on mutually exclusive domain interactions allow antibody detection directly in solution.
Detection of antibodies is essential for the diagnosis of many diseases including infections, allergies, and autoimmune diseases. Current heterogeneous immunoassays require multiple time-consuming binding and washing steps, which limits their application in point-of-care diagnostics and high-throughput screening. Here, we report switchable reporter enzymes that allow simple colorimetric detection of antibodies directly in solution. Our approach is based on the antibody-induced disruption of an intramolecular interaction between TEM1 β-lactamase and its inhibitor protein BLIP. Using the HIV1-p17 antibody as an initial target, the interaction between enzyme and inhibitor was carefully tuned to yield a reporter enzyme whose activity increased 10-fold in the presence of pM antibody concentrations. Reporter enzymes for two other antibodies (HA-tag and Dengue virus type I) were obtained by simply replacing the epitope sequences. This new sensor design represents a modular and generic approach to construct antibody reporter enzymes without the cumbersome optimization required by previous engineering strategies.
Laboratory of Chemical Biology, Department of Biomedical Engineering, Eindhoven University of Technology , P.O. Box 513, 5600MB Eindhoven, The Netherlands.
Sambashiva
S
Banala
Stijn J A
SJ
Aper
Werner
W
Schalk
Maarten
M
Merkx
D000906:Antibodies / Q000032:analysis*; D003124:Colorimetry; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D057230:Limit of Detection; D017434:Protein Structure, Tertiary; D012996:Solutions; D065093:beta-Lactamase Inhibitors / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D001618:beta-Lactamases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D000906:Antibodies; D012996:Solutions; D065093:beta-Lactamase Inhibitors; D001618:beta-Lactamases