Phage display antibody-based proteomic device using resonance-enhanced detection.
The combination of phage display antibody arrays with a novel nanotransducer technique based on resonant nanoparticles in a nanosandwiched film enables the sensitive parallel screening of proteins. Using the resonance of nanoparticles with their induced mirror dipoles in a thin-film structure, limitations of fluorophores, such as unspecific background and nonvisibility to the eye, can be overcome, thereby leading to an optical signal significantly more sensitive than that of standard colloid techniques. The signal can be both directly observed as a color change of a microdot at the sensor surface and tuned throughout the visible range of the spectrum. Here we report the application of an optical chip using scFv-antibody-antigen interactions. Artificial scFv-antibodies against a variety of proteins, including yeast enzymes and bovine serum albumin (as a standard), were constructed via Phage Display. These scFv-antibodies were then coated onto metal nanoclusters and bound to their antigens that were arrayed as nanodroplets at the resonance layer of the chip. ScFv-Antibody-antigen interaction resulted in a visible array of microdots. Using resonance-enhanced absorption, the absorption signal of the spots was amplified by one to two orders of magnitude (compared to colloid-based techniques). For quantitative analysis, either an 8-micron scanner or a CCD camera (resolution 4 microns) was employed to gain direct-reflection spectra rather than unspecific scatter data (prone to dust and unspecific interaction). Our results demonstrate that this device enables high-throughput proteomics to overcome some limitations of fluorescence, enzyme labels, and colloid techniques.
Stich
Norbert
N
Kluyver Laboratory for Biotechnology, TU-Delft, 2628BC Delft, The Netherlands.
Gandhum
Arminder
A
Matyushin
Viatcheslav
V
Raats
Jos
J
Mayer
Christian
C
Alguel
Yilmaz
Y
Schalkhammer
Thomas
T
D000906:Antibodies; D000936:Antigen-Antibody Complex / Q000032:analysis; D020099:Coated Materials, Biocompatible / Q000138:chemical synthesis; D004926:Escherichia coli / Q000276:immunology / Q000378:metabolism; D005240:Feasibility Studies; D006046:Gold; D008863:Microspheres; D036103:Nanotechnology / Q000295:instrumentation* / Q000379:methods; D019151:Peptide Library; D040081:Protein Array Analysis / Q000295:instrumentation* / Q000379:methods; D011506:Proteins / Q000032:analysis* / Q000737:chemistry; D040901:Proteomics / Q000295:instrumentation / Q000379:methods; D020349:Surface Plasmon Resonance / Q000295:instrumentation* / Q000379:methods
D000906:Antibodies; D000936:Antigen-Antibody Complex; D020099:Coated Materials, Biocompatible; D019151:Peptide Library; D011506:Proteins; D006046:Gold