Highly sensitive immunoassay of protein molecules based on single nanoparticle fluorescence detection in a nanowell

Jin Hee Han, Hee Joo Kim, Sudheendra Lakshmana, Shirley J. Gee, Bruce D. Hammock, Ian M. Kennedy

Research output: Chapter in Book/Report/Conference proceedingConference contribution

Abstract

A nanoarray based-single molecule detection system was developed for detecting proteins with extremely high sensitivity. The nanoarray was able to effectively trap nanoparticles conjugated with biological sample into nanowells by integrating with an electrophoretic particle entrapment system (EPES). The nanoarray/EPES is superior to other biosensor using immunoassays in terms of saving the amounts of biological solution and enhancing kinetics of antibody binding due to reduced steric hindrance from the neighboring biological molecules. The nanoarray patterned onto a layer of PMMA and LOL on conductive and transparent indium tin oxide (ITO)-glass slide by using e-beam lithography. The suspension of 500 nm-fluorescent (green emission)-carboxylated polystyrene (PS) particles coated with protein-A followed by BDE 47 polyclonal antibody was added to the chip that was connected to the positive voltage. The droplet was covered by another ITO-coated-glass slide and connected to a ground terminal. After trapping the particles into the nanowells, the solution of different concentrations of anti-rabbit- IgG labeled with Alexa 532 was added for an immunoassay. A single molecule detection system could quantify the anti-rabbit IgG down to atto-mole level by counting photons emitted from the fluorescent dye bound to a single nanoparticle in a nanowell.

Original languageEnglish (US)
Title of host publicationProgress in Biomedical Optics and Imaging - Proceedings of SPIE
Volume7908
DOIs
StatePublished - 2011
EventNanoscale Imaging, Sensing, and Actuation for Biomedical Applications VIII - San Francisco, CA, United States
Duration: Jan 24 2011Jan 27 2011

Other

OtherNanoscale Imaging, Sensing, and Actuation for Biomedical Applications VIII
CountryUnited States
CitySan Francisco, CA
Period1/24/111/27/11

Keywords

  • ebeam lithography
  • electrophoretic particle entrapment system (EPES)
  • nanoarray
  • photons
  • single molecule detection system

ASJC Scopus subject areas

  • Atomic and Molecular Physics, and Optics
  • Electronic, Optical and Magnetic Materials
  • Biomaterials
  • Radiology Nuclear Medicine and imaging

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  • Cite this

    Han, J. H., Kim, H. J., Lakshmana, S., Gee, S. J., Hammock, B. D., & Kennedy, I. M. (2011). Highly sensitive immunoassay of protein molecules based on single nanoparticle fluorescence detection in a nanowell. In Progress in Biomedical Optics and Imaging - Proceedings of SPIE (Vol. 7908). [79080S] https://doi.org/10.1117/12.874259