Tagged antibodies are one of most widely used tools in biology and medicine. Methods such as analyte immunoassays, flow cytometry, and protein immunolocalization are used on an enormous scale in both pure research and clinical, diagnostic applications. The development of fluorescently labeled antibodies has extended the range and power of these techniques by permitting simultaneous detection of several entities while at the same time increasing overall sensitivity. However, despite intense technical maximization efforts, current fluorescent probes do not provide the multiplexing capacity and sensitivity desired for many applications. These limitations result from i) the use of the visible spectrum for fluorescence detection and ii) the inherent properties of the fluorophores used. The studies proposed are aimed at overcoming these limitations by developing antibodies tagged with novel fluorophores that emit in a broad, unused region of the spectrum - the shortwave infrared (SWIR) (850- 1200 nm). Fluorescence emission can be detected with greater sensitivity in this region because, in contrast to the visible region, biological materials show negligible autofluorescence at these wavelengths. More importantly, doubling of the spectral region that can be used for detection will greatly enhance capacity for multiplexed analysis. The fluorophores to be used are individual species of single walled carbon nanotubes (SWCNTs). Additional unique properties of the fluorescent emission from these novel carbon structures will contribute further to their usefulness in these applications. Individual SWCNT species show sharp distinct emission spectra, providing greater multiplexing possibilities than the commonly used visible-range fluors. Unlike the commonly used fluors, SWCNT fluorescence shows no photobleaching or intermittent emission (""""""""blinking""""""""). The sensitivity for SWCNT fluorescence detection is also remarkable: individual SWCNTs can be detected in biological material. The successful generation of these SWCNT-based immunoprobes will enhance widely used clinical applications and basic research. In particular, the certainty of detection of metastatic cells by simultaneous probing for multiple cell markers will be significantly improved.
Fluorescently labeled antibodies are widely used in clinical applications for detection of hormones, blood proteins, disease organisms, and cancerous cells. However, the current versions of these antibodies have limitations in terms of sensitivity of detection and the number that can be used simultaneously on a single clinical sample. The experiments proposed will create novel fluorescent antibodies that will overcome these limitations and thus improve the quality and efficiency of medical diagnoses.
| Lin, Ching-Wei; Bachilo, Sergei M; Vu, Michael et al. (2016) Spectral triangulation: a 3D method for locating single-walled carbon nanotubes in vivo. Nanoscale 8:10348-57 |
