Spraying Quantum Dot Conjugates in the Colon of Live Animals Enabled Rapid and Multiplex Cancer Diagnosis Using Endoscopy

Abstract

The detection of colon cancer using endoscopy is widely used, but the interpretation of the diagnosis is based on the clinician's naked eye. This is subjective and can lead to false detection. Here we developed a rapid and accurate molecular fluorescence imaging technique using antibody-coated quantum dots (Ab-QDs) sprayed and washed simultaneously on colon tumor tissues inside live animals, subsequently excited and imaged by endoscopy. QDs were conjugated to matrix metalloproteinases (MMP) 9, MMP 14, or carcinoembryonic antigen (CEA) Abs with zwitterionic surface coating to reduce nonspecific bindings. The Ab-QD probes can diagnose tumors on sectioned mouse tissues, fresh mouse colons stained ex vivo and also in vivo as well as fresh human colon adenoma tissues in 30 min and can be imaged with a depth of 100 μm. The probes successfully detected not only cancers that are readily discernible by bare eyes but also hyperplasia and adenoma regions. Sum and cross signal operations provided postprocessed images that can show complementary information or regions of high priority. This multiplexed quantum dot, spray-and-wash, and endoscopy approach provides a significant advantage for detecting small or flat tumors that may be missed by conventional endoscopic examinations and bestows a strategy for the improvement of cancer diagnosis.

Considerations in producing preferentially reduced half-antibody fragments

“…Employing  antibodies for surface modification has become a staple technique in the development of immunosensors (Virzonis et al 2014, Chrouda et al 2015, drug-delivery systems (Wu et al, Shahbazi et al 2014) and high-sensitivity biological labels (Jaiswal et al 2003, Park et al 2014. The simplest method of coupling antibodies to surfaces is physical adsorption.…”

Clinically compatible flexible wide-field multi-color fluorescence endoscopy with a porcine colon model

“…However, the emission spectrum of hfQDs is still somewhat broader than that of cadmium-based QD nanoparticles. While we used the incubation time of 30 min for tumor detection based on our previous study [18] that is relatively long duration for human clinical studies, this incubation time could be reduced with further optimization.…”

“…To investigate the applicability of biocompatible MMP14 Ab-hfQD probes in the spraying and washing method [18], fresh tumor tissue was harvested from a subcutaneous mouse CT26 (colon carcinoma cell line) xenograft model, and embedded in an ex vivo porcine colon tissue. The fresh tumor tissues were sprayed by 300 nM MMP14 Ab-hfQD probes in PBS solution with only PBS solution used as control.…”

“…In addition to their resistance to photobleaching, quantum dots (QDs) are ideal for multispectral imaging where different types of cells or tissues can be imaged simultaneously by labeling with distinct QDs at different wavelengths. Recently, specific molecular targeting with mouse colonic tumor and human colon adenoma was performed with QD-conjugates that contain heavy metal [18]. However, the inherent potential toxicity prevents cadmium-or lead-based QDs from being used for clinical imaging.…”

Biomarkers in Colorectal Cancer Screening

“…For example, light of certain wavelengths, in addition to being absorbed or reflected, can produce either autofluorescence 59 or stimulate a fluorescent response from exogenously introduced fluorophores. 60 Variation in fluorescence patterns between normal and abnormal intestinal mucosa, including neoplastic change, are well described. These types of molecular imaging represent areas of intense research, but to date have not yielded clinically important advantages.…”