Breaking Through Bead-Supported Assay: Integration of Optical Tweezers Assisted Fluorescence Imaging and Luminescence Confined Upconversion Nanoparticles Triggered Luminescent Resonance Energy Transfer (LRET).

Herein, a conceptual approach for significantly enhancing a bead-supported assay is proposed. For the fluorescence imaging technology, optical tweezers are introduced to overcome the fluid viscosity interference and immobilize a single tested bead at the laser focus to guarantee a fairly precise imaging condition. For the selection of fluorescent materials and the signal acquisition means, a type of innovative luminescence confined upconversion nanoparticle with a unique sandwich structure is specially designed to act as an efficient energy donor to trigger the luminescent resonance energy transfer (LRET) process. By further combining the double breakthrough with a molecular beacon model, the newly developed detection strategy allows for achieving a pretty high LRET ratio (≈ 88%) to FAM molecules and offering sound assay performance toward miRNA analysis with a detection limit as low as the sub-fM level, and is capable of well identifying single-base mismatching. Besides, this approach not only is able to accurately qualify the low-abundance targets from as few as 30 cancer cells but also can be employed as a valid cancer early warning tool for performing liquid biopsy.

Combining Holographic Optical Tweezers with Upconversion Luminescence Encoding: Imaging-Based Stable Suspension Array for Sensitive Responding of Dual Cancer Biomarkers.

Establishment of a stable analytical methodology with high-quality results is an urgent need for screening cancer biomarkers in early diagnosis of cancer. In this study, we incorporate holographic optical tweezers with upconversion luminescence encoding to design an imageable suspension array and apply it to conduct the detection of two liver cancer related biomarkers, carcinoembryonic antigen and alpha fetal protein. This bead-based assay is actualized by forming a bead array with holographic optical tweezers and synchronously exciting the upconversion luminescence of corresponding trapped complex beads fabricated with a simple one-step sandwich immunological recognition. Owing to the fact that these flowing beads are stably trapped in the focal plane of the objective lens which tightly converges the array of the laser beams by splitting a 980 nm beam using a diffraction optical element, a fairly stable excitation condition is achieved to provide reliable assay results. By further taking advantage of the eminent encoding capability of upconversion nanoparticles and the extremely low background signals of anti-Stokes luminescence, the two targets are well-identified and simultaneously detected with quite sound sensitivity and specificity. Moreover, the potential on-demand clinical application is presented by employing this approach to respond the targets toward complex matrices such as serum and tissue samples, offering a new alternative for cancer diagnosis technology.

Integrating optical tweezers with up-converting luminescence: a non-amplification analytical platform for quantitative detection of microRNA-21 sequences.

We report a single-microsphere based imaging assay method by integrating up-converting luminescence with optical tweezers for detecting microRNA-21 sequences. This method achieves a competitive detection limit of 12 fM with good selectivity and no dedicated signal amplification designs.

Dual Amplification Fluorescence Assay for Alpha Fetal Protein Utilizing Immunohybridization Chain Reaction and Metal-Enhanced Fluorescence of Carbon Nanodots.

As an emerging fascinating fluorescent nanomaterial, carbon nanodots (CDs) have attracted much attention owing of their unique properties such as small size, antiphotobleaching, and biocompatibility. However, its use in biomedical analysis is limited because of its low quantum yield. Herein, we constructed a dual amplification fluorescence sensor by incorporating immunohybridization chain reaction (immuno-HCR) and metal-enhanced fluorescence (MEF) of CDs for the detection of alpha fetal protein (AFP). The immunoplasmonic slide and detection antibodies-conjugated oligonucleotide initiator are served to capture and probe AFP molecules, respectively. Then, CD-tagged hairpin nucleic acids were introduced to trigger the HCR, in which the hairpin nucleic acid and oligonucleotide initiator are complementary. The interaction between CDs and the gold nanoisland film greatly improves the radiative decay rate, increases the quantum yield, and enhances the fluorescence emission of the CDs. Furthermore, the HCR provides secondary amplification of fluorescence intensity. Therefore, the MEF-capable immunohybridization reactions provide obvious advantages and result in exceptional sensitivity. In addition, the sandwich immunoassay method offers high specificity. The results show a wide linearity between the fluorescence intensity and AFP concentration over 5 orders of magnitude (0.0005-5 ng/mL), and the detection limit reaches as low as 94.3 fg/mL. This method offers advantages of high sensitivity and reliability, wide detection range, and versatile plasmonic chips, thus presenting an alternative for the technologies in biomedical analysis and clinical applications.