Complex target SELEX-based identification of DNA aptamers against Bungarus caeruleus venom for the detection of envenomation using a paper-based device.

Complex target SELEX always have been an intriguing approach to the scientific community, as it offers the potential discovery of novel biomarkers. We herein successfully performed SELEX on Bungarus caeruleus venom to develop a panel of highly affine aptamers that specifically recognizes the B. caeruleus (common krait) venom and was able to discriminate the B. caeruleus venom from Cobra, Russell's, and Saw-scaled viper's venom. The aptamers generated against the crude venom also lead to the identification of the specific component of the venom, which is ß-Bungarotoxin, a toxin uniquely present in the B. caeruleus venom. The best performing aptamer candidates were used as a molecular recognition element in a paper-based device and were able to detect as low as 2 ng krait venom in human serum background. The developed aptamer-based paper device can be used for potential point-of-care venom detection applications due to its simplicity and affordability.

GOLD SELEX: a novel SELEX approach for the development of high-affinity aptamers against small molecules without residual activity.

GOLD SELEX, a novel SELEX approach has been developed that obviates the need for target immobilization for aptamer development. The approach purely relies on the affinity of the aptamers towards its target, to get detached from the gold nanoparticle (GNP) surface (weak attraction) after binding with its target. Thus, only the completely detached aptamers are selected for the next round of SELEX. This, in-process, also addresses the issue of residual binding and thus improves the sensitivity of the developed aptamers. As a proof of concept for establishing the utility of the approach for small molecules, we have developed aptamers against dichlorvos (DV), a pesticide in just 8 rounds. Using these aptamer candidates, we have developed an aptamer-NanoZyme (GNP having peroxidase mimic activity) based colorimetric assay. The developed aptamer displayed high affinity (Kd in sub micromolar range) and selectivity for DV. The developed assay could detect as low as 15 µM DV. The best-performing aptamer was also able to work in real samples like river water and commercial apple juice. The GOLD SELEX approach developed in this study, we believe, can act as a template for future SELEX strategy development and can replace the conventional SELEX strategy.

Application of aptamers as molecular recognition elements in lateral flow assays.

Owing to their ease in operation and fast turnaround time, lateral flow assays (LFAs) are increasingly being used as point-of-care diagnostic tests for variety of analytes. In a majority of these LFAs, antibodies are used as a molecular recognition element. Antibodies have a number of limitations such as high batch-to-batch variation, poor stability, long development time, difficulty in functionalization and need for ethical approval and cold chain. All these factors pose a great challenge to scale up the antibody-based tests. In recent years, the advent of aptamer technology has made a paradigm shift in the point-of-care diagnostics owing to the various advantages of aptamers over antibodies that favour their adaptability on a variety of sensing platforms including the lateral flow. In this review, we have highlighted the advantages of aptamers over antibodies, suitability of aptamers for lateral flow platforms, different types of aptamer-based LFAs and various labels for aptamer-based LFAs. We have also provided a summary of the applications of aptamer technology in LFAs for analytical applications.

Developing single-entity theranostic: drug-based fluorescent nanoclusters with augmented cytotoxicity.

AIM: To develop methotrexate (MTX) templated luminescent gold nanoclusters (NCs) as a single unit nanotheranostic for cancer therapy and to assess its potential as an alternative to the parent drug, for drug delivery vehicles (DDVs). METHODS: Theranostics were synthesized and extensively characterized. The stability of the theranostic and its bioimaging aptitude were evaluated. The antiproliferative propensity of the theranostic was gauged with cell viability assays and was supplemented with cytometry-based assays. Feasibility of delivering the MTX NCs instead of parent drug on a DDV was also checked. RESULTS: MTX NCs displayed remarkable physical characteristics and augmented cytotoxicity with a robust stability in phosphate-buffered saline and serum. MTX NCs also demonstrated their amenability to being loaded on a DDV (chitosan folic acid nanoparticles) while retaining their physical and cytotoxic profile. CONCLUSION: Generation of next level drug-based theranostics with the potential of replacing the free drug in drug delivery platforms.

dGTP-Templated Luminescent Gold Nanocluster-Based Composite Nanoparticles for Cancer Theranostics.

dGTP-templated facile synthesis of luminescent gold nanoclusters (Au NCs) in the presence of cisplatin following PEG coating developed spherical composite NPs. The composite NPs delivered cisplatin efficiently into HeLa cells to induce apoptosis-mediated cell death, and simultaneously bioimaged the cellular uptake.