Biocompatible thiolate protected copper nanoclusters for an efficient imaging of lung cancer cells.

We report, the one-pot synthesis of water-soluble and biocompatible 3-mercaptopropylsulfonate (MPS) protected novel copper nanoclusters (CuNCs). Interestingly, the TEM image of MPS protected CuNCs exhibits an ultrasmall nanoclusters of particle size <2-nm, similar to its Au and Ag analogue. The hydrophilic and biocompability property of thiolate protected CuNCs. i.e., MPS stabilized CuNCs and its luminescent nature gave rise to maximum quantum yield of 1.5%. Further, as achieved CuNCs was investigated for haemocompatibility, cell viability and fluorescent microscopic analysis with A549 lung cancer cell line. Haemolytic study was examined using human RBCs in the concentration range of 4 to 22 µg/mL for which 7.5% of haemolysis was obtained for an optimum concentration of 22 µg/mL of CuNCs. The cell viability analysis was carried out by MTT assay using A549 lung cancer cells for the minimum (10 µg/mL) and maximum (45 µg/mL) concentration of CuNCs which reports 93.1% and 38.2% cell viability respectively. The inverted light microscopic images from the control and CuNCs treated (20 µg/mL) cells exhibited an excellent biocompatibility with a normal morphology. Upon increasing the concentration of CuNCs upto 45 µg/mL, the cell viability trends to decrease and the cell morphology also denature gradually. Further, the bio-imaging application of CuNCs was analyzed with A549 lung cancer cells. The efficient imaging with CuNCs treated (20 µg/mL) A549 cells resulted in a green colour emission using FITC filter (460- 490 nm). Thereby the obtained results confirm the applicability of CuNCs for the biomedical and cancer diagnosis applications.

Scanometric Detection of Tomato Leaf Curl New Delhi Viral DNA Using Mono- and Bifunctional AuNP-Conjugated Oligonucleotide Probes.

Scanometric detection of tomato leaf curl New Delhi viral DNA using AuNP-conjugated mono- and bifunctional oligo probes through direct DNA hybridization assay (DDH assay) and sandwich DNA hybridization assay (SDH assay) with silver enhancement was developed. Tomato leaf curl New Delhi virus (ToLCNDV) coat protein gene-specific thiol-modified ssoligo probes were used for the preparation of mono- and bifunctional AuNP-ssoligo probe conjugates (signal probes). ssDNA arrays were prepared using polymerase chain reaction (PCR), rolling circle amplification (RCA), genomic DNAs fragments, and phosphate-modified positive control/capture probes through 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide/1-methylimidazole conjugation on the amine-modified glass slide (GS) surface. In the DDH assay, signal probes were directly hybridized with ssDNA array of positive control and ToLCNDV DNA samples and the detection signals were amplified by silver enhancement. Dark black/gray colors were developed on the GS by the result of Ag enhancement, which can be visualized and discriminated by the naked eye. The images were captured using a simple flatbed scanner, and the determined amounts of signal probes were hybridized with their target DNA. Similarly, the SDH assay also performed through two rounds of hybridization between capture probes and target DNA; target DNA and signal probes followed by silver enhancement. The detection signals were found higher in the PCR sample than the RCA and genomic DNA samples because of the presence of increased copy numbers of complementary DNAs in PCR samples. Further, bifunctional AuNP-ssoligo probe shows higher intensity of detection signal than monofunctional probes because it can be hybridized with both strands of dsDNA targets. Moreover, the DDH-based scanometric method showed higher detection sensitivity than the SDH assay-based scanometric method. Overall, bifunctional signal probes showed more detection sensitivity than monofunctional probes in scanometric methods based on both DDH and SDH assays. The limit of detection of this developed scanometric method was optimized (100 zM to 100 pM concentration). Further, DDH assay-based scanometric method shows significant advantages over the SDH assay method, such as cost-effectiveness, because it requires only single probes (signal probes), less time-consuming by the need of only single-step hybridization, and higher detection sensitivity (up to zM). To the best of our knowledge, this is the first attempt made to develop a scanometric-based nanoassay method for the detection of plant viral DNA. This approach will be a remarkable milestone for the application of nanotechnology in the development of nanobiosensor for plant pathogen detection.

Developing a Programmable, Self-Assembling Squash Leaf Curl China Virus (SLCCNV) Capsid Proteins into "Nanocargo"-like Architecture.

A new era has begun in which pathogens have become useful scaffolds for nanotechnology applications. In this research/study, an attempt has been made to generate an empty cargo-like architecture from a plant pathogenic virus named Squash leaf curl China virus (SLCCNV). In this approach, SLCCNV coat protein monomers are obtained efficiently by using a yeast Pichia pastoris expression system. Further, dialysis of purified SLCCNV-CP monomers against various pH modified (5-10) disassembly and assembly buffers produced a self-assembled "Nanocargo"-like architecture, which also exhibited an ability to encapsulate magnetic nanoparticles in vitro. Bioinformatics tools were also utilized to predict the possible self-assembly kinetics and bioconjugation sites of coat protein monomers. Significantly, an in vitro biocompatibility study using SLCCNV-Nanocargo particles showed low toxicity to the cells, which eventually proved as a potential nanobiomaterial for biomedical applications.

Auric Chloride Induced Micellization on Fractal Patterned Dicationic Amphiphiles and Stabilization of Gold Nanoparticles.

The present article reports the development of sunlight-mediated rapid synthesis of bile acid derived dicationic amphiphiles, namely, dicationic cysteamine-conjugated cholic acid (DCaC), dicationic cysteamine-conjugated deoxycholic acid (DCaDC), and dicationic cysteamine-conjugated lithocholic acid (DCaLC) by adopting thiol-yne click chemistry approach. The auric chloride (AuHCl4) induced micellization of amphiphiles from fractal pattern to chainlike aggregates was examined by critical micelle concentration measurements, quenching studies, field emission scanning electron microscopy, and optical microscopy techniques. The micelles thus formed act as ideal templates for the stabilization of gold nanoparticles (AuNPs) and exhibit good stability for more than 6 months. The synthesized AuNPs were characterized using UV-visible spectroscopy, high-resolution transmission electron microscopy, DLS, zeta potential, and contact angle measurements. These NPs showed high salt tolerance, and the levels were found to be 420, 460, and 580 mM for DCaC-, DCaDC-, and DCaLC-capped AuNPs, respectively.

Role of Surface Hydrophobicity of Dicationic Amphiphile-Stabilized Gold Nanoparticles on A549 Lung Cancer Cells.

Herein, we report the surface functionality of dicationic cysteamine conjugated cholic acid (DCaC), dicationic cysteamine conjugated deoxycholic acid (DCaDC), and dicationic cysteamine conjugated lithocholic acid (DCaLC) templated gold nanoparticles (AuNPs) on mammalian cells. The haemocompatibility of the synthesized NPs was evaluated by in vitro hemolysis and erythrocyte sedimentation rate using human red blood cells (RBCs). In all of the systems, no toxicity was observed on human erythrocytes (RBCs) up to the concentration of 120 µg/mL. The anticancer activity of these dicationic amphiphile-stabilized AuNPs on A549 lung cancer cells was demonstrated by in vitro cell viability assay, intracellular reactive oxygen species estimation by DCFH-DA, apoptosis analysis using AO-EtBr fluorescence staining, DNA fragmentation analysis by agarose gel electrophoresis, and western blot analysis of caspase-3 expression. These results suggest that the cytotoxicity of AuNPs to A549 cells increase with the dose and hydrophobicity of amphiphiles and were found to be in the order: DCaLC-AuNPs > DCaDC-AuNPs > DCaC-AuNPs.