Raghuraj S. Chouhan

Quantum dot conjugated S. cerevisiae as smart nanotoxicity indicators for screening the toxicity of nanomaterials

In this study, we have evaluated the toxicity of different forms of carbon nanotubes (CNTs) using S. cerevisiae-QD (SQD) bioconjugates as a novel fluorescent biological nanotoxicity indicator. A CNT mediated effect in SQD bioconjugates was used as an indicator for the changes occurring at the cell-membrane interfaces that induced disruption of membrane bound QDs resulting in the loss of fluorescence. Single, double and multiwalled carbon nanotubes (SWCNTs, DWCNTs and MWCNTs) were tested for their toxicities imposed on SQD bioconjugates. Bioconjugates exposed to varying concentrations of different forms of CNTs exhibited different modes of toxicities on SQD bioconjugates. SQD bioconjugates were highly responsive in the 0.1-10 μg mL-1 CNT concentration range after 1 h of exposure. The toxicity of CNTs was linked to the number of CNT walls. These results were further confirmed by SEM analysis and cell-viability tests that were consistent with the toxicity assays using fluorescent bioconjugates with different types of CNTs. SWCNTs imposed more severe cellular toxicity followed by MWCNTs and DWCNTs and the order of increasing cellular-damage by CNTs followed DWCNTs < MWCNTs < SWCNTs. This study speculates that the cell-injury by CNTs depends on their physical properties, such as layers of walls, non-covalent forces and dispersion states. Our results demonstrated a facile optical strategy that enables rapid and real-time cytotoxicity screening with yeast as model living-cells for engineering nanomaterials.

E. coli–quantum dot bioconjugates as whole-cell fluorescent reporters for probing cellular damage

A quantum dot (QD) conjugated whole-cell E. coli biosensor (E. coli-QD bioconjugates) was developed as a new molecular tool for probing cellular damage. The E. coli-QD bioconjugates were viable and exhibited fluorescence emission at 585 nm. Scanning electron microscopy (SEM) analysis of E. coli-QD bioconjugates revealed that the QDs were immobilized on the cell-surfaces and the fluorescence emission from QDs present on cell-surfaces was visualized by confocal microscopic examination. The E. coli-QD bioconjugates were employed as whole-cell fluorescent reporters that were designed to function as fluorescence switches that turn-off when cellular damage occurs. In this study, multi-walled carbon nanotubes (CNTs) were utilized as a model nanomaterial to probe cellular damage. Fluorescence spectra were recorded after the exposure of E. coli-QD bioconjugates with CNTs. We observed a strong correlation between fluorescence emission spectra, SEM and confocal microscopic analysis demonstrating that CNTs induced a dose and exposure time-dependent cellular toxicity. This toxicity mainly occurred by the physical interaction and cellular trafficking mechanisms that led to the collapse of the cellular structure and thus loss of fluorescence. The responses of E. coli-QD bioconjugates against CNTs were also visualized by simply exposing the cells to UV light and therefore rapid toxicity analysis and screening can be made. Our study demonstrated an easy and simple method to determine an important mechanistic perspective for the biological toxicity of chemicals or nanomaterials (NMs).

Nanomaterial resistant microorganism mediated reduction of graphene oxide

In this study, soil bacteria were isolated from nanomaterials (NMs) contaminated pond soil and enriched in the presence of graphene oxide (GO) in mineral medium to obtain NMs resistant bacteria. The isolated resistant bacteria were biochemically and genetically identified as Fontibacillus aquaticus. The resistant bacteria were allowed to interact with engineered GO in order to study the biotransformation in GO structure. Raman spectra of GO extracted from culture medium revealed decreased intensity ratio of ID/IG with subsequent reduction of CO which was consistent with Fourier transform infrared (FTIR) results. The structural changes and exfoliatied GO nanosheets were also evident from transmission electron microscopy (TEM) images. Ultraviolet-visible spectroscopy, high resolution X-ray diffraction (XRD) and current-voltage measurements confirmed the reduction of GO after the interaction with resistant bacteria. X-ray photoelectron spectroscopy (XPS) analysis of biotransformed GO revealed reduction of oxygen-containing species on the surface of nanosheets. Our results demonstrated that the presented method is an environment friendly, cost effective, simple and based on green approaches for the reduction of GO using NMs resistant bacteria.

Development of an immunoblot assay for carcinoembryonic antigen (CEA) in human serum using a portable UV illuminator

A rapid quantum dot (QD) based immunoblot assay is developed to detect CEA in human serum using a portable UV illuminator. Here, photostable QD-conjugated antibodies were utilized that bind CEA in human serum, previously blotted on a nitrocellulose membrane for point-of-care applications.

Quantum Dots Conjugated E. coli Living Cells as Fluorescent Reporters to Detect Cytotoxicity of Chemicals

Quantum dots (QDs) have attracted much of research interest in recent years for imaging, diagnostics, and therapy due to their unique optical properties, such as broad excitation spectra and long fluorescence stability. In this study, a controlled bioconjugation using CdTe QDs with gram negative E. coli cells was performed to develop QD-E. coli bioconjugates. These bioconjugates were used as whole cell living baits to determine cytotoxicity of model toxic chemicals, such as oxidative stress inducer (H2O2) and a pesticide (methyl viologen or paraquat). These chemicals over a wide concentration ranges were exposed to QD-E. coli bioconjugates that interacted with cells and the real time fluorescence responses, with QD-E. coli bioconjugates, were analyzed. The results showed that the fluorescent ability of QD-E. coli bioconjugates tend to diminish with increasing concentration of toxic chemicals. This stress is attributed to the damages occurred as a result of interaction of toxic chemicals to the cell-wall or membrane of cells that resulted in the loss of fluorescence signal. This loss in the fluorescence (signal off phenomena) of QD-E. coli bioconjugates can be used as probes to develop a variety of fluorescence-based detection kits for the rapid determination of toxic drugs or food sample testing.