Monitoring the biochemical changes occurring to human keratinocytes exposed to solar radiation by Raman spectroscopy.

Solar radiation exposure is recognised to be a significant contributor to the development of skin cancer. Monitoring the simultaneous and consecutive mechanisms of interaction could provide a greater understanding of the process of photocarcinogenesis. This work presents an analysis of the biochemical and morphological changes occurring to immortalised human epithelial keratinocyte (HaCaT) cell cultures exposed to simulated solar radiation (SSR). Cell viability was monitored with the aid of the Alamar Blue assay, morphological examination was done with haematoxylin and eosin staining (H&E) and changes to the biochemical constituents (nucleic acids and proteins) as a result of the radiation insult were demonstrated through a combination of Raman microspectroscopy and multivariate analysis of spectral patterns. The spectral results suggest that SSR induces changes to the conformational structure of DNA as an immediate result of the radiation, whereas alteration in the protein signature is mostly seen as a later response.

Developing Gold Nanoparticles-Conjugated Aflatoxin B1 Antifungal Strips.

Lateral flow immunochromatographic assays are a powerful diagnostic tool for point-of-care tests, based on their simplicity, specificity, and sensitivity. In this study, a rapid and sensitive gold nanoparticle (AuNP) immunochromatographic strip is produced for detecting aflatoxin B1 (AFB1) in suspicious fungi-contaminated food samples. The 10 nm AuNPs were encompassed by bovine serum albumin (BSA) and AFB1 antibody. Thin-layer chromatography, gel electrophoresis and nuclear magnetic resonance spectroscopy were employed for analysing the chemical complexes. Various concentrations of AFB1 antigen (0-16 ng/mL) were tested with AFB1 antibody-BSA-AuNPs (conjugated AuNPs) and then analysed by scanning electron microscopy, ultraviolet-visible spectroscopy, and Zetasizer. The results showed that the AFB1 antibody was coupled to BSA by the N-hydroxysuccinimide ester method. The AuNPs application has the potential to contribute to AFB1 detection by monitoring a visible colour change from red to purple-blue, with a detection limit of 2 ng/mL in a 96-well plate. The lateral flow immunochromatographic strip tests are rapid, taking less than 10 min., and they have a detection capacity of 10 ng/g. The smartphone analysis of strips provided the results in 3 s, with a detection limit of 0.3 ng/g for AFB1 when the concentration was below 10 ng/g. Excellent agreement was found with AFB1 determination by high-performance liquid chromatography in the determination of AFB1 among 20 samples of peanuts, corn, rice, and bread.

Design and Simple Assembly of Gold Nanostar Bioconjugates for Surface-Enhanced Raman Spectroscopy Immunoassays.

Immunoassays using Surface-Enhanced Raman Spectroscopy are especially interesting on account not only of their increased sensitivity, but also due to its easy translation to point-of-care formats. The bases for these assays are bioconjugates of polyclonal antibodies and anisotropic gold nanoparticles functionalized with a Raman reporter. These bioconjugates, once loaded with the antigen analyte, can react on a sandwich format with the same antibodies immobilized on a surface. This surface can then be used for detection, on a microfluidics or immunochromatographic platform. Here, we have assembled bioconjugates of gold nanostars functionalized with 4-mercaptobenzoic acid, and anti-horseradish peroxidase antibodies. The assembly was by simple incubation, and agarose gel electrophoresis determined a high gold nanostar to antibody binding constant. The functionality of the bioconjugates is easy to determine since the respective antigen presents peroxidase enzymatic activity. Furthermore, the chosen antibody is a generic immunoglobulin G (IgG) antibody, opening the application of these principles to other antibody-antigen systems. Surface-Enhanced Raman Spectroscopy analysis of these bioconjugates indicated antigen detection down to 50 µU of peroxidase activity. All steps of conjugation were fully characterized by ultraviolet-visible spectroscopy, dynamic light scattering, ζ -Potential, scanning electron microscopy, and agarose gel electrophoresis. Based on the latter technique, a proof-of-concept was established for the proposed immunoassay.