Controlling the surface functionalities of nanoporous alumina membranes

A new approach for controlling and to functionalise nanoporous anodic alumina oxide (AAO) membranes is described. Our approach is predicated on the remarkable stability of the silanised AAO surface during anodisation. Well ordered nanoporous AAO membranes with different external surface properties compared to the internal pore surface properties were prepared and characterised

Quantitative analysis of nanoparticle internalization in mammalian cells by high resolution X-ray microscopy

<p>Abstract</p> <p>Background</p> <p>Quantitative analysis of nanoparticle uptake at the cellular level is critical to nanomedicine procedures. In particular, it is required for a realistic evaluation of their effects. Unfortunately, quantitative measurements of nanoparticle uptake still pose a formidable technical challenge. We present here a method to tackle this problem and analyze the number of metal nanoparticles present in different types of cells. The method relies on high-lateral-resolution (better than 30 nm) transmission x-ray microimages with both absorption contrast and phase contrast -- including two-dimensional (2D) projection images and three-dimensional (3D) tomographic reconstructions that directly show the nanoparticles.</p> <p>Results</p> <p>Practical tests were successfully conducted on bare and polyethylene glycol (PEG) coated gold nanoparticles obtained by x-ray irradiation. Using two different cell lines, EMT and HeLa, we obtained the number of nanoparticle clusters uptaken by each cell and the cluster size. Furthermore, the analysis revealed interesting differences between 2D and 3D cultured cells as well as between 2D and 3D data for the same 3D specimen.</p> <p>Conclusions</p> <p>We demonstrated the feasibility and effectiveness of our method, proving that it is accurate enough to measure the nanoparticle uptake differences between cells as well as the sizes of the formed nanoparticle clusters. The differences between 2D and 3D cultures and 2D and 3D images stress the importance of the 3D analysis which is made possible by our approach.</p

Nanoparticle-Based Radiosensitization

Radiotherapy is a highly affordable treatment and provides many excellent outcomes [...

Nanoparticle-Based Radiosensitization

Radiotherapy is a highly multidisciplinary field with respect to its foundations of research and development, and in its clinical utility [...

The MTT Assay: Utility, Limitations, Pitfalls, and Interpretation in Bulk and Single-Cell Analysis

The MTT assay for cellular metabolic activity is almost ubiquitous to studies of cell toxicity; however, it is commonly applied and interpreted erroneously. We investigated the applicability and limitations of the MTT assay in representing treatment toxicity, cell viability, and metabolic activity. We evaluated the effect of potential confounding variables on the MTT assay measurements on a prostate cancer cell line (PC-3) including cell seeding number, MTT concentration, MTT incubation time, serum starvation, cell culture media composition, released intracellular contents (cell lysate and secretome), and extrusion of formazan to the extracellular space. We also assessed the confounding effect of polyethylene glycol (PEG)-coated gold nanoparticles (Au-NPs) as a tested treatment in PC-3 cells on the assay measurements. We additionally evaluated the applicability of microscopic image cytometry as a tool for measuring intracellular MTT reduction at the single-cell level. Our findings show that the assay measurements are a result of a complicated process dependant on many of the above-mentioned factors, and therefore, optimization of the assay and rational interpretation of the data is necessary to prevent misleading conclusions on variables such as cell viability, treatment toxicity, and/or cell metabolism. We conclude, with recommendations on how to apply the assay and a perspective on where the utility of the assay is a powerful tool, but likewise where it has limitations

DNA Double Strand Break and Response Fluorescent Assays: Choices and Interpretation

Accurately characterizing DNA double-stranded breaks (DSBs) and understanding the DNA damage response (DDR) is crucial for assessing cellular genotoxicity, maintaining genomic integrity, and advancing gene editing technologies. Immunofluorescence-based techniques have proven to be invaluable for quantifying and visualizing DSB repair, providing valuable insights into cellular repair processes. However, the selection of appropriate markers for analysis can be challenging due to the intricate nature of DSB repair mechanisms, often leading to ambiguous interpretations. This comprehensively summarizes the significance of immunofluorescence-based techniques, with their capacity for spatiotemporal visualization, in elucidating complex DDR processes. By evaluating the strengths and limitations of different markers, we identify where they are most relevant chronologically from DSB detection to repair, better contextualizing what each assay represents at a molecular level. This is valuable for identifying biases associated with each assay and facilitates accurate data interpretation. This review aims to improve the precision of DSB quantification, deepen the understanding of DDR processes, assay biases, and pathway choices, and provide practical guidance on marker selection. Each assay offers a unique perspective of the underlying processes, underscoring the need to select markers that are best suited to specific research objectives