Green-emitting water-soluble amino-ketoenole dye AmyGreen is proposed as an efficient fluorescent stain for visualization of bacterial amyloids in biofilms and the detection of pathological amyloids in vitro. This dye is almost non-fluorescent in solution, displays strong green emission in the presence of amyloid fibril of proteins. AmyGreen is also weakly fluorescent in presence to biomolecules that are components of cells, extracellular matrix or medium: nucleic acids, polysaccharides, lipids, and proteins. Thus, the luminescence turn-on behavior of AmyGreen can be utilized for visualization of amyloid components of bacterial biofilm extracellular matrix. Herein we report the application of AmyGreen for fluorescent staining of a number of amyloid-contained bacteria biofilms produced by Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Bordetella avium, and Staphylococcus aureus. The effectiveness of AmyGreen was compared to traditional amyloid sensitive dye Thioflavine T. The main advantage of AmyGreen (concentration 10–5 M) is a higher sensitivity in the visualization of amyloid biofilm components over Thioflavine T (10–4 M) as it was revealed when staining E. coli and K. pneumoniae bacterial biofilms. Besides, AmyGreen displays lower cross-selectivity to nucleic acids as demonstrated both in in-solution experiments and upon staining of eukaryotic human mesenchymal stem cells used as amyloid-free negative control over amyloid-rich bacterial biofilms. The results point to a lower risk of false-positive response upon determination of amyloid components of bacterial biofilm using AmyGreen. Co-staining of biofilm by AmyGreen and cellulose sensitive dye Calcofluor White show difference in their staining patterns and localization, indicating separation of polysaccharide-rich and amyloid-rich regions of investigated biofilms. Thus, we suggest the new AmyGreen stain for visualization and differentiation of amyloid fibrils in bacterial biofilms to be used solely and in combination with other stains for confocal and fluorescence microscopy analysis.

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An application of 2D-BN nanoparticles as a spectroscopic marker, weak luminescent marker and anticancer drug (doxorubicin, DOX) delivery system with protection properties was studied for the LNCaP strains of cancer cells using FTIR and Raman spectroscopy for analysing the cancer cells, cells with BN, the cancer cells with DOX, and the cancer cells with BN nanoparticles loaded by DOX. Study of IR absorption and Raman spectra of the LNCaP strains of cancer cells incubated with 2D-BN nanoparticles for 1 hour showed that the 2D-BN nanoparticles could pass through the cell membrane and localize inside the membrane or close to the membrane in the cytoplasm of the cells. We registered the spectra of the disturbed lipids during the DOX-2D-BN passing through the membrane. After incubation for 2 hours and more, spectral changes in other structural components of the cell (nuclei, cytoplasm, mitochondria) can be registered. Confocal microscopy showed that a gold nanostructured support enhances the fluorescence of the cancer cells with 2D-BN as well as that with DOX, however the double action of 2D-BN and DOX on the cancer cells aggravates the emission property of the studied system. An MTT test showed that the toxicity of DOX on the 2D-BN nanoparticles is less than that on the reference cells, and at the same time the efficiency of the DOX action on the cancer cells does not change.