High-throughput and green optical sensing of thiocyanate in human saliva based on microplates and an overhead book scanner as detector.

An instrumental-free, high-throughput assay has been developed for the quantification of thiocyanate in human saliva. The proposed green method is based on the rapid reaction of the analyte with Fe(III) under acidic pH in a microplates format to form a colored complex that is captured as an image by an overhead book scanner. Optimization included the effects of the amount concentration of Fe(III), acidity and reaction time / complex stability using a total volume of 300 µL per well. Validation towards the matrix effect was focused on the specific application and was performed using both artificial and human saliva. The linearity of the developed assay was up to 500 µM thiocyanate offering a lower limit of quantification (LLOQ) of 30 µM. The green potentials were evaluated by both the Green Analytical Procedure (GAPI) and Blue Applicability Grade (BAGI) indexes. The thiocyanate content in the saliva of non-smoking volunteers ranged between 750 and 1350 µΜ, while elevated concentrations were verified in smoking individuals (1860-3080 µΜ). Statistical agreement with a corroborative method was assessed using the Bland-Altman plot.

Integrated Skin Transcriptomics and Serum Multiplex Assays Reveal Novel Mechanisms of Wound Healing in Diabetic Foot Ulcers.

Nonhealing diabetic foot ulcers (DFUs) are characterized by low-grade chronic inflammation, both locally and systemically. We prospectively followed a group of patients who either healed or developed nonhealing chronic DFUs. Serum and forearm skin analysis, both at the protein expression and the transcriptomic level, indicated that increased expression of factors such as interferon-γ (IFN-γ), vascular endothelial growth factor, and soluble vascular cell adhesion molecule-1 were associated with DFU healing. Furthermore, foot skin single-cell RNA sequencing analysis showed multiple fibroblast cell clusters and increased inflammation in the dorsal skin of patients with diabetes mellitus (DM) and DFU specimens compared with control subjects. In addition, in myeloid cell DM and DFU upstream regulator analysis, we observed inhibition of interleukin-13 and IFN-γ and dysregulation of biological processes that included cell movement of monocytes, migration of dendritic cells, and chemotaxis of antigen-presenting cells pointing to an impaired migratory profile of immune cells in DM skin. The SLCO2A1 and CYP1A1 genes, which were upregulated at the forearm of nonhealers, were mainly expressed by the vascular endothelial cell cluster almost exclusively in DFU, indicating a potential important role in wound healing. These results from integrated protein and transcriptome analyses identified individual genes and pathways that can potentially be targeted for enhancing DFU healing.