Phase Behaviour and Miscibility Studies of Collagen/Silk Fibroin Macromolecular System in Dilute Solutions and Solid State

Miscibility is an important issue in biopolymer blends for analysis of the behavior of polymer pairs through the detection of phase separation and improvement of the mechanical and physical properties of the blend. This study presents the formulation of a stable and one-phase mixture of collagen and regenerated silk fibroin (RSF), with the highest miscibility ratio between these two macromolecules, through inducing electrostatic interactions, using salt ions. For this aim, a ternary phase diagram was experimentally built for the mixtures, based on observations of phase behavior of blend solutions with various ratios. The miscibility behavior of the blend solutions in the miscible zones of the phase diagram was confirmed quantitatively by viscosimetric measurements. Assessing the effects of biopolymer mixing ratio and salt ions, before and after dialysis of blend solutions, revealed the importance of ion-specific interactions in the formation of coacervate-based materials containing collagen and RSF blends that can be used in pharmaceutical, drug delivery, and biomedical applications. Moreover, the conformational change of silk fibroin from random coil to beta sheet, in solution and in the final solid films, was detected by circular dichroism (CD) and Fourier transform infrared spectroscopy (FTIR), respectively. Scanning electron microscopy (SEM) exhibited alterations of surface morphology for the biocomposite films with different ratios. Surface contact angle measurement illustrated different hydrophobic properties for the blended film surfaces. Differential scanning calorimetry (DSC) showed that the formation of the beta sheet structure of silk fibroin enhances the thermal stability of the final blend films. Therefore, the novel method presented in this study resulted in the formation of biocomposite films whose physico-chemical properties can be tuned by silk fibroin conformational changes by applying different component mixing ratios.info:eu-repo/semantics/publishedVersio

Phase behaviour and miscibility studies of collagen/silk fibroin macromolecular system in dilute solutions and solid state

Miscibility is an important issue in biopolymer blends for analysis of the behavior of polymer pairs through the detection of phase separation and improvement of the mechanical and physical properties of the blend. This study presents the formulation of a stable and one-phase mixture of collagen and regenerated silk fibroin (RSF), with the highest miscibility ratio between these two macromolecules, through inducing electrostatic interactions, using salt ions. For this aim, a ternary phase diagram was experimentally built for the mixtures, based on observations of phase behavior of blend solutions with various ratios. The miscibility behavior of the blend solutions in the miscible zones of the phase diagram was confirmed quantitatively by viscosimetric measurements. Assessing the effects of biopolymer mixing ratio and salt ions, before and after dialysis of blend solutions, revealed the importance of ion-specific interactions in the formation of coacervate-based materials containing collagen and RSF blends that can be used in pharmaceutical, drug delivery, and biomedical applications. Moreover, the conformational change of silk fibroin from random coil to beta sheet, in solution and in the final solid films, was detected by circular dichroism (CD) and Fourier transform infrared spectroscopy (FTIR), respectively. Scanning electron microscopy (SEM) exhibited alterations of surface morphology for the biocomposite films with different ratios. Surface contact angle measurement illustrated different hydrophobic properties for the blended film surfaces. Differential scanning calorimetry (DSC) showed that the formation of the beta sheet structure of silk fibroin enhances the thermal stability of the final blend films. Therefore, the novel method presented in this study resulted in the formation of biocomposite films whose physico-chemical properties can be tuned by silk fibroin conformational changes by applying different component mixing ratios228sem informaçãosem informaçã

A New Label-Free Technique for Analysing Evaporation Induced Self-Assembly of Viral Nanoparticles Based on Enhanced Dark-Field Optical Imaging

Nanoparticle self-assembly is a complex phenomenon, the control of which is complicated by the lack of appropriate tools and techniques for monitoring the phenomenon with adequate resolution in real-time. In this work, a label-free technique based on dark-field microscopy was developed to investigate the self-assembly of nanoparticles. A bio-nanoparticle with complex shape (T4 bacteriophage) that self-assembles on glass substrates upon drying was developed. The fluid flow regime during the drying process, as well as the final self-assembled structures, were studied using dark-field microscopy, while phage diffusion was analysed by tracking of the phage nanoparticles in the bulk solutions. The concentrations of T4 phage nanoparticles and salt ions were identified as the main parameters influencing the fluid flow, particle motion and, consequently, the resulting self-assembled structure. This work demonstrates the utility of enhanced dark-field microscopy as a label-free technique for the observation of drying-induced self-assembly of bacteriophage T4. This technique provides the ability to track the nano-sized particles in different matrices and serves as a strong tool for monitoring self-assembled structures and bottom-up assembly of nano-sized building blocks in real-time

Single Extracellular VEsicle Nanoscopy

Abstract Extracellular vesicles (EVs) and their cargo constitute novel biomarkers. EV subpopulations have been defined not only by abundant tetraspanins (e.g., CD9, CD63 and CD81) but also by specific markers derived from their source cells. However, it remains a challenge to robustly isolate and characterize EV subpopulations. Here, we combined affinity isolation with super‐resolution imaging to comprehensively assess EV subpopulations from human plasma. Our Single Extracellular VEsicle Nanoscopy (SEVEN) assay successfully quantified the number of affinity‐isolated EVs, their size, shape, molecular tetraspanin content, and heterogeneity. The number of detected tetraspanin‐enriched EVs positively correlated with sample dilution in a 64‐fold range (for SEC‐enriched plasma) and a 50‐fold range (for crude plasma). Importantly, SEVEN robustly detected EVs from as little as ∼0.1 μL of crude plasma. We further characterized the size, shape and molecular tetraspanin content (with corresponding heterogeneities) for CD9‐, CD63‐ and CD81‐enriched EV subpopulations. Finally, we assessed EVs from the plasma of four pancreatic ductal adenocarcinoma patients with resectable disease. Compared to healthy plasma, CD9‐enriched EVs from patients were smaller while IGF1R‐enriched EVs from patients were larger, rounder and contained more tetraspanin molecules, suggestive of a unique pancreatic cancer‐enriched EV subpopulation. This study provides the method validation and demonstrates that SEVEN could be advanced into a platform for characterizing both disease‐associated and organ‐associated EV subpopulations