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Multiplexed cyclic imaging in expandable tissue gels has been extensively studied to visualize numerous biomolecules at a nanoscale resolution in situ. Previous studies have employed sparse labels, such as DAPI or lectin staining, as registration markers. However, these sparse labels do not adequately capture the full extent of deformation across the entire region of interest. To overcome this challenge, we propose the use of dense labels, specifically fluorophore N -hydroxysuccinimide (NHS)-ester staining, as registration markers to achieve highly accurate image registration. We first tested several NHS-functionalized fluorophores as fiducial markers and identified the proper candidates for three-dimensional (3D) multiplexed cyclic imaging. We analyzed the registration accuracy between DAPI and NHS-ester staining and illustrated that dense label-based registration provides a more accurate registration performance. In the multiplexed imaging of expanded specimens, we observed that repetitive expansion/shrinking processes and chemical treatments for signal elimination can induce 3D nonlinear distortion. This sample distortion can be mitigated by re-embedding the tissue gel or replacing the chemical de-staining process with photobleaching-based signal removal or computational signal unmixing. With such an optimized experimental setup, we demonstrated 3D multiplexed cyclic imaging with nanoscale precision image registration. Finally, we prove that dense biological structures, such as actin, can be used as registration markers to achieve high registration accuracy. We anticipate that the proposed dense labeling strategy will overcome the technical limitations of multiplexed cyclic imaging in expandable tissue gels, offering high-precision registration. We expect it to be widely adopted by the biological and medical communities.

Inside living organisms, proteins are self‐assembled into diverse 3D structures optimized for specific functions. This structure‐function relationship can be exploited to synthesize functional materials through biotemplating and depositing functional materials onto protein structures. However, conventional biotemplating faces limitations due to the predominantly intracellular existence of proteins and associated challenges in achieving tunability while preserving functionality. In this study, Conversion to Advanced Materials via labeled Biostructures (CamBio), an integrated biotemplating platform that involves labeling target protein structures with antibodies followed by the growth of functional materials, ensuring outstanding nanostructure tunability is proposed. Protein‐derived plasmonic nanostructures created by CamBio can serve as precise quantitative tools for assessing target species is demonstrated. The assessment is achieved through highly tunable and efficient surface‐enhanced Raman spectroscopy (SERS). CamBio enables the formation of dense nanogap hot spots among metal nanoparticles, templated by diverse fibrous proteins comprising densely repeated monomers. Furthermore, iterative antibody labeling strategies to adjust the antibody density surrounding targets, amplifying the number of nanogaps and consequently improving SERS performance are employed. Finally, cell‐patterned substrates and whole meat sections as SERS substrates, confirming their easily accessible, cost‐effective, scalable preparation capabilities and dimensional tunability are incorporated. This work demonstrates an integrated biotemplating that utilizes intracellular proteins to fabricate highly functional nanomaterials, called Conversion to advanced materials via labeled Biostructures (CamBio). This work exploits the structural characteristics of cells and tissues, employing them as Surface‐Enhanced Raman Spectroscopy (SERS) substrates by combining conventional fabrication and bio‐related techniques. This work offers the capability to transform advanced materials by adding biostructural benefits for a myriad of applications.

This chapter discusses the processing and rheological properties of carbon nanotubes (CNTs)‐based polymer nanocomposites, which provide more insights into structure‐property relationship. The two most important factors, which control the rheology and processing of the CNT‐based nanocomposites, are their dispersion state and concentration of CNTs. This chapter reviews the viscoelastic properties of polymer/CNT composites, which provide critical information regarding the state of dispersion of the nanotube and its mesoscale structure in the polymer matrix. The most familiar method for preparing polymer/CNT nanocomposites involves the mixing of CNTs and polymer using a suitable solvent. Furthermore, the viscoelastic properties also provide critical insight into the processing characteristics of these materials and could help predict the effect of nanoparticles on the final properties. Detailed understandings of the linear and nonlinear rheological properties and their relation with the underlying structure are essential for industrial success of polymer/CNT composites manufacturing and applications.

To avoid rhythm disturbance, sutures for ventricular septal defect closure have been traditionally placed 2∼5 mm or more away from the edge of the ventricular septal defect. However, the traditional suturing method appears to induce right bundle branch block and tricuspid valve regurgitation after ventricular septal defect closure more than our alternative technique, shallow suturing just at the edge of the ventricular septal defect (shallower bites at the postero-inferior margin). We aimed to verify our clinical experience of perimembranous ventricular septal defect repair. The alternative shallow suturing method has been applied since 2003 at our institution. We retrospectively reviewed the clinical data of 556 isolated perimembranous ventricular septal defect patients who underwent surgical closure from 2000 to 2019. We investigated the postoperative occurrence of right bundle branch block or progression of tricuspid regurgitation and analysed risk factors for right bundle branch block and tricuspid regurgitation. Traditional suturing method (Group T) was used in 374 cases (66.8%), and alternative suturing method (Group A) was used in 186 cases (33.2%). The right bundle branch block occurred more frequently in Group T (39.6%) than in Group A (14.9%). In multivariable logistic regression analysis, Group T and patch material were significant risk factors for late right bundle branch block. More patients with progression of tricuspid regurgitation were found in Group T. Shallow suturing just at the edge of the ventricular septal defect may reduce the rate of right bundle branch block occurrence and tricuspid regurgitation progression without other complications.

Aim： To determine the presence of voltage-gated K＋ （Kv） channels in bone marrow-derived human mesenchymal stem cells （hMSCs） and their impact on differentiation of hMSCs into adipocytes. Methods： For adipogenic differentiation, hMSCs were cultured in adipogenic medium for 22 d. The degrees of adipogenic differentiation were examined using Western blot, Oil Red 0 staining and Alamar assay. The expression levels of Kv channel subunits Kvl.1, Kvl.2, Kvl.3, Kvl.4, Kv2.1, Kv3.1, Kv3.3, Kv4.2, Kv4.3, and Kv9.3 in the cells were detected using RT-PCR and Western blot analysis. Results： The expression levels of Kv2.1 and Kv3.3 subunits were markedly increased on d 16 and 22. in contrast, the expression levels of other Kv channel subunits, including Kvl.1, Kvl.2, Kvl.3, Kvl.4, Kv4.2, Kv4.3, and Kv9.3, were decreased as undifferentiated hMSCs differentiated into adipocytes. Addition of the Kv channel blocker tetraethylammonium （TEA, 10 mmol/L） into the adipogenic medium for 6 or 12 d caused a significant decrease, although not complete, in lipid droplet formation and adipocyte fatty acid-binding protein 2 （aP2） expressions. Addition of the selective Kv2.1 channel blocker guangxitoxin （GxTX-1, 40 nmol/L） into the adipogenic medium for 21 d also suppressed adipogenic differentiation of the cells. Conclusion： The results demonstrate that subsets of Kv channels including Kv2.1 and Kv3.3 may play an important role in the differentiation of hMSCs into adipocytes.