In this study, large-area hexagonal-packed Si nanorod (SiNR) arrays in conjunction with Au nanoparticles (AuNPs) were fabricated for surface-enhanced Raman spectroscopy (SERS). We have achieved ultrasensitive molecular detection with high reproducibility and spatial uniformity. A finite-difference time-domain simulation suggests that a wide range of three-dimensional electric fields are generated along the surfaces of the SiNR array. With the tuning of the gap and diameter of the SiNRs, the produced long decay length (>130 nm) of the enhanced electric field makes the SERS substrate a zero-gap system for ultrasensitive detection of large biomolecules. In the detection of R6G molecules, our SERS system achieved an enhancement factor of >10 with a relative standard deviation as small as 3.9-7.2% over 30 points across the substrate. More significantly, the SERS substrate yielded ultrasensitive Raman signals on long amyloid-β fibrils at the single-fibril level, which provides promising potentials for ultrasensitive detection of amyloid aggregates that are related to Alzheimer's disease. Our study demonstrates that the SiNRs functionalized with AuNPs may serve as excellent SERS substrates in chemical and biomedical detection.

In this work, maltodextrin-modified magnetic microspheres Fe3O4@SiO2-Maltodextrin (Fe3O4@SiO2-MD) with uniform size and fine morphology were synthesized through a facile and low-cost method. As the maltodextrins on the surface of microspheres were combined with maltose binding proteins (MBP), the magnetic microspheres could be applied to enriching standard MBP fused proteins. Then, the application of Fe3O4@SiO2-MD in one-step purification and immobilization of MBP fused proteins was demonstrated. For the model protein we examined, Fe3O4@SiO2-MD showed excellent binding selectivity and capacity against other Escherichia coli proteins in the crude cell lysate. Additionally, the maltodextrin-modified magnetic microspheres can be recycled for several times without significant loss of binding capacity.

Plant-hormone-initiated signaling pathways are extremely vital for plant growth, differentiation, development, and adaptation to environmental stresses. Hormonal perception by receptors induces downstream signal transduction mechanisms that lead to plant responses. However, conventional techniques—such as genetics, biochemistry, and physiology methods—that are applied to elucidate these signaling pathways can only provide qualitative or ensemble-averaged quantitative results, and the intrinsic molecular mechanisms remain unclear. The present study developed novel methodologies based on in vitro single-molecule fluorescence assays to elucidate the complete and detailed mechanisms of plant hormone signal transduction pathways. The proposed methods are based on multicolor total internal reflection fluorescence microscopy and a flow cell model for gas environment control. The methods validate the effectiveness of single-molecule approaches for the extraction of abundant information, including oligomerization, specific gas dependence, and the interaction kinetics of different components.

Cryptochromes are blue light receptors that mediate circadian rhythm and magnetic sensing in various organisms. A typical cryptochrome consists of a conserved photolyase homology region domain and a varying carboxyl-terminal extension across species. The structure of the flexible carboxyl-terminal extension and how carboxyl-terminal extension participates in cryptochrome’s signaling function remain mostly unknown. In this study, we uncover the potential missing link between carboxyl-terminal extension conformational changes and downstream signaling functions. Specifically, we discover that the blue-light induced opening of carboxyl-terminal extension in C. reinhardtii animal-like cryptochrome can structurally facilitate its interaction with Rhythm Of Chloroplast 15, a circadian-clock-related protein. Our finding is made possible by two technical advances. Using single-molecule Förster resonance energy transfer technique, we directly observe the displacement of carboxyl-terminal extension by about 15 Å upon blue light excitation. Combining structure prediction and solution X-ray scattering methods, we propose plausible structures of full-length cryptochrome under dark and lit conditions. The structures provide molecular basis for light active conformational changes of cryptochrome and downstream regulatory functions.

Subvisible particles are an ongoing problem in biotherapeutic injectable pharmaceutical formulations, and their identification is an important prerequisite for tracing them back to their source and optimizing the process. Flow imaging microscopy (FIM) is a favored imaging technique, mainly because of its ability to achieve rapid batch imaging of subvisible particles in solution with excellent imaging quality. This study used VGG16 after transfer learning to identify subvisible particle images acquired using FlowCam. We manually prepared standards for seven classes of particles, acquired the image information through FlowCam, and fed the images over 5 µm into VGG16 consisting of a convolutional base of VGG16 pre-trained with ImageNet data and a custom classifier for training. An accuracy of 97.51% was obtained for the test set data. The study also demonstrated that the recognition method using transfer learning outperforms machine learning methods based on morphological parameters in terms of accuracy, and has a significant training speed advantage over scratch-trained CNN. The combination of transfer learning and FIM images is expected to provide a general and accurate data-analysis method for identifying subvisible particles.

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