The application of optical microscopy in four-dimensional (spatial and temporal) super-resolution imaging poses challenges because of the requirement of a long acquisition time or high illumination intensity. In this paper, we introduce simultaneous two-angle axial ratiometry (STARII) for <20 nm axial super-resolution imaging and for fast and long-term imaging of live cells up to hundreds of frames per second. This method involves recording two raw images in two incident angle channels in the context of evanescent wave illumination and obtaining the corresponding intensity ratio. Furthermore, we demonstrate the combination of STARII with the lateral superresolution method to resolve three-dimensional nanoscale structures of microtubules and to visualize the long-term dynamical plasma membrane curvature and fast remodeling of endoplasmic reticulum tubule meshwork and three-way junctions. These demonstrations indicate an important potential application of STARII in investigating nanoscale cellular complex processes in the native state.
In this contribution, we conduct a multi-angular analysis of the interdisciplinarity of Nobel Prize winning research compared to non-Nobel Prize winning articles, based on a large data set. Here interdisciplinarity is measured by the diversity of references, using two true diversity indicators.Articles mentioned by the Nobel Prize committee in Physiology or Medicine (in short: NP articles) awarded during the period from 1900 to 2016 are the focus of our research. These articles are compared with those in a dataset of articles that do not include a Nobel Prize winner among their authors.Moreover, these non-NPs articles were not only published in the same year and in the same research field as the NP ones but were also dealing with the same research topic (such articles are referred to as non-NP articles).The results suggest that the topic-related knowledge included in Nobel Prize winning work is higher than that in non-NPs, hence with lower interdisciplinarity than the latter. Our findings provide useful clues to better understand the characteristics of transformative research, here represented 2 by key publications by Nobel Prize laureates in Physiology or Medicine, and their pattern of knowledge integration.
Being the established imaging tool for cell membrane-associated studies, total internal reflection fluorescence microscopy (TIRFM) still has some limitations. The most important one is the inhomogeneous evanescent excitation field mainly caused by the large-angle and fixed-azimuth illumination scheme, which can be eliminated by using ring-shaped illumination (ring TIRFM). However, it is challenging in assembling a ring TIRFM system with precise parameter control that works well. Here we emphasize the quantification of the ring TIRFM system and introduce a robust calibration routine to simultaneously rectify the asymmetry of the spinning light beam and determine the crucial experimental parameter, i.e., the incident angle. The calibration routine requires no specific sample preparation and is entirely based on the automatic back focal plane manipulation, avoiding possible errors caused by the sample difference and manual measurement. Its effectiveness is experimentally demonstrated by both the qualitative and quantitative comparisons of the images acquired using different samples, illumination schemes, and calibration approaches. These characteristics should enable our approach to greatly improve the practicability of TIRFM in life sciences.
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