FISH-TAMB, a Fixation-Free mRNA Fluorescent Labeling Technique to Target Transcriptionally Active Members in Microbial Communities.

Keystone species or ecological engineers are vital to the health of an ecosystem; however, often, their low abundance or biomass present challenges for their discovery, identification, visualization and selection. We report the development of fluorescent in situ hybridization of transcript-annealing molecular beacons (FISH-TAMB), a fixation-free protocol that is applicable to archaea and bacteria. The FISH-TAMB method differs from existing FISH methods by the absence of fixatives or surfactants in buffers, the fast hybridization time of as short as 15 min at target cells' growth temperature, and the omission of washing steps.

Comparative and practical aspects of localization-based super-resolution imaging.

Super-resolution microscopy overcomes diffraction to generate images with superior resolution compared to conventional light microscopy. Localization-based super-resolution methods result in up to ten-fold improvement in resolution by determining the positions of fluorescent molecules with sub-pixel accuracy. This process critically depends on controlled emission at the level of individual fluorophores so that fluorescence is non-overlapping, allowing for accurate centroid determination of diffraction-limited spots by Gaussian fitting of the pixel intensities.

Multimodal optical microscope for detecting viability of mouse embryos in vitro.

We present a multimodal optical microscope that incorporates six imaging modalities on one common platform. The imaging modalities include three staring modes, optical quadrature microscopy (OQM), differential interference contrast (DIC) microscopy, and epi-fluorescence microscopy, and three scanning modes, confocal reflectance microscopy (CRM), confocal fluorescence microscopy (CFM), and two-photon microscopy (2PM). OQM reconstructs the amplitude and phase of an optically transparent specimen within a modified Mach-Zehnder configuration.