Journal: The Review of scientific instruments

Article Title: Atom Probe Tomography Using an Extreme Ultraviolet Trigger Pulse

doi: 10.1063/5.0160797

Figure Lengend Snippet: (a) Schematic of the EUV beamline. The EUV light is generated collinearly with the NIR driving field in a noble gas filled capillary. The EUV light is isolated from the NIR light by means of two flat mirrors (M1 and M2) and Al filters (i.e. foils). When a toroid (T1) is inserted into the beam path, the light is reflected from a grating (G1) and focused onto a CCD camera for spectral analysis. When a toroid (T1) is removed from the beam path the EUV light transmits past the toroid and through a circular aperture (A1) before reflecting off a flat mirror (M3) and then a toroid (T2) which focuses the light inside the APT analysis chamber and onto the APT specimen apex. During alignments utilizing NIR light, the ultra high vacuum region is separated from the beamline via a sapphire window mounted on a gate valve (GV1). During experiments with EUV light, the ultra high vacuum analysis region is separated from the beamline via an Al filter mounted on a gate valve (GV2). When the instrument is idle, a standard steel gate valve (GV3) is closed to provide robust isolation between the ultrahigh vacuum chamber and the beamline. Two EUV sensitive photodiodes can be inserted into the beampath (PD1, PD2) for power measurements. The focal spot is steered onto the tip by a computer controlled tip/tilt actuator on which a toroid (T2) is mounted. (b) The beamline setup with each of the major components identified.

Article Snippet: The imaging spectrometer consists of the toroid (T1), grating (G1) set up for conical diffraction (500/mm and plane-ruled at 4.7°), and CCD camera (Greateyes, ALEX-s 2k512 BI) set up to image the exit of the capillary to the CCD camera in a 1:1 imaging configuration.

Techniques: Generated, Isolation

Journal: The Review of scientific instruments

Article Title: Atom Probe Tomography Using an Extreme Ultraviolet Trigger Pulse

doi: 10.1063/5.0160797

Figure Lengend Snippet: (a) Calculated reflectivity (p-pol, 6° grazing angle-of-incidence) of the coatings used on the grazing incidence optics present in the beamline. Note that nominally Si coated optics are expected to have a native oxide after exposure to air around 1 nm thick. (b) Calculated transmission (normal angle of incidence) for potentially oxidized Al filters present in the beamline. The dashed lines are calculated for 200 nm and 1 μm thick unoxidized Al. The solid lines are for the same filters with 16 nm and 33 nm of Al2O3 added, respectively. The hollow circles and squares show the measured transmission of two Al filters present in the beamline that are nominally 200 nm and 1 μm thick, respectively. (c) Calculated beamline throughput for various configurations including at the CCD camera and at the APT specimen location. Note that these calculations are smoothed by the spacing of the EUV harmonics (3 eV). (d) The spectra ‘as collected’ on the CCD for EUV light generated with different noble gases with two 1 μm Al filters (Xe) and three 1 μm filters (Kr and Ar). (e) The ‘at source’ spectra calculated by dividing the ‘as collected’ spectra by the beamline throughput to the CCD. (f) The ‘at APT specimen’ spectra calculated by multiplying the ‘at source’ spectra by the beamline throughput to the APT chamber (with a 1 μm Al filter inserted).

Article Snippet: The imaging spectrometer consists of the toroid (T1), grating (G1) set up for conical diffraction (500/mm and plane-ruled at 4.7°), and CCD camera (Greateyes, ALEX-s 2k512 BI) set up to image the exit of the capillary to the CCD camera in a 1:1 imaging configuration.

Techniques: Transmission Assay, Generated

Journal: The Review of scientific instruments

Article Title: Atom Probe Tomography Using an Extreme Ultraviolet Trigger Pulse

doi: 10.1063/5.0160797

Figure Lengend Snippet: (a) Schematic of a 2D beam profile measurement. The EUV light strikes a Ce:YAG crystal and the fluorescence is imaged onto a CCD camera at a ≈ 45° viewing angle. (b) An example 2D beam profile after the image is corrected for viewing angle. Here the beam size is comparable to the resolution of the camera imaging optics making quantitative analysis difficult. (c) Schematic of a ‘knife-edge’ beam profile measurement. Here a grounded local electrode puck was modified to have a flat metal surface intersect the beam. A piece of razor blade is mounted on a specimen puck held at +2.5 kV and slowly translated through the beam as the photoemission current is measured. The resulting one-dimensional profiles (d-e) are fit to a Gaussian beam model to extract the fwhm parameters.

Article Snippet: The imaging spectrometer consists of the toroid (T1), grating (G1) set up for conical diffraction (500/mm and plane-ruled at 4.7°), and CCD camera (Greateyes, ALEX-s 2k512 BI) set up to image the exit of the capillary to the CCD camera in a 1:1 imaging configuration.

Techniques: Fluorescence, Imaging, Modification

Journal: bioRxiv

Article Title: Super-resolved, three-dimensional spatial transcriptomics reveals cell-type and brain-region-specific modulation of key epitranscriptomic switches following adolescent alcohol exposure

doi: 10.64898/2026.01.22.698893

Figure Lengend Snippet: (a) Schematics showing the experimental workflow of RESOLution enhanced Visualization using Expansion-coupled FISH (RESOLVE-FISH). (i) Brain tissue undergoes Hybridization Chain Reaction (HCR)-RNA-FISH and immunostaining to label the target RNAs and proteins, respectively. A multifunctional anchor, glycidyl methacrylate (GMA), is introduced to covalently retain the target biomolecules and labels (rectangular box). (ii) The tissue is embedded in a swellable hydrogel to form a tissue-hydrogel composite. (iii) The tissue is homogenized and expanded, with the target biomolecules and labels retained (circular box). (b) Schematics showing the computational and analysis workflow of RESOLVE-FISH. (i) Cytoplasmic labels are used to segment and generate cell masks. (ii) The segmented cells are classified based on the cell-type markers. (iii) Individual RNA puncta are counted in three-dimensions (3D) within each cell mask, enabling spatially-resolved quantification of RNA in single cells across different cell types. (c) Fluorescent images from rat basolateral amygdala (BLA) demonstrating the computational and analysis workflow of RESOLVE-FISH. (i) The cytoplasm is labeled using ribosomal RNA (rRNA, magenta). (ii-1) A cell-type marker, NeuN (cyan, neuronal), is used to determine the cell types. (ii-2) Cell segmentation is performed using the cytoplasmic label to generate the cell masks. (iii) RNA puncta (green) are detected and quantified in 3D within each segmented cell, allowing for cell-type-specific quantification of RNA. Scale bars, 10 µm (35 µm). Here and after, scale bars are provided at the pre-expansion scale (with the corresponding post-expansion size indicated in brackets). (d) A 3D rendered volumetric image (left) of a neuronal cell from rat BLA and a two-dimensional (2D) Z-slice image (right) of the same cell. The 2D image roughly corresponds to the cross-section indicated by the dashed lines in the 3D rendering. Nuclear label (DAPI, blue), cytoplasmic label (rRNA, magenta), and Mettl3 RNA puncta (green) are shown. Scale bars, 5 µm (17.5 µm) (e) Schematics showing a rat brain coronal section highlighting three brain regions used in this study: the central amygdala (CeA), BLA, and hippocampus. The inset shows a zoomed-in view of the hippocampus (dashed box) highlighting the subregions used in the study: CA1, CA3, and dentate gyrus (DG).

Article Snippet: In parallel, fluorescently labeled HCR hairpins (B1 labeled with Alexa Fluor 488, B3 labeled with Alex Fluor 594, Molecular Instruments, Inc.) were snap-cooled by heating to 95°C for 90 seconds, then allowed to cool at RT in the dark for 30 minutes.

Techniques: Hybridization, Immunostaining, Labeling, Marker

Journal: The Plant Journal

Article Title: Single-nucleotide and long-patch base excision repair of DNA damage in plants

doi: 10.1111/j.1365-313X.2009.03994.x

Figure Lengend Snippet: Repair of base DNA damage by Arabidopsis cell extracts. (a) Schematic diagram of molecules used as DNA substrates. Double-stranded oligonucleotides contained a lesion (X = uracil or AP site) at an Hpa II site on the upper strand. The Alexa Fluor-labeled 5′ end of the lower strand is indicated by a star. The size of the 5′ end-labeled fragment generated after Hpa II digestion of the fully repaired product is indicated on the right. (b) DNA duplexes containing U, a natural AP site, or THF opposite G were incubated with Arabidopsis cell extract at 30°C for 3 h in a reaction mixture containing either dCTP or all four dNTPs, as indicated. Reaction products were digested with Hpa II, separated in a 12% denaturing polyacrylamide gel and quantified by fluorescence scanning. (c) A DNA duplex containing a U:G mispair was incubated with Arabidopsis cell extracts at 30°C in a reaction mixture containing either dCTP or all four dNTPs. Reactions were stopped at the indicated times and products were analyzed as described above. The percentage of fully repaired DNA product for reactions containing only dCTP (•) or all four dNTPs (○) is shown in the bottom panel. Values are means with standard errors from two independent experiments.

Article Snippet: Fluorescein- or Alexa-labeled DNA was visualized in an FLA-5100 imager, and data were analyzed using M ultigauge (Fujifilm, http://www.fujifilm.com ).

Techniques: Labeling, Generated, Incubation, Fluorescence

Journal: The Plant Journal

Article Title: Single-nucleotide and long-patch base excision repair of DNA damage in plants

doi: 10.1111/j.1365-313X.2009.03994.x

Figure Lengend Snippet: Uracil excision repair involves single-nucleotide insertion and long-patch DNA synthesis. Arabidopsis cell extract was incubated with duplex DNA that contained a U residue in the upper strand, and which was labeled at the 5′ end of the upper strand with fluorescein and at the 5′ end of the lower strand with Alexa Fluor. Reactions were incubated at 30°C for 3 h in a reaction mixture containing either dCTP or all four dNTPs, as indicated. Reaction products were separated in a 12% denaturing polyacrylamide gel either before (lanes 2–5) or after (lanes 6–9) Hpa II digestion. Fluorescein- (lanes 2–5) and Alexa Fluor-labeled fragments (lanes 6–9) were detected by fluorescence scanning.

Article Snippet: Fluorescein- or Alexa-labeled DNA was visualized in an FLA-5100 imager, and data were analyzed using M ultigauge (Fujifilm, http://www.fujifilm.com ).

Techniques: DNA Synthesis, Incubation, Labeling, Fluorescence