Title: Data from: Video-rate raman-based metabolic imaging by airy light-sheet illumination and photon-sparse detection Dataset DOI: ### Pending DOI from DOE Data, code and/or products within this dataset support the follwoing manuscript: Manuscript Title: Video-rate raman-based metabolic imaging by airy light-sheet illumination and photon-sparse detection Journal: PNAS DOI: ### IN REVIEW Description/Abstract: Data supporting manuscript submitted to PNAS: Video-Rate Raman-based Metabolic Imaging by Airy Light-Sheet Illumination and Photon-Sparse Detection. The data set includes: [1] raw data and [2] related images used in the analyses described within the manuscript. Despite its massive potential, Raman imaging represents just a modest fraction of all research and clinical microscopy to-date. This is due to the ultralow Raman scattering cross-sections of most biomolecules that impose low-light or photon-sparse conditions. Bioimaging under such conditions is suboptimal, as it either results in ultralow frame-rates or requires increased levels of irradiance. Here, we overcome this tradeoff by introducing Raman imaging that operates at both video-rates and 1000-fold lower irradiance than state-of-the-art methods. To accomplish this, we deployed a judicially de-signed Airy light-sheet microscope to efficiently image large specimen regions. Further, we implemented sub-photon per pixel image acquisition and reconstruction to confront issues arising from photon sparsity at just msec integrations. We demonstrate the versatility of our approach by imaging a variety of samples, including the 3D metabolic activity of single microbial cells and the underlying cell-to-cell variability. To image such small-scale targets, we again harnessed photon-sparsity to increase magnification with-out a field-of-view penalty, thus, overcoming another key limitation in modern light-sheet microscopy. **Data Use**: *License*: [CC-BY 4.0 (https://creativecommons.org/licenses/by/4.0/)] *Recommended Citation*: Vasdekis AE (2023) Data from: Video-rate raman-based metabolic imaging by airy light-sheet illumination and photon-sparse detection [Dataset]. University of Idaho. ###Pending DOI LINK### Resource URL: https://data.nkn.uidaho.edu/dataset/data-video-rate-raman-based-metabolic-imaging-airy-light-sheet-illumination-and-photon Creator(s): 1. Full Name: Andreas E. Vasdekis Unique identifier: https://orcid.org/0000-0003-4315-1047 Affiliation(s): University of Idaho Other Contributor(s): NULL Publisher: University of Idaho Publication Year: 2023 Language(s): American English Subject(s): 1. NATURAL SCIENCES 1.3 Physical Science 1.6 Biological Science Keywords/Tags: Raman imaging, photon-sparse imaging, light-sheet microscopy Resource Type General: Dataset Dates: NULL Date available for the public: ### Pending release upon manuscript publication Sizes: 273.9 MB Format(s): Compressed data deirectory (zip) with txt, csv and tiff files. Version: NULL Funding References: U.S. Department of Energy Award: DE-SC0022282 URL: NULL Title: Integrative Imaging of Plant Roots during Symbiosis with Mycorrhizal Fungi Spatial/Geographical Coverage Location: NULL Temporal Coverage: NULL Granularity of the Data: NULL Contact Info: Contact Name: Andreas Vasdekis Contact Email: andreasv@uidaho.edu Related Content: Peer-reviewed manuscript-PNAS | ### Pending link to manuscript Project associated code-GitHub | https://github.com/aevasdekis/photon-sparse#photon-sparse Data Files: Figure 4: file directory containing data files necessary to compile figure 4 of associated manuscript Figure_4a.csv: data for compiling the histogram presented in figure 4a cell: ID of the cell observed sig_D2O: intensity values of a single cell grown in D2O bg_D2O: intensity values of background in D2O sig-bg_D2O: intensity difference between signal and background for a single cell grown in D2O sig_H2O: intensity values of a single cell grown in H2O [data only available for 1st 10 rows of data (cells h1-h10)] bg_H2O: intensity values of background in H2O [data only available for 1st 10 rows of data (cells h1-h10)] sig-bg_H2O: intensity difference between signal and background for a single cell grown in H2O [data only available for 1st 10 rows of data (cells h1-h10)] Figure_4b.csv: data for compiling the time-trace presented in figure 4b time_min: temporal axis values (in sec) cell: intensity values for a single cell grown in D2O cell_norm: intensity values for a single cell grown in D2O, normalized at t = 1 min Figure 5: file directory containing data files necessary to compile figure 5 of associated manuscript 220121_cloud_40x_90pct_focus.tif: 3D tif stack (xyt) representing single photon clouds as a function of time (t) of a single cell at focus Figure_5_error_analysis.csv: data of cell area error value represented in the insets of figure 5 photons-per-voxel: number of photons per unit voxel xy-area: cell area in the x-y plane in pixel units xy-area-per_norm: cell area in the x-y plane in pixel units normalized at 10 photons per voxel xz-area: cell area in the x-z plane in pixel units xz-area-per_norm: cell area in the x-z plane in pixel units normalized at 10 photons per voxel Figure 6: file directory containing data files necessary to compile figure 6 of associated manuscript cell_1x1_figure_6a.tif: 2D tif image of a Y. lipolytica cell (as per Fig. 5) under pixel-level projection cell_2x2_figure_6b.tif: 2D tif image of a Y. lipolytica cell (as per Fig. 5) under sub-pixel projection Figure_6_traces.csv: data for compiling the traces presented in the insets of figure 6a and 6b Header Key: distance-pixel: x-axis distance in pixel units distance-μm: x-axis distance in micron units trace_1x1: trace of the Y. lipolytica cell under pixel-level projection trace_2x2: trace of the Y. lipolytica cell under sub-pixel projection