Raw microscopy data accompanying "Applying coherent Raman microscopy to biological phenotyping"

Description This deposition contains the raw microscopy images underlying the Arcadia Science pub "Applying coherent Raman microscopy to biological phenotyping." The study compares coherent Raman microscopy (CRM) against conventional confocal fluorescence imaging for label-free phenotyping of the unicellular alga Chlorella vulgaris, using two strains: Wild type (EV_CTL) — the parent strain (211/11p) stably expressing Cas9 and carrying an empty vector; used as the control. ADA1 knockout (ADA_mut / ADAKO) — a mutant in which a C. vulgaris ortholog of human ADA1 (UniProt KAI3436177.1) was knocked out by targeting the first exon with S. pyogenes Cas9. A frameshift was confirmed by Sanger sequencing. ADA1 is a lipid-metabolism gene, and the mutant was expected to show elevated lipid content detectable by CRS. Analysis code, figure-generation notebooks, and the committed processed outputs (per-cell CSV tables and per-FOV spectra) live in the companion GitHub repository: https://github.com/Arcadia-Science/chlorella-crs-phenotyping. Content Two archives, each holding raw, instrument-native image files. Only raw data are deposited here; processed/derived tables are version-controlled in the GitHub repository. Archive Contents Format fluorescence.zip Spinning-disk confocal fluorescence acquisitions (Nikon CSU-W1) Nikon ND2 raman.zip Hyperspectral coherent Raman scattering acquisitions (Leica STELLARIS 8 CRS) Leica LIF After downloading, unzip both archives into the data/ directory of the GitHub repository so the raw files sit alongside the committed processed/ folders. The intended layout is: data/ ├── fluorescence/ │ └── 2026-05-12/ │ ├── processed/*.csv│ └── Well<XX>_Point<XX>_<FOV>_ChannelDIC,FITC BP,Cy5 BP_Seq<NNNN>.nd2 └── raman/ ├── 2026-04-13/ │ ├── processed/.../spectra/*.csv │ └── chlorella_<...>.lif ├── 2026-04-27/ │ ├── processed/.../spectra/*.csv │ └── chlorella_<...>.lif └── 2026-05-12/ ├── processed/.../spectra/*.csv └── chlorella_<...>.lif fluorescence.zip — confocal fluorescence (Nikon CSU-W1) Spinning-disk confocal acquisitions. Each ND2 file is a single field of view with three channels: Channel Reports DIC Brightfield / morphology FITC BP BODIPY-stained neutral (storage) lipid Cy5 BP Chlorophyll autofluorescence A single 96-well plate was imaged across rows C and D, columns 1–12, with 25 fields of view per well (_0000–_0024), for 600 ND2 files total (session 2026-05-12/). Strain, staining, and dilution are assigned by plate position: Factor Layout Strain columns 1–3, 7–9 → ADA1 knockout (ADAKO); columns 4–6, 10–12 → wild type (EV_CTL) BODIPY staining columns 1–6 → stained; columns 7–12 → unstained Dilution row C → 1:9; row D → 1:27 So for the imaged rows: ADA1 KO: WellC01–C03, WellC07–C09, WellD01–D03, WellD07–D09 Wild type: WellC04–C06, WellC10–C12, WellD04–D06, WellD10–D12 Stained (columns 1–6) vs unstained (columns 7–12) The full well → strain / staining / dilution map is provided as plate-map.csv in the GitHub repository. raman.zip — coherent Raman scattering (Leica STELLARIS 8 CRS) Hyperspectral CRS acquisitions using the forward stimulated Raman scattering (F-SRS) and epi coherent anti-Stokes Raman scattering (E-CARS) detectors, collected across three independent sessions in April–May 2026. Each LIF is a LAS X Navigator lambda scan: a dual-channel SRS + CARS image stack acquired at successive wavenumber positions, with multiple cells per field of view at subcellular resolution. Two spectral regions were acquired (encoded in series/folder names): High-wavenumber (HWN), 2,800–3,100 cm⁻¹ — Lambda_HWN_3100_2800_* series, 0.2–0.4 nm step size. Fingerprint (FP), ~600–1,800 cm⁻¹ — Lambda_fingerprint_* series, 0.5–1.0 nm step size. (Step size appears in names as e.g. 0pt4 = 0.4 nm; Nav1/Nav2 are LAS X Navigator acquisition indices.) Session Raw files Notes 2026-04-13/ chlorella_slide{1,2}_B1_low_lipid.lif, chlorella_slide{1,2}_B8_high_lipid.lif Pilot session. Filenames encode slide number and lipid condition; B1 (low lipid) corresponds to wild type (EV_CTL). 2026-04-27/ chlorella_slide{1,2}_A1.lif, chlorella_slide{1,2}_C12.lif Filenames encode slide number and well. 2026-05-12/ chlorella_EV_CTL_slide{1,2,3}.lif, chlorella_ADA_mut_slide{1,2,3}.lif Main strain comparison; filenames name the strain explicitly. chlorella_EV_CTL_slide1_after_reboot.lif is a re-acquisition after an instrument reboot. Processing pipelines Confocal ND2 files were segmented and measured with arcadia-microscopy-tools (Cellpose segmentation), producing per-cell measurement tables. CRS LIF files were processed with the companion pipeline crs-spectral-pipeline, producing per-cell SRS spectra. The resulting processed tables are committed to the GitHub repository, so the analysis notebooks can run on the processed data alone; this raw deposition is needed to reproduce the full pipeline end to end and the figure panels built directly from raw images.