Microchamber analysis

This repo contains code for quantifying and comparing the movement of Chlamydomonas reinhardtii in 384 well plates and custom-developed microchambers. These analyses are included in the publication Microchamber slide design for cell confinement during imaging.

Repository contents

• code/FIJI/ — ImageJ/Fiji macros for pre-processing raw timelapse movies (see code/FIJI/README.md).
• code/CellProfiler/ — CellProfiler pipelines that segment/track cells and export per-cell measurements (see code/CellProfiler/README.md).
• code/R/ — R notebook that analyzes the exported measurements and produces the figures (see code/R/README.md).

The full image-to-figure workflow is: Fiji macro (raw .nd2 movies → processed frames) → CellProfiler pipeline (chlamy_swim_track.cppipe, tracks cells and exports measurements_Cells.csv) → R notebook (statistics and figures). The exported measurements_Cells.csv.gz is also archived on Zenodo, so the R analysis can be reproduced without rerunning the image-processing steps.

Reproducing the R analysis

The R analysis lives in code/R/analyzing_chlamydomonas_movement_384_vs_microchambers.ipynb and can be run on its own from the published data.

1. Install R 4.x and Jupyter with the IRkernel so the notebook can run R cells.
2. Install the only R package dependency, vioplot (developed/tested with vioplot v0.3.7):

install.packages("vioplot")

3. Download the input data, measurements_Cells.csv.gz, from Zenodo (DOI: 10.5281/zenodo.6999004). See code/R/README.md for the exact curl command.
4. Open the notebook in Jupyter and update the data path cell to point at the downloaded CSV, then run all cells.

Associated timelapse image data

The following Zenodo repositories contain image data associated with the "Microchamber slide design for cell confinement during imaging" pub and the project to dissect mechanisms of environmental adaptation in protists.

Colpoda steinii cyst rotation.

Colpoda steinii early encystment.

Colpoda steinii in 384 well or microwells.

Colpoda steinii reanimation experiment.

Tetrahymena rostrata swimming.

Versions and platforms

Fiji macro was used with ImageJ2 Version: 2.3.0/1.53q Build: d544a3f481. Cellprofiler pipeline was run on CellProfiler version 5.

Computation was performed on an image analysis workstation with the following specifications:

Operating system: Windows 10 Pro Version 21H2 OS Build 19044.1889

Component	Specification
Platform	Intel Xeon C621E EATX
Motherboard	Supermicro X12SPA-TF 64L
CPU	Intel Xeon W-3365 2.7GHz Thirty-Two Core 48MB 270W
RAM	8 x DDR4-3200 64GB ECC Reg.
Video Card	NVIDIA RTX A6000 48GB PCI-E
Primary Hard Drive	Samsung 980 Pro 2TB Gen4 M.2 SSD
Secondary Hard Drive	Samsung 870 EVO 4TB SATA3 2.5inch SSD
Tertiary Hard Drive	Western Digital Ultrastar 18TB SATA3
Case	Fractal Design Define 7 XL
Power Supply	Super Flower LEADEX Platinum 850W
Additional Cooling	Case Fans Upgrade Kit (PWM Ramping)