• Contains materials, documentation, and slides used in the 2023/2024 Bioimage Analysis Course for NanoCell Interactions Lab.

• This course is recorded in Portuguese, and the materials are available in English.

• All materials were prepared and presented by fefossa.

Our aim is to provide a detailed material on how to plan, organize, and perform a bioimage analysis protocol using CellProfiler [1] for segmentation and feature extraction, pycytominer [2] for profiling, and Python for data visualization and interpretation.

For each day of the course, we will provide the written documentation, slides, and recording (in Portuguese).

ATTENTION: Videos are in Portuguese (PT). Slides and most materials in English (EN).

Most notebooks and scripts used for this course is available at scripts_notebooks_fossa repo. To learn how to use this repository properly for your analysis, watch this tutorial on how to create a submodule of a GitHub repository in PT.

1. Folders and file naming for bioimage analysis

   1.1 Practicing: shell and bash basic commands

2. Using Cellpose to train your own model

   2.1 Installing and using Cellpose human-on-a-loop Video in PT

   2.2 Using RunCellpose (Cellpose + CellProfiler integration). Video in EN

3. Installing CellProfiler with RunCellpose

   Install CellProfiler+RunCellpose on Windows in PT

4. Generate Metadata

   4.1 Presentation about metadata in bioimage analysis in PT here and Q&A

   4.2 Using the Load Data Generator and Layout to CSV apps demonstration and Q&A

   4.3 Using CellProfiler LoadData module here

5. Assay development and Analysis with CellProfiler

   5.1 Assaydev pipeline

   5.2 Analysis pipeline

   • Practicing: CellProfiler

6. Profiling using Pycytominer

   6.1 What's profiling?

   6.2 Profiling evaluation using mean Average Precision

   6.3 Practicing: Create environment & Jupyter notebooks

7. Batch correction with PyCombat

   Batch effect and plate layout design in PT

8. Visualizing and interpreting profiling data

   Morpheus part 1 in EN and Morpheus part 2 in EN

   Morpheus part 1 in PT and Morpheus part 2 in PT

   Visualize single-cells notebook https://youtu.be/55uu1YcmtH0

9. Machine learning

   Videos explaining the ideas and mathematics behind machine learning are freely provided by StatQuest in EN

   A basic explanation of Machine Learning in PT is also available.

   Notebooks for feature importance using Random Forest, Linear Regression, and T-test

EXTRAS

• Using CellProfiler Analyst for Quality Control analysis: see video in PT

[1] Carpenter, A.E., Jones, T.R., Lamprecht, M.R. et al. CellProfiler: image analysis software for identifying and quantifying cell phenotypes. Genome Biol 7, R100 (2006). https://doi.org/10.1186/gb-2006-7-10-r100

[2] Way, G., Chandrasekaran, S. N., Bornholdt, M., Fleming, S., Tsang, H., Adeboye, A., Cimini, B., Weisbart, E., Ryder, P., Stirling, D., Jamali, N., Carpenter, A., & Singh, S. Pycytominer: Python package for processing image-based profiling data (Version 0.3.0) [Computer software]. https://github.com/cytomining/pycytominer

[3] PyCombat