Unified Olympus OIR Batch Processing Macro (Fiji)

Overview

Fiji / ImageJ macros for automated batch processing of Olympus OIR microscopy files. Handles channel splitting, Z-stack projection, and composite image generation across nested directory structures with no manual interaction.

Repository Structure

.
├── unified fiji.ijm              # Original macro (with upscaling)
├── unified fiji_no_scale.ijm     # Revised macro (native resolution, quality-fixed)
├── README.md
├── input_data/                   # Raw .oir files (user-provided)
└── output_data/                  # Auto-generated outputs
    ├── Individual_Channels/
    └── Composite_Images/

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Macros

unified fiji.ijm

The original processing macro. Opens each .oir file, detects whether it is a Z-stack or channel-only image, splits channels, generates a composite, and saves TIFFs. Applies a 10× upscale before export and uses a hardcoded inch-based scale.

unified fiji_no_scale.ijm (recommended)

Revised version with the following changes over the original:

• No upscaling — exports at native camera resolution; removes the 10× Bilinear scale that caused pixelation
• Contrast normalization — applies Enhance Contrast per channel before 16→8-bit RGB conversion to preserve dynamic range
• LUT-preserving channel export — duplicates each channel individually from the composite hyperstack so fluorescence colors are retained in saved TIFFs
• Correct composite rendering — uses Flatten instead of Stack to RGB, which renders the composite exactly as displayed including LUT assignments
• Calibration preserved — removes the hardcoded Set Scale override; Bio-Formats reads the correct µm/pixel calibration from OIR metadata automatically

Both macros mirror the input subfolder structure in the output directories and support arbitrarily nested input folders.

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Requirements

• Fiji (ImageJ ≥ 1.53 recommended)
• Bio-Formats plugin — included in standard Fiji; handles Olympus OIR import

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How to Run

1. Launch Fiji
2. Go to Plugins → Macros → Edit… and open the .ijm file
3. Click Run and choose input/output directories when prompted
4. Monitor progress in the Log window

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Output

Each run produces:

• Individual_Channels/ — one TIFF per fluorescence channel per file, with LUT color applied
• Composite_Images/ — merged composite TIFF with the transmission/brightfield channel excluded

Output folder structure mirrors the input directory tree.