Giorgio
e7e4db264d
Reorganized flat 41-file directory into structured layout with: - scripts/ for Python analysis code with shared config.py - notebooks/ for Jupyter analysis notebooks - data/ split into raw/, metadata/, processed/ - docs/ with analysis summary, experimental design, and bimodal hypothesis tutorial - tasks/ with todo checklist and lessons learned - Comprehensive README, PLANNING.md, and .gitignore Co-Authored-By: Claude Opus 4.6 <noreply@anthropic.com>
98 lines
5.1 KiB
Markdown
98 lines
5.1 KiB
Markdown
# Experimental Design: Barrier-Opening Social Interaction Assay
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## Overview
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This document describes the experimental design of a Drosophila behavioral tracking experiment conducted as part of the Cupido project. The experiment uses a barrier-opening assay to measure social interaction patterns between trained and untrained flies.
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**Date**: July 15, 2025
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**Species**: *Drosophila melanogaster*, Canton-S (CS) wild-type strain
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## Assay Description
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The barrier-opening assay places two flies in each Region of Interest (ROI), separated by a physical barrier. After a configurable delay from the start of recording, the barrier is manually opened, allowing the flies to interact socially. The primary behavioral metric is the **distance between the two flies over time** following barrier opening, which serves as a proxy for social engagement.
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## Experimental Groups
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- **Trained**: Flies that received prior social experience before the assay.
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- **Untrained**: Socially naive flies with no prior social experience.
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> **Note**: The exact training protocol (duration, conditions, group sizes) should be documented separately. "Trained" refers to flies with prior social experience; "untrained" refers to socially naive individuals.
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## Equipment and Recording Parameters
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| Parameter | Value |
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|-----------|-------|
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| Resolution | 1920 x 1088 pixels |
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| Frame rate | 25 fps |
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| Video codec | H.264 |
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| Quality | 28q |
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| ROIs per session | 6 (each containing a pair of flies) |
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| Tracking output | SQLite databases (one per session) |
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## Machines and Recording Sessions
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Three ethoscope machines were used for tracking, with a fourth (Machine 139) having metadata but no tracking data.
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| Machine | Session Start Time | Barrier Opening (s) | Status |
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|---------|--------------------|----------------------|--------|
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| ETHOSCOPE_076 | 16:03:10 | 52 | OK |
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| ETHOSCOPE_076 | 16:31:34 | 25 | OK |
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| ETHOSCOPE_145 | 16:03:27 | 42 | OK |
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| ETHOSCOPE_145 | 16:31:41 | 20 | OK |
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| ETHOSCOPE_268 | 16:32:05 | 75 | OK |
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| ETHOSCOPE_139 | 16:31:52 | Not recorded | **DATA MISSING** |
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**Total sessions**: 6 (5 with tracking data, 1 missing)
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## ROI-to-Group Mapping
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Each session contains 6 ROIs. The assignment of trained/untrained groups to ROIs varies across sessions.
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| Machine | Session | ROI 1 | ROI 2 | ROI 3 | ROI 4 | ROI 5 | ROI 6 |
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|---------|---------|-------|-------|-------|-------|-------|-------|
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| 076 | 16:03:10 | Trained | Untrained | Trained | Untrained | Trained | Untrained |
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| 076 | 16:31:34 | Trained | Trained | Trained | Untrained | Untrained | Untrained |
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| 145 | 16:03:27 | Trained | Trained | Trained | Untrained | Untrained | Untrained |
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| 145 | 16:31:41 | Trained | Trained | Trained | Untrained | Untrained | Untrained |
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| 268 | 16:32:05 | Untrained | Untrained | Untrained | Trained | Trained | Trained |
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| 139 | 16:31:52 | Trained | Trained | Trained | Untrained | Untrained | Untrained |
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## Sample Sizes
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| Group | ROIs (total) | ROIs (with data) | ROIs (missing) |
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|-------|-------------|-------------------|----------------|
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| Trained | 18 | 15 | 3 (Machine 139) |
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| Untrained | 18 | 15 | 3 (Machine 139) |
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| **Total** | **36** | **30** | **6** |
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## Tracking Database Schema
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Each recording session produces a SQLite database file containing tables `ROI_1` through `ROI_6`. Each table has the following columns:
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| Column | Type | Description |
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|--------|------|-------------|
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| `id` | INTEGER | Row identifier |
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| `t` | INTEGER | Timestamp in **milliseconds** from start of recording |
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| `x` | REAL | Horizontal position in pixels |
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| `y` | REAL | Vertical position in pixels |
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| `w` | REAL | Width of detected object (pixels) |
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| `h` | REAL | Height of detected object (pixels) |
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| `phi` | REAL | Angle/orientation of detected object |
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| `is_inferred` | INTEGER | Whether the position was inferred (not directly detected) |
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| `has_interacted` | INTEGER | Whether an interaction was detected |
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## Known Issues and Data Caveats
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1. **Machine 139 missing data**: Metadata entries exist for ETHOSCOPE_139 (session 16:31:52) in the metadata CSV, but no corresponding tracking database file is present and no barrier opening time was recorded. This accounts for 6 missing ROIs (3 trained, 3 untrained). The cause needs investigation.
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2. **Time unit mismatch between files**: The tracking databases store time (`t`) in **milliseconds**, while `2025_07_15_barrier_opening.csv` stores barrier opening times in **seconds**. The analysis pipeline converts barrier opening times to milliseconds for alignment.
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3. **Machine name type inconsistency**: The metadata CSV stores machine identifiers as integers (e.g., `76`, `145`, `268`), while `2025_07_15_barrier_opening.csv` also stores them as integers (e.g., `076`, `145`, `268`). String conversion with zero-padding is required when matching between files and when constructing tracking database filenames (e.g., `ETHOSCOPE_076`).
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## Source Files
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| File | Description |
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|------|-------------|
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| `2025_07_15_metadata_fixed.csv` | ROI-to-group mapping (trained/untrained) |
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| `2025_07_15_barrier_opening.csv` | Barrier opening times per machine/session |
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| `*_tracking.db` | SQLite tracking databases (one per session) |
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