Neuropixels Analysis

# Neuropixels Data Analysis

## Routing Boundary

Use this skill only for Neuropixels, SpikeGLX, Kilosort, Open Ephys, spike sorting, probe/channel maps, or electrophysiology recording workflows tied to these tools. Do not use it for generic neuroscience literature, clinical EEG, fMRI, flow cytometry, or single-cell RNA analysis.

## Overview

Comprehensive toolkit for analyzing Neuropixels high-density neural recordings using current best practices from SpikeInterface, Allen Institute, and International Brain Laboratory (IBL). Supports the full workflow from raw data to publication-ready curated units.

## When to Use This Skill

This skill should be used when:
- Working with Neuropixels recordings (.ap.bin, .lf.bin, .meta files)
- Loading data from SpikeGLX, Open Ephys, or NWB formats
- Preprocessing neural recordings (filtering, CAR, bad channel detection)
- Detecting and correcting motion/drift in recordings
- Running spike sorting (Kilosort4, SpykingCircus2, Mountainsort5)
- Computing quality metrics (SNR, ISI violations, presence ratio)
- Curating units using Allen/IBL criteria
- Creating visualizations of neural data
- Exporting results to Phy or NWB

## Supported Hardware & Formats

| Probe | Electrodes | Channels | Notes |
|-------|-----------|----------|-------|
| Neuropixels 1.0 | 960 | 384 | Requires phase_shift correction |
| Neuropixels 2.0 (single) | 1280 | 384 | Denser geometry |
| Neuropixels 2.0 (4-shank) | 5120 | 384 | Multi-region recording |

| Format | Extension | Reader |
|--------|-----------|--------|
| SpikeGLX | `.ap.bin`, `.lf.bin`, `.meta` | `si.read_spikeglx()` |
| Open Ephys | `.continuous`, `.oebin` | `si.read_openephys()` |
| NWB | `.nwb` | `si.read_nwb()` |

## Quick Start

### Basic Import and Setup

```python
import spikeinterface.full as si
import neuropixels_analysis as npa

# Configure parallel processing
job_kwargs = dict(n_jobs=-1, chunk_duration='1s', progress_bar=True)
```

### Loading Data

```python
# SpikeGLX (most common)
recording = si.read_spikeglx('/path/to/data', stream_id='imec0.ap')

# Open Ephys (common for many labs)
recording = si.read_openephys('/path/to/Record_Node_101/')

# Check available streams
streams, ids = si.get_neo_streams('spikeglx', '/path/to/data')
print(streams)  # ['imec0.ap', 'imec0.lf', 'nidq']

# For testing with subset of data
recording = recording.frame_slice(0, int(60 * recording.get_sampling_frequency()))
```

### Complete Pipeline (One Command)

```python
# Run full analysis pipeline
results = npa.run_pipeline(
    recording,
    output_dir='output/',
    sorter='kilosort4',
    curation_method='allen',
)

# Access results
sorting = results['sorting']
metrics = results['metrics']
labels = results['labels']
```

## Standard Analysis Workflow

### 1. Preprocessing

```python
# Recommended preprocessing chain
rec = si.highpass_filter(recording, freq_min=400)
rec = si.phase_shift(rec)  # Required for Neuropixels 1.0
bad_ids, _ = si.detect_bad_channels(rec)
rec = rec.remove_channels(bad_ids)
rec = si.common_reference(rec, operator='median')

# Or use our wrapper
rec = npa.preprocess(recording)
```

### 2. Check and Correct Drift

```python
# Check for drift (always do this!)
motion_info = npa.estimate_motion(rec, preset='kilosort_like')
npa.plot_drift(rec, motion_info, output='drift_map.png')

# Apply correction if needed
if motion_info['motion'].max() > 10:  # microns
    rec = npa.correct_motion(rec, preset='nonrigid_accurate')
```

### 3. Spike Sorting

```python
# Kilosort4 (recommended, requires GPU)
sorting = si.run_sorter('kilosort4', rec, folder='ks4_output')

# CPU alternatives
sorting = si.run_sorter('tridesclous2', rec, folder='tdc2_output')
sorting = si.run_sorter('spykingcircus2', rec, folder='sc2_output')
sorting = si.run_sorter('mountainsort5', rec, folder='ms5_output')

# Check available sorters
print(si.installed_sorters())
```

### 4. Postprocessing

```python
# Create analyzer and compute all extensions
analyzer = si.create_sorting_analyzer(sorting, rec, sparse=True)

analyzer.compute('random_spikes', max_spikes_per_unit=500)
analyzer.compute('waveforms', ms_before=1.0, ms_after=2.0)
analyzer.compute('templates', operators=['average', 'std'])
analyzer.compute('spike_amplitudes')
analyzer.compute('correlograms', window_ms=50.0, bin_ms=1.0)
analyzer.compute('unit_locations', method='monopolar_triangulation')
analyzer.compute('quality_metrics')

metrics = analyzer.get_extension('quality_metrics').get_data()
```

### 5. Curation

```python
# Allen Institute criteria (conservative)
good_units = metrics.query("""
    presence_ratio > 0.9 and
    isi_violations_ratio < 0.5 and
    amplitude_cutoff < 0.1
""").index.tolist()

# Or use automated curation
labels = npa.curate(metrics, method='allen')  # 'allen', 'ibl', 'strict'
```

### 6. AI-Assisted Curation (For Uncertain Units)

When using this skill with Claude Code, Claude can directly analyze waveform plots and provide expert curation decisions. For programmatic API access:

```python
from anthropic import Anthropic

# Setup API client
client = Anthropic()

# Analyze uncertain units visually
uncertain = metrics.query('snr > 3 and snr < 8').index.tolist()

for unit_id in uncertain:
    result = npa.analyze_unit_visually(analyzer, unit_id, api_client=client)
    print(f"Unit {unit_id}: {result['classification']}")
    print(f"  Reasoning: {result['reasoning'][:100]}...")
```

**Claude Code Integration**: When running within Claude Code, ask Claude to examine waveform/correlogram plots directly - no API setup required.

### 7. Generate Analysis Report

```python
# Generate comprehensive HTML report with visualizations
report_dir = npa.generate_analysis_report(results, 'output/')
# Opens report.html with summary stats, figures, and unit table

# Print formatted summary to console
npa.print_analysis_summary(results)
```

### 8. Export Results

```python
# Export to Phy for manual review
si.export_to_phy(analyzer, output_folder='phy_export/',
                 compute_pc_features=True, compute_amplitudes=True)

# Export to NWB
from spikeinterface.exporters import export_to_nwb
export_to_nwb(rec, sorting, 'output.nwb')

# Save quality metrics
metrics.to_csv('quality_metrics.csv')
```

## Common Pitfalls and Best Practices

1. **Always check drift** before spike sorting - drift > 10μm significantly impacts quality
2. **Use phase_shift** for Neuropixels 1.0 probes (not needed for 2.0)
3. **Save preprocessed data** to avoid recomputing - use `rec.save(folder='preprocessed/')`
4. **Use GPU** for Kilosort4 - it's 10-50x faster than CPU alternatives
5. **Review uncertain units manually** - automated curation is a starting point
6. **Combine metrics with AI** - use metrics for clear cases, AI for borderline units
7. **Document your thresholds** - different analyses may need different criteria
8. **Export to Phy** for critical experiments - human oversight is valuable

## Key Parameters to Adjust

### Preprocessing
- `freq_min`: Highpass cutoff (300-400 Hz typical)
- `detect_threshold`: Bad channel detection sensitivity

### Motion Correction
- `preset`: 'kilosort_like' (fast) or 'nonrigid_accurate' (better for severe drift)

### Spike Sorting (Kilosort4)
- `batch_size`: Samples per batch (30000 default)
- `nblocks`: Number of drift blocks (increase for long recordings)
- `Th_learned`: Detection threshold (lower = more spikes)

### Quality Metrics
- `snr_threshold`: Signal-to-noise cutoff (3-5 typical)
- `isi_violations_ratio`: Refractory violations (0.01-0.5)
- `presence_ratio`: Recording coverage (0.5-0.95)

## Bundled Resources

### scripts/preprocess_recording.py
Automated preprocessing script:
```bash
python scripts/preprocess_recording.py /path/to/data --output preprocessed/
```

### scripts/run_sorting.py
Run spike sorting:
```bash
python scripts/run_sorting.py preprocessed/ --sorter kilosort4 --output sorting/
```

### scripts/compute_metrics.py
Compute quality metrics and apply curation:
```bash
python scripts/compute_metrics.py sorting/ preprocessed/ --output metrics/ --curation allen
```

### scripts/export_to_phy.py
Export to Phy for manual curation:
```bash
python scripts/export_to_phy.py metrics/analyzer --output phy_export/
```

### assets/analysis_template.py
Complete analysis template. Copy and customize:
```bash
cp assets/analysis_template.py my_analysis.py
# Edit parameters and run
python my_analysis.py
```

### reference/standard_workflow.md
Detailed step-by-step workflow with explanations for each stage.

### reference/api_reference.md
Quick function reference organized by module.

### reference/plotting_guide.md
Comprehensive visualization guide for publication-quality figures.

## Detailed Reference Guides

| Topic | Reference |
|-------|-----------|
| Full workflow | [references/standard_workflow.md](reference/standard_workflow.md) |
| API reference | [references/api_reference.md](reference/api_reference.md) |
| Plotting guide | [references/plotting_guide.md](reference/plotting_guide.md) |
| Preprocessing | [references/PREPROCESSING.md](reference/PREPROCESSING.md) |
| Spike sorting | [references/SPIKE_SORTING.md](reference/SPIKE_SORTING.md) |
| Motion correction | [references/MOTION_CORRECTION.md](reference/MOTION_CORRECTION.md) |
| Quality metrics | [references/QUALITY_METRICS.md](reference/QUALITY_METRICS.md) |
| Automated curation | [references/AUTOMATED_CURATION.md](reference/AUTOMATED_CURATION.md) |
| AI-assisted curation | [references/AI_CURATION.md](reference/AI_CURATION.md) |
| Waveform analysis | [references/ANALYSIS.md](reference/ANALYSIS.md) |

## Installation

```bash
# Core packages
pip install spikeinterface[full] probeinterface neo

# Spike sorters
pip install kilosort          # Kilosort4 (GPU required)
pip install spykingcircus     # SpykingCircus2 (CPU)
pip install mountainsort5     # Mountainsort5 (CPU)

# Our toolkit
pip install neuropixels-analysis

# Optional: AI curation
pip install anthropic

# Optional: IBL tools
pip install ibl-neuropixel ibllib
```

## Project Structure

```
project/
├── raw_data/
│   └── recording_g0/
│       └── recording_g0_imec0/
│           ├── recording_g0_t0.imec0.ap.bin
│           └── recording_g0_t0.imec0.ap.meta
├── preprocessed/           # Saved preprocessed recording
├── motion/                 # Motion estimation results
├── sorting_output/         # Spike sorter output
├── analyzer/               # SortingAnalyzer (waveforms, metrics)
├── phy_export/             # For manual curation
├── ai_curation/            # AI analysis reports
└── results/
    ├── quality_metrics.csv
    ├── curation_labels.json
    └── output.nwb
```

## Additional Resources

- **SpikeInterface Docs**: https://spikeinterface.readthedocs.io/
- **Neuropixels Tutorial**: https://spikeinterface.readthedocs.io/en/stable/how_to/analyze_neuropixels.html
- **Kilosort4 GitHub**: https://github.com/MouseLand/Kilosort
- **IBL Neuropixel Tools**: https://github.com/int-brain-lab/ibl-neuropixel
- **Allen Institute ecephys**: https://github.com/AllenInstitute/ecephys_spike_sorting
- **Bombcell (Automated QC)**: https://github.com/Julie-Fabre/bombcell
- **SpikeAgent (AI Curation)**: https://github.com/SpikeAgent/SpikeAgent