ICCS

ICCS quantifies nanoscale spatial organization and interactions of functional nuclear sites by applying super-resolved image cross-correlation spectroscopy to dual-color stimulated emission depletion (STED) nanoscopy data at 20–250 nm resolution.

Key Features:

• Non-Presegmentation Analysis: Analyzes multicolor images directly without pre-segmentation, avoiding object-based segmentation steps.
• Dynamic Interaction Detection: Identifies and quantifies dynamic interactions and spatiotemporal coordination between functional nuclear sites.
• Nanoscale Resolution (STED-ICCS): Integrates dual-color STED nanoscopy with ICCS to measure colocalized fractions and characteristic distances at 20–250 nm resolution.
• Validation through Experimental Data: Applied to MCF10A cells, showing lack of correlation between transcription foci and H3K9me3 and strong proximity between nascent DNA replication foci and PCNA.
• Spatial Segregation Insights: Resolves spatial segregation of nuclear functions, exemplified by a measured 130 nm distance between transcription and replication foci.

Scientific Applications:

• Nuclear architecture and genome maintenance: Quantifies spatial relationships among nuclear components to study genome maintenance mechanisms.
• Transcription regulation: Measures colocalization and distances involving transcription foci to inform studies of transcriptional regulation.
• DNA replication: Assesses proximity of nascent DNA replication foci and PCNA at nanometer scales to study replication dynamics.
• Chromatin organization: Evaluates spatial segregation of chromatin marks such as H3K9me3 relative to functional nuclear sites.

Methodology:

Combines dual-color STED nanoscopy with image cross-correlation spectroscopy to analyze multicolor images without pre-segmentation and to calculate colocalized fractions and characteristic distances.