PRISM Wet-Lab Protocol¶
Safety First
Always wear appropriate PPE when handling reagents, especially PFA and formamide.
Shared Protocol¶
The PRISM wet-lab workflow follows the standard RCA protocol for the shared preprocessing steps (fixation through post-amplification). See the full bench-ready SOPs:
- RCA (Fresh Frozen) — Standard protocol for fresh frozen tissue (v5.3)
- RCA (FFPE) — For FFPE tissue samples (v1.0)
PRISM-Specific Steps¶
After the shared RCA protocol, PRISM uses single-round fluorescence imaging with radius vector encoding:
- Fluorescent Probe Hybridization: Hybridize detection probes carrying unique fluorescent barcodes to the RCA products.
- Single-Round Imaging: Capture all barcode signals in a single imaging round across multiple fluorescence channels.
- Decoding: Use radius vector encoding to decode gene identity from the fluorescent barcode pattern.
Related Resources¶
- Bill of Materials — Reagent catalog numbers and vendors
- RCA Mechanism — Conceptual overview of Rolling Circle Amplification
- Imaging Guide — Microscope configuration and acquisition settings
- PRISM Analysis Pipeline — Bioinformatics SOP for downstream analysis