Fluent BioSciences PIPseq workflow
1. Sample Preparation
- During sample preparation, cell suspension of interest is mixed with our core template particles and segregated into Particle-Templated Instant Partitions (PIPs) by simple vortexing
- The cells in PIPs are then lysed on a thermal device and the mRNA is captured by barcoded oligonucleotides incorporated with the template particles
2. Library Preparation
- The captured mRNA undergoes reverse transcription and cDNA libraries are then generated via fragmentation, end repair and A-tailing
- This is then followed by adapter ligation and cleanup
- Sample indices (compatible with Illumina sequencing) are then added, followed by a final cleanup
3. Next generation sequencing (NGS)
- Libraries can then be processed on the appropriate Illumina NGS instrument.
- The sequencing reagent kit (and chip) is chosen based on the number of samples multiplexed, cell count and desired read depth
4. Data Analysis
- The Fluent PIPseekerTM Software enables primary analysis of the PIPseq sequencing libraries
- Upload your FASTQ files into the software and obtain summary metrics, diagnostic plots, clustering and differential gene expression tables
- Also generate standard feature-barcode count matrices, which are compatible with widely-used open-source tertiary analysis packages
Select from the below resources to learn more about our protocols (user guides) or customer applications (posters).