Integration of the Element AVITITM System with BD RhapsodyTM SingleCell Assays creates a powerful solution for the immuno-oncology researcher Introduction Advances in single-cell isolation and barcoding technologies combined with high-quality next-generation sequencing (NGS) offer exceptional opportunities to profile biomolecules at single-cell resolution. The BD Rhapsody™ Single-Cell Analysis System, with its proprietary microwell-based partitioning technology, enables capture and barcoding of thousands of single cells, and the custom library prep protocols enable a multiomic NGS readout with single-cell resolution. Element Biosciences' benchtop sequencing platform, Element AVITITM System, is based on its proprietary Avidity SequencingTM chemistry, which enables a combination of accuracy, low cost and operational efficiency. In this application note, we show compatibility of Element’s novel sequencing system with BD RhapsodyTM Single-Cell Assays. Single-cell capture Assay library prep Data analysis Element AVITI™ BD Pipeline SeqGeq™ Software Figure 1. Element AVITI™ sequencing is seamlessly incorporated into the existing BD RhapsodyTM System workflow workflow To demonstrate platform compatibility, mRNA and surface protein markers were isolated by loading 10,000 and 2,000 AbSeqstained human PBMCs onto a BD RhapsodyTM Cartridge and using the BD RhapsodyTM System and Enhanced Cartridge Reagent Kit (Cat. No. 664887). Following cDNA synthesis, WTA (Cat. No. 633801), BD® AbSeq Immune Discovery Panel (Cat. No. 625970) and Targeted Human Immune Response Panel (Cat. No. 633774, Cat. No. 633750) libraries were made compatible for sequencing on the AVITITM System using the Element Adept™ Compatibility Kit (Cat. No. 830-00003), a simple 75-minute protocol in which libraries are circularized (without the use of PCR). After circularization and cleanup, libraries are quantified by qPCR using included standards and loaded at the appropriate concentration onto the AVITITM System fo

The sequencing results showed compatibility between the two systems and resulted in high-quality, reproducible data from all library combinations tested. Figure 2. Second derivative from the WTA assay shows well-defined cell calling for high and low cell input The Whole Transcriptome Analysis libraries resulted in high detection of cells and cell labels (Table 1), high gene correlation between the high and low cell input samples (Figure 2) and tSNE plots with cell clustering by type. WTA + AbSeq profile and quality metrics Library type intended cell input Putative cell count % Sequencing saturation...

WTA AbSeq 2,000 vs 10,000 PBMCs WTA + + AbSeq 2,000 vs 10,000PBMCs Subset Name Figure 3. A shows a high gene expression (UMI) correlation between the high kmeans_k10 subset-5 3343 kmeans_k10 subset-4 626 and low cell input WTA + AbSeq libraries. tSNE plots from the WTA + AbSeq kmeans_k10 subset-3 1822 kmeans_k10 subset-2 assay in B illustrate the distinct clustering of PBMCs by cell type with no181 batch kmeans_k10 subset-1 373 kmeans_k10 subset 1794 effect between high and low cell input. QualityCells Subset Name Subset Name Targeted Human Immune Response Panel (HIRP) The Targeted assay quality...

Targeted 2,000 10,000 PBMCs Targeted 2,000 vs vs 10,000 PBMCs 1000 900 Subset Name Figure 4. A shows a high gene expression (UMI) correlation between the high and low cell input targeted mRNA HIRP libraries. tSNE plots from targeted libraries in B illustrate the distinct clustering of PBMCs by cell type with no batch effect between high and low cell input. Conclusions The results of this study demonstrate clear compatibility between the two systems, as high-quality results were shown across all combinations of BD RhapsodyTM System libraries tested. This suggests the AVITITM System offers an alternative...