BD^® OMICS-One Immune Profiler Protein Panel

Similar performance between the BD^® OMICS-One IP Protein Panel versus freshly mixed BD^® AbSeq Antibodies. Following isolation from whole blood, peripheral blood mononuclear cells (PBMC) were split into resting (untreated) and activated (treated with CD3/CD28 for 24 hours) groups. A 1:1 mixture of the resting and activated cells were stained with either the BD^® OMICS-One IP Protein Panel or a freshly prepared mixture of the same AbSeq specificities. Equal number of cells were then loaded onto the BD Rhapsody^™ Cartridges and AbSeq and WTA libraries were generated and sequenced (n = 2 individual experiments for this study). Data were analyzed using SeqGeq^™ Software.

A. UMAP demonstrated strong overlap in the cell groups identified between the BD^® OMICS-One IP Protein Panel and fresh BD^® AbSeq Antibody-stained samples.

B. The total number of AbSeq molecules detected by the BD^® OMICS-One IP Protein Panel and fresh BD^® AbSeq Antibody mixture showed a high correlation with R² greater than 0.98.

Performance of all 30 AbSeq specificities included in the BD^® OMICS-One IP Protein Panel. PBMCs were activated and prepared as described in the figure above. After staining, cells were captured on the BD Rhapsody^™ System and AbSeq and WTA libraries were generated and sequenced. To obtain over 80% sequencing saturation, the libraries were sequenced at 20,000 reads/cell for WTA and 30,000 reads/cell for AbSeq using the Illumina^™ NextSeq^™ High-Output Kit. Data were analyzed using SeqGeq™ Software. We repeated the above experiments with at least two different donors. The representative figures from one donor are shown here.

A. Cell annotation of UMAP of resting + activated PBMCs resolved by the BD^® OMICS-One IP Protein Panel antibodies and the WTA mRNA profile.

B. Heat maps of each AbSeq clone from BD^® OMICS-One IP Protein Panel on UMAP from (A) showing the specificity of AbSeq detection for individual cell type.

The BD^® OMICS-One IP Protein Panel is a flexible backbone panel and accommodates additional AbSeq specificities. Three BD^® AbSeq Antibodies were added and mixed with the reconstituted BD^® OMICS-One IP Protein Panel pellet (n = 2).

A. The BD^® OMICS-One IP Protein Panel performance was not impacted by drop-ins as shown by high correlation (R² over 0.99 with or without drop-ins).

B. The BD^® OMICS-One IP Protein Panel specificities of CD8 (SK1) and CD45RA (top row) were assessed against the specificity of drop-ins CD8 [RPA-T8], CD45RO and CD38 (bottom row) and are shown in UMAP. Drop-in for CD38 detected cell types that are expected to be positive for CD38. Drop-in clone for CD8 (RPA-T8) showed a staining pattern very similar to the BD^® OMICS-One IP Protein Panel clone (SK1) suggesting the high specificity of drop-in antibody as well as the compatibility of two clones for the same antigen. The contrasting expression pattern of CD45RO (drop-in) compared to CD45RA (BD^® OMICS-One IP Protein Panel) further confirmed that adding the AbSeq specificities to the BD^® OMICS-One IP Protein Panel had no adverse impact on experimental outcomes.

The BD^® OMICS-One IP Protein Panel is designed to work with RNA and multiplexing assays

A. WTA and AbSeq libraries from BD^® OMICS-One IP Protein Panel-stained cells (1:1 mixture of resting and activated PBMCs) were generated and sequenced. To illustrate the power of multiomic analysis, we analyzed the WTA data only (mRNA analysis) and compared with WTA + AbSeq data (mRNA and protein analysis). UMAP coordinates and unbiased clustering (Phenograph) using only WTA (mRNA) data are shown on the left, while coordinates and annotations using WTA + AbSeq (mRNA and protein) data are shown on the right. With a multiomics approach, additional cell types were revealed offering deeper biological insights.

B. To test the compatibility of the BD^® Single-Cell Multiplexing Kit (SMK) and the BD^® OMICS-One IP Protein Panel, we performed cell staining with the SMK and the BD^® OMICS-One IP Protein Panel together and generated WTA, AbSeq and SMK libraries for sequencing. The expression of markers in the BD^® OMICS-One IP Protein Panel was then compared to data generated in the absence of the SMK. These data showed that addition of the SMK does not impact the BD^® OMICS-One IP Protein Panel as demonstrated by high correlation (R² >0.99) between the BD^® OMICS-One IP Protein Panel + WTA versus BD^® OMICS-One IP Protein Panel + WTA + SMK.