Aggregate of Human Whole Leukocytes Isolated from Fixed Whole Blood of a Diseased Donor and a Healthy Donor (Next GEM)

Whole blood samples were obtained from a donor with no reported illness and from a donor diagnosed with multiple myeloma. The blood samples from each donor were fixed overnight at room temperature or for 7 days at 4°C, following the Demonstrated Protocol for Blood Fixation and Cell Isolation for Chromium Fixed RNA Profiling (CG000721). After fixation, whole leukocytes were isolated in accordance with the protocol and were processed in a 4 or 16-sample multiplexed experiment. In this experiment, the cells were hybridized with probe barcodes alongside cells from other isolated leukocyte samples (not described here). Following hybridization, the samples were pooled in equal proportions, washed, and then processed in a single GEM well to generate Fixed RNA Gene Expression libraries targeting 5,000 cells per sample.

Fixed RNA Gene Expression libraries were generated as outlined in the Chromium Fixed RNA Profiling for Multiplexed Samples User Guide (CG000527). The resulting libraries were sequenced on an Illumina NovaSeq 6000 and analyzed using the cellranger multi pipeline. To accurately capture granulocytes/neutrophils in the data, we adjusted the default cell calling algorithm by running cellranger multi with the --force-cells=5000 option, following the tutorial available on the Sofware Support website.

The outputs from each sample were then aggregated and normalized to an equal sequencing depth (approximately 32k read pairs per cell) using the cellranger aggr pipeline, generating a unified set of output files that contain all data from the individual samples.

This dataset is licensed under the Creative Commons Attribution license.