Enrichment of viable cells from patient samples with LeviCell technology as input into the ResolveOMETM single cell multiomic workflow

Alison Rojas, Swetha D. Velivela, Jessica N. Borja, Jesse A. Ramirez, Tatiana V. Morozova, Jeff G. Blackinton, Jon S. Zawistowski

BioSkryb Genomics, Durham, NC, USA

Single cell multiomic analysis is at the forefront of studies driving insight into tumor clonal evolution and somatic mosaicism in normal tissues, as well as into edited cell line surveillance. The ability to generate unified genomic and transcriptomic information with fidelity is wholly dependent on the input of viable cells into the amplification chemistry, otherwise concurrent genotype and phenotype are not accurately captured. We present the coupling of the Levitas LeviCell 1.0 instrument, where viable vs dying/dead cells differentially levitate in a microfluidic cartridge within a magnetic field, to single cell dispensing technologies providing input into ResolveOME multiomic amplification technology. Primary cryopreserved bronchoalveolar lavage cells were either directly inputted into FACS or HP D100 instruments for dispensing into 96 well PCR plates or inputted into the LeviCell 1.0 prior to dispensing. For FACS dispensing, even with the use of Calcein-AM and propidium iodide dual viability staining, levitation of primary cells (2 independent samples, 2 replicates each) prior to dispensing increased the percentage of cells yielding genomic amplification product from 51.25% to 88.75% and, for transcriptomic yield, from 52.50% to 97.50%. A similar trend of reduced reaction yield dropouts was obtained for HP D100 dispensing for both genomic amplification and cDNA yield, albeit with a more modest mean improvement of 17%--yet LeviCell viability enrichment importantly reduced the amount of sporadic clogging of the HP D100 microfluidic cartridge. Low-coverage genomic sequencing metrics assessing library complexity and amplification uniformity were overall comparable between pre- and post-levitation, as were the transcriptomic sequencing metrics of mitochondrial transcript percentage, exonic:intergenic read ratio, and number of expressed genes detected. The marked reduction of reaction failures or sub-optimal reaction yields while maintaining robust sequencing metrics with this LeviCell-dispensing-ResolveOME workflow is a facile solution for maximizing the utility of rare and valuable samples, as well as for maximizing the success rate of a given ResolveOME run with often challenging clinical samples.