GSM5833305: PBMC-03-1-ADT-S3; Homo sapiens; RNA-Seq
Libraries of antibody-derived tags (ADT) from feature barcoding antibodies were prepared by repeating size purification on the supernatant obtained from the prior size purification of gene expression cDNA libraries (Step 2.3.d in the manufacturer's instructions https://assets.ctfassets.net/an68im79xiti/1eX2FPdpeCgnCJtw4fj9Hx/7cb84edaa9eca04b607f9193162994de/CG000204_ChromiumNextGEMSingleCell3_v3.1_Rev_D.pdf), using a 7:8 volumetric ratio of 2.0X SPRIselect reagent (Beckman Coulter, Cat# B23317) to sample. Indexing amplification was performed using Kapa Hifi HotStart ReadyMix (Kapa Biosystems, Cat# KK2601) and TruSeq Small RNA RPI primers (Illumina) with the following thermocycling conditions: (1) 98 °C, 2 min; (2) 15 × (98 °C, 20 sec; 60 °C, 30 sec; 72 °C, 20 sec); (3) 72 °C, 5 min. Size purification was then repeated on amplified libraries using a 5:6 volumetric ratio of 1.2X SPRIselect reagent to sample. Peripheral blood was collected from each subject in Vacutainer ACD tubes. PBMCs were isolated using a standard Ficoll method and stored in liquid nitrogen.