Anti cd34 pe

For newly sequenced samples with low tumor purity (below 60%), the leukemic cell population was enriched either by flow cytometric sorting or T cell depletion by magnetic beads (EasySep Human CD3 Positive Selection Kit II, 17851, StemCell Technologies). For flow cytometric sorting, CD45^dimCD33^dim positive population was sorted using anti-CD45 PerCP-Cyanine5.5 (eBioscience cat# 8045–9459-120) and anti-CD33 APC (eBioscience cat# 17–0338-42). CD34 gating using anti-CD34 PE (Beckman cat# IM1459U) was added depending on the positivity of each patient sample. Enrichment of the tumor population was confirmed flow cytometric analysis of the post-sorting samples (generally > 90%). Libraries were constructed using the TruSeq Stranded Total RNA Kit, with Ribozero Gold (20020598, Illumina) for RNA-Seq, the TruSeq DNA PCR-Free Library Prep Kit (20015963, Illumina) for WGS, and the TruSeq Exome Kit v1 (20020614, Illumina) for WES according to the manufacturer’s instructions. After library quality and quantity assessment, samples were sequenced on HiSeq2000 or 2500 (Illumina, RRID:SCR_020132, RRID:SCR_016383) instruments with paired-end (2 × 101 bp, 2 × 126 bp, or 2 × 151 bp) sequencing using TruSeq SBS Kit v3-HS (FC-401–3001, Illumina) or TruSeq Rapid SBS Kit (FC-402–4023, Illumina).

Flow cytometric analysis was performed to confirm the MSCs surface markers. Two cell lines of hWJ-MSCs were performed. The adherent MSCs cells were treated with trypsin-EDTA and collected after being centrifuged at 350× g for 5 min. MSCs were incubated in antibodies in the dark for 20 min. All antibodies are listed in Table S1. Each antibody was diluted in 100 mL of PBS(-)—the following antibodies: anti-CD34-PE (dilution 1:10, Beckman Coulter, Brea, CA, USA), anti-CD45-FITC (dilution 1:20, Biolegend, San Diego, CA, USA), anti-CD73-APC, anti-CD90-APC/A750, and anti-CD105-PE (dilution 1:100, Biolegend). The samples were analyzed by an Attune™ NxT Flow Cytometer (Attune™ NXT, Thermo Fisher Scientific, Cleveland, OH, USA). The percentage of CD34-, CD45-, CD73+, CD90+, and CD105+ positive or negative cell populations were calculated using the FCS Express™ Software.