Sample 36
Homo sapiens
Protocols: Viably frozen bone marrow aspirates were thawed in DMEM + 10% FCS according to the 10X Genomics protocol 'Fresh Frozen Human Peripheral Blood Mononuclear Cells for Single Cell RNA sequencing'. Only samples containing >200x10E6 MNCs were included. Fc receptors were blocked in 10% normal human AB serum (Sigma-Aldrich). To enrich for stromal fibroblasts, samples were incubated with biotinylated antibodies against CD45 (clone HI30, BioLegend), CD235a (clone HIR2, BioLegend) and CD38 (from human CD38 MicroBead Kit, Miltenyi Biotec) followed by depletion using magnetic anti-biotin beads (Miltenyi Biotec) and the iMag (BD Biosciences). After depletion, cells were stained for sorting in PBS containing 2% FCS at 4°C with the following antibodies: CD271-PE (ME20.4), CD235a-PECy7 (HI264), CD31-APCCy7 (WM59), CD44-BV711 (IM7), streptavidin-BV421 (all BioLegend), CD34-PECF610 (4H11, eBioscience), CD45-APC (2D1, eBioscience), CD38-FITC (MHCD3801, Life Technologies), CD71-Alexa Fluor 700 (MEM-75, ExBio) and CD105-BV510 (266, BD Biosciences). 7AAD (Beckman Coulter) or DAPI (Life Technologies) were used for dead cell exclusion. Cells were sorted in DMEM + 10% FCS using a FACSAria III (BD Biosciences). Prior to depletion, 10x106 cells were used to sort CD45+CD38- and CD38+ hematopoietic populations using the same antibody panel. Sorted CD45- cells were combined with sorted CD45+CD38- cells and processed as one sample. Single cells were encapsulated for cDNA synthesis and barcoded using the Chromium Single Cell 3' Reagent Kit v3 (10X Genomics) Libraries were constructed according to the manufacturer's recommendations (10X Genomics)
ENA first public
2020-11-19
ENA last update
2020-11-18
External Id
SAMEA7587646
INSDC center alias
Dept. of Hematology Erasmus MC
INSDC center name
Dept. of Hematology Erasmus MC
INSDC first public
2020-11-19T04:05:23Z
INSDC last update
2020-11-18T19:54:39Z
INSDC status
public
Submitter Id
E-MTAB-91391605724980:Sample 36
age
75
broker name
ArrayExpress
cell type
bone marrow cell
clinical history
Coronary disease
common name
human
developmental stage
adult
disease
normal
immunophenotype
Non-hematopoietic cells
individual
CBM7
inferred cell type
bone marrow microenvironment
organism part
bone marrow
sample name
E-MTAB-91391605724980:Sample 36
sex
male