title: GSM4565852: CD14_4; Homo sapiens; RNA-Seq
library_construction_protocol: Tumor-draining lymph nodes (tdLN) and primary tumors were collected in CO2 independent medium (Gibco) within few hours after the primary surgery. Tissue were cut into small fragments and submitted to enzymatic digestion using 0.1 mg/ml of Liberase TL (Roche) and 0.1 mg/ml of DNase (Roche) for 30 min. Cells were filtered on 40-μm cell strainer (BD), washed using CO2 independent medium (Gibco) containing 0.4g/ml of human albumin and resuspended for cell counting. T and B lymphocytes, NK cells, erythrocytes and myelomonocytic cells were depleted using monoclonal antibodies against: CD3, CD19, CD56, CD235a and CD15, respectively. Subsequently, cell suspensions were stained for 30 min with antibody-conjugated as the following: HLA-DR, CD11c, CD14, CD1c, CD304, CD1a, CD206, CD141. Around 1,000 cells of each DC subset were sorted by flow cytometry using BD FACS ARIA II cell sorter, (purity > 98%). Cells were centrifuged and lysed with TCL buffer (Qiagen) containing 1% of beta-mercaptoethanol before storage at -80°C. RNA were extracted and isolated using the Single Cell RNA purification kit (Norgen)  according to the manufacturer's instructions and the RNA integrity number was evaluated with an Agilent RNA 6000 pico kit.
attributes.tag: GEO Accession
attributes.value: GSM4565852