Cd19 magnetic microbeads kit

This study was approved by the Medical Ethical Committee of Shandong Provincial Hospital affiliated to Shandong University. The paraffin-embedded archived samples from 50 newly diagnosed DLBCL patients and 20 reactive hyperplasia patients were collected. Samples of patients with reactive hyperplasia were referred as control. Histological diagnoses were established according to the WHO classification [26 ]. Peripheral blood mononuclear cells (PBMCs) from the whole blood of healthy donors were isolated using Ficoll-Hypaque density gradient centrifugation (TBD science, Tianjin, China). Normal peripheral blood CD19+ B cells were purified from freshly isolated PBMCs using CD19+ magnetic microbeads kit (Miltenyi Biotec, Bergisch Gladbach, Germany). Cells were incubated with beads for 15 min at 4 °C while rotating. Purified CD19+ B cells were selected according to the manufacturer’s protocols. The purity of isolated populations was assessed by FACS analysis, and cells with >90% purity were collected. All samples were obtained with informed consent in accordance with the Declaration of Helsinki.

Blood samples of 96 de novo patients of CLL were collected at the Department of Hematology in Shandong Provincial Hospital after informed consent as approved by the ethics commission. The leukemic clone proportion in peripheral blood of the enrolled CLL patients was > 90%, which could be represented by peripheral blood mononuclear cells [19 (link)]. All the participants were aged between 29 and 85 years (mean age = 60.29 years, s. d. = 11.45). Peripheral blood mononuclear cells (PBMCs) were isolated by Ficoll density gradient centrifugation, which was described in previous study [20 (link), 21 (link)]. CD19 + B cells from healthy donors were purified using CD19 + magnetic microbeads kit (Miltenyi Biotec, Bergisch Gladbach, Germany). Clinical characteristics of CLL patients, including age, stage, immunoglobulin mutational status, and cytogenetics (FISH) were recorded.

Lymph node samples from 60 de novo DLBCL patients and 30 reactive lymphoid hyperplasia patients were collected. Histological diagnoses were established according to the World Health Organization (WHO) classification [19 ]. Normal peripheral blood mononuclear cells (PBMCs) from healthy volunteers were isolated by the Ficoll-Hypaque density gradient centrifugation method (TBD Science, Tianjin, China). Normal B cells were purified from freshly isolated PBMCs using CD19+ magnetic microbeads kit (Miltenyi Biotec, Bergisch Gladbach, Germany) as previously reported [20 (link), 21 (link)]. This study was approved by the Medical Ethical Committee of Shandong Provincial Hospital Affiliated to Shandong University. All samples were obtained with informed consent in accordance with the Declaration of Helsinki.