Dataset features


Application: Gene expression microarray analysis
Number of samples: 18
Release date: Dec 11 2008
Last update date: Oct 29 2018
Access: Public
Diseases: Leukemia, Leukemia, Lymphocytic, Chronic, B-Cell
Dataset link Analysis of gene expression response of CLL cells to co-culture with Nurse like cells

Experimental Protocol

The microarray part of this study included samples of CLL cells from 9 different patients, analyzed directly after purification and after 14 days of co-culture with Nurse like cells . In detail, RNA was isolated from CD19-purified CLL cells from 9 different patients’ peripheral blood mononuclear cells (PBMC) after Ficoll separation and subsequent purification with CD19 MicroBeads and the MACS® technology according to the manufacturer’s instructions (Miltenyi Biotec, Bergisch Gladbach, Germany). For comparison, the same CLL cell samples were co-cultured for 14 days with NLC ("14d NLC")."). For co-culture with NLC, PBMC from patients with CLL were suspended in complete RPMI medium (RPMI1640 with 10% FCS, penicillin-streptomycin-glutamine, Gibco-BRL, Grand Island, NY) to a concentration of 1 x 107/ml (total 20 ml) and incubated for 14 days in 75 cm2 tissue culture flasks (Techno Plastic Products AG) as described previously3. Nonadherent lymphoid cells then were removed and the NLC layer was washed two times with phosphate-buffered saline (PBS). The complete removal of lymphocytes was verified by phase-contrast microscopy. The nonadherent cells together with the wash-fractions were then used for RNA preparation. In order to purify the CLL B cells prior to RNA isolation, CLL PBMC were passed through a 30 µm nylon mesh to obtain a single-cell suspension. Then CLL B cells were purified with CD19 MicroBeads. Subsequently RNA was extracted.










Andreas Rosenwald