il10 blocking antibody  (R&D Systems)

Journal: Cellular immunology

Article Title: IL10 restrains autoreactive B cells in transgenic mice expressing inactive RAG1

doi: 10.1016/j.cellimm.2018.06.004

Figure Lengend Snippet: (A). Splenocytes from WT or dnRAG1 mice were cultured for 72h with or without LPS (10 μg/mL) in the absence or presence of the following: recombinant IL10 (rIL10), IL10 blocking antibody (α-IL10 Ab) or isotype control (Iso Ab), or ruxolitinib (Rux), SH-4–54, triamcinolone acetonide (TA) or vehicle (DMSO), and plasma cell subsets were analyzed by flow cytometry as in Fig. 2B. Summary data for n=3 animals/genotype are presented in bar graph format and represented as mean +/− SEM. Statistically significant differences using the Tukey correction are indicated (a, vs LPS; b, vs LPS+Iso Ab; c, vs LPS+DMSO). (B) Concentrations of IgM and IgG in supernatants from splenocyte cultures shown in panel A were measured by ELISA.

Article Snippet: Cell culture Single-cell suspensions from spleen were adjusted to 4.0×10 6 cells/mL in sterile RF10M and 2.0×10 5 cells dispensed into wells of a sterile 96-well flat bottom plate (Corning #353072) The cells were mixed with an equal volume of RF10M lacking or containing the following reagents (source and final concentration shown in parentheses): LPS (Sigma, L2880, 10 μg/mL), IL10 blocking antibody (R&D Systems, MAB417, 10 ng/mL), Rat IgG1 control for IL10 blocking antibody (R&D Systems, MAB005, 10 ng/mL in PBS), recombinant mouse IL10 (Biolegend, 575802, 100 ng/mL), ruxolitinib (Selleckchem, S1378, 10 nM), SH-4–54 (Selleckchem, S7337, 10 nM), or triamcinolone acetonide (Alfa Aesar, J6354803, 10 nM).

Techniques: Cell Culture, Recombinant, Blocking Assay, Control, Flow Cytometry, Enzyme-linked Immunosorbent Assay