Blocking Assay:
other:
Concentration Assay:Article Title: α-Synuclein aggregates induce mitochondrial damage and trigger innate immunity to drive neuron–microglia communication
Article Snippet: .. For inflammatory modulations, following reagents were used with associated treatment concentration and durations: STING activator – diABZI (SelleckChem – compound 3, S8796; 2.5 μM for 2h towards the end of experiment), STING inhibitor – H-151 (Invivogen InvitroFit TM , inh-h151; 0.5 μg/mL for 16h for TNT assessment experiments, and 24h for α-Syn transfer experiments), NF-κB inhibitor – JSH-23 (Sigma, J4455; 10 μM for 16h for TNT assessment experiments, and 24h for α-Syn transfer experiments), human cGAS inhibitor – G140 (Invivogen InvitroFit TM , inh-g140; 10 μM for 16h), Bcl-2 inhibitor – ABT-737 (Sigma, 197333; 10 μM for 6h, 12h, or 24h) co-treated with pan-caspase inhibitor – Q-VD-OPh (Non-O- methylated, Sigma, 551476, 10 μM), human IL-1α (Sigma, I2778; 3 ng/mL for 24h), human IL-1β (Sigma, H6291; 20 ng/mL for 24h), human TNF-α (Merck Millipore, 654205; 50 ng/mL for 24h), TLR4 inhibitor – TAK-242 (Tocris, 6587; 10 μM for 16h), TLR2 inhibitor – TL2-C29 (Invivogen InvitroFit TM , inh-C29; 100 μM for 16h), lysosomotropic agent LLOMe (Sigma, L7393; 1 mM for 1h towards the end of experiment), mitochondrial electron transport chain uncoupler CCCP (Sigma, C2759; 20 μM for 2h). .. The following dyes were used: MitoTracker Red CMX Ros (Invitrogen, M7512; 500 nM for 30 minutes at 37°C) and TMRM (Invitrogen, T668; 100 nM for 30 minutes at 37°C).
Article Title: α-Synuclein aggregates induce mitochondrial damage and trigger innate immunity to drive neuron-microglia communication.
Article Snippet: .. For inflammatory modulations, following reagents were used with associated treatment concentration and durations: STING activator – diABZI (SelleckChem – compound 3, S8796; 2.5 μM for 2h towards the end of experiment), STING inhibitor – H-151 (Invivogen InvitroFitTM, inhh151; 0.5 μg/mL for 16h for TNT assessment experiments, and 24h for α-Syn transfer experiments), NF-κB inhibitor – JSH-23 (Sigma, J4455; 10 μM for 16h for TNT assessment AR TI CL E IN P RE SS experiments, and 24h for α-Syn transfer experiments), human cGAS inhibitor – G140 (Invivogen InvitroFitTM, inh-g140; 10 μM for 16h), Bcl-2 inhibitor – ABT-737 (Sigma, 197333; 10 μM for 6h, 12h, or 24h) co-treated with pan-caspase inhibitor – Q-VD-OPh (Non-O-methylated, Sigma, 551476, 10 μM), BAX inhibitor – BAI1 (MedChemExpress, HY-103269, 1 μM for 1h followed by a wash and subsequently 100 nM for 16h), human IL-1α (Sigma, I2778; 3 ng/mL for 24h), human IL-1β (Sigma, H6291; 20 ng/mL for 24h), human TNF-α (Merck Millipore, 654205; 50 ng/mL for 24h), TLR4 inhibitor – TAK-242 (Tocris, 6587; 10 μM for 16h), TLR2 inhibitor – TL2-C29 (Invivogen InvitroFitTM, inh-C29; 100 μM for 16h), TLR9 inhibitor – TLR9-IN-1 (MedChemExpress, HY-148045, 10 μM for 16h), RIG1 inhibitor – RIG012 (MedChemExpress, HY-147124, 0.1 μM for 16h), lysosomotropic agent LLOMe (Sigma, L7393; 1 mM for 1h towards the end of experiment), mitochondrial electron transport chain uncoupler CCCP (Sigma, C2759; 20 μM for 2h). .. NPCs at 80-100% confluency were cultured on POLAM-coated (poly-L-ornithine Sigma-aldrich, P4957; laminin, Sigma-Aldrich, L2020) wells in DAn induction medium [DMEM/F12 (Life, 21331-20), 1% N2 (Life, 17502048), 1% Penicillin/Streptomycin (ScienCell, 0503)] supplemented with 200 ng/ml Sonic Hedgehog (PeproTech, 100-45) and 100 ng/ml FGF8 (PeproTech, 100-25) for 6 days.
Methylation:Article Title: α-Synuclein aggregates induce mitochondrial damage and trigger innate immunity to drive neuron–microglia communication
Article Snippet: .. For inflammatory modulations, following reagents were used with associated treatment concentration and durations: STING activator – diABZI (SelleckChem – compound 3, S8796; 2.5 μM for 2h towards the end of experiment), STING inhibitor – H-151 (Invivogen InvitroFit TM , inh-h151; 0.5 μg/mL for 16h for TNT assessment experiments, and 24h for α-Syn transfer experiments), NF-κB inhibitor – JSH-23 (Sigma, J4455; 10 μM for 16h for TNT assessment experiments, and 24h for α-Syn transfer experiments), human cGAS inhibitor – G140 (Invivogen InvitroFit TM , inh-g140; 10 μM for 16h), Bcl-2 inhibitor – ABT-737 (Sigma, 197333; 10 μM for 6h, 12h, or 24h) co-treated with pan-caspase inhibitor – Q-VD-OPh (Non-O- methylated, Sigma, 551476, 10 μM), human IL-1α (Sigma, I2778; 3 ng/mL for 24h), human IL-1β (Sigma, H6291; 20 ng/mL for 24h), human TNF-α (Merck Millipore, 654205; 50 ng/mL for 24h), TLR4 inhibitor – TAK-242 (Tocris, 6587; 10 μM for 16h), TLR2 inhibitor – TL2-C29 (Invivogen InvitroFit TM , inh-C29; 100 μM for 16h), lysosomotropic agent LLOMe (Sigma, L7393; 1 mM for 1h towards the end of experiment), mitochondrial electron transport chain uncoupler CCCP (Sigma, C2759; 20 μM for 2h). .. The following dyes were used: MitoTracker Red CMX Ros (Invitrogen, M7512; 500 nM for 30 minutes at 37°C) and TMRM (Invitrogen, T668; 100 nM for 30 minutes at 37°C).
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