[IF=2.464] Zhou, Taocheng. et al. Leonurine Alleviates Alcoholic Steatohepatitis Through the TLR4/NF-κB Signalling Pathway. Revista Brasileira de Farmacognosia-Brazilian Journal of Pharmacognosy. 2022 Aug;:1-15  WB ;  Mouse.  

PubMed:10.1007/s43450-022-00284-4

Sample:
Lane 1: A431 (Human) Cell Lysate at 30 ug
Lane 2: Hela (Human) Cell Lysate at 30 ug
Primary:
Anti-TNF alpha (bs-41192R) at 1/1000 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 36/17/26 kD
Observed band size: 36 kD

Blank control:Raw264.7.
Primary Antibody (green line): Rabbit Anti-TNF alpha antibody (bs-41192R)
Dilution: 2ug/Test;
Secondary Antibody : Goat anti-rabbit IgG-FITC
Dilution: 0.5ug/Test.
Protocol
The cells were treated with LPS (1 ug/mL, 18 hr/6 hr) and Brefeldin A (300 ng/mL, last 3 hr of stimulation).The cells were fixed with 4% PFA (10min at room temperature)and then permeabilized with 0.1% PBST for 20 min at room temperature.The cells were then incubated in 5%BSA to block non-specific protein-protein interactions for 30 min at room temperature .Cells stained with Primary Antibody for 30 min at room temperature. The secondary antibody used for 40 min at room temperature. Acquisition of 20,000 events was performed.

Blank control:Raw264.7.
Primary Antibody (green line): Rabbit Anti-TNF alpha antibody (bs-41192R)
Dilution: 2ug/Test;
Secondary Antibody : Goat anti-rabbit IgG-FITC
Dilution: 0.5ug/Test.
Protocol
The cells were treated with LPS (1 ug/mL, 18 hr/6 hr) and Brefeldin A (300 ng/mL, last 3 hr of stimulation).The cells were fixed with 4% PFA (10min at room temperature)and then permeabilized with 0.1% PBST for 20 min at room temperature.The cells were then incubated in 5%BSA to block non-specific protein-protein interactions for 30 min at room temperature .Cells stained with Primary Antibody for 30 min at room temperature. The secondary antibody used for 40 min at room temperature. Acquisition of 20,000 events was performed.

Blank control:THP-1.
Primary Antibody (green line): Rabbit Anti-TNF alpha antibody (bs-41192R)
Dilution: 1ug/Test;
Secondary Antibody : Goat anti-rabbit IgG-FITC
Dilution: 0.5ug/Test.
Protocol
The cells were treated with TPA (80 nM, overnight) and then treated with LPS (1 ug/mL, 18 hr/6 hr) and Brefeldin A (300 ng/mL, last 3 hr of stimulation) .The cells were fixed with 4% PFA (10min at room temperature)and then permeabilized with 0.1% PBST for 20 min at room temperature.The cells were then incubated in 5%BSA to block non-specific protein-protein interactions for 30 min at room temperature .Cells stained with Primary Antibody for 30 min at room temperature. The secondary antibody used for 40 min at room temperature. Acquisition of 20,000 events was performed.

Blank control:THP-1.
Primary Antibody (green line): Rabbit Anti-TNF alpha antibody (bs-41192R)
Dilution:2ug/Test;
Secondary Antibody : Goat anti-rabbit IgG-FITC
Dilution: 0.5ug/Test.
Protocol
The cells were treated with TPA (80 nM, overnight) and then treated with LPS (1 ug/mL, 18 hr/6 hr) and Brefeldin A (300 ng/mL, last 3 hr of stimulation) .The cells were fixed with 4% PFA (10min at room temperature)and then permeabilized with 0.1% PBST for 20 min at room temperature.The cells were then incubated in 5%BSA to block non-specific protein-protein interactions for 30 min at room temperature .Cells stained with Primary Antibody for 30 min at room temperature. The secondary antibody used for 40 min at room temperature. Acquisition of 20,000 events was performed.