IL-10 Recombinant Rabbit Monoclonal Antibody [JE59-71]

☑ Cell treatment (CT)

Western blot analysis of IL-10 on different lysates with Rabbit anti-IL-10 antibody (HA722032) at 1/1,000 dilution.

Lane 1: THP-1 cell lysate (30 µg/Lane)
Lane 2: THP-1 treated with 80nM TPA overnight then treated with 100ng/mL LPS for 6 hours add 300ng/mL BFA for 3 hours cell lysate (30 µg/Lane)
Lane 3: Mouse spleen tissue lysate (40 µg/Lane)
Lane 4: Rat spleen tissue lysate (40 µg/Lane)

Predicted band size: 21 kDa
Observed band size: 21 kDa

Exposure time: 1 minutes 59 seconds;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA722032) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

☑ Cell treatment (CT)

Immunocytochemistry analysis of THP-1 cells treated with 80nM TPA overnight then treated with 100ng/mL LPS for 6 hours add 300ng/mL BFA for 3 hours labeling IL-10 with Rabbit anti-IL-10 antibody (HA722032) at 1/500 dilution.

Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-IL-10 antibody (HA722032) at 1/500 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

Immunohistochemical analysis of paraffin-embedded rat spleen tissue with Rabbit anti-IL-10 antibody (HA722032) at 1/1,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (HA722032) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.