Anti-Tight junction protein ZO-2 TJP2 Antibody

Figure 1. Western blot analysis of TJP2 using anti-TJP2 antibody (PA1957).
Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 30 ug of sample under reducing conditions.
Lane 1: human RT4 whole cell lysates,
Lane 2: human A431 whole cell lysates,
Lane 3: human HepG2 whole cell lysates,
Lane 4: human Hacat whole cell lysates,
Lane 5: human MCF-7 whole cell lysates,
Lane 6: human U20S whole cell lysates,
Lane 7: human Hela whole cell lysates,
Lane 8: human PC-3 whole cell lysates,
Lane 9: rat liver tissue lysates,
Lane 10: rat RH35 whole cell lysates,
Lane 11: mouse liver tissue lysates,
Lane 12: mouse HEPA1-6 whole cell lysates.
After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% non-fat milk/TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-TJP2 antigen affinity purified polyclonal antibody (Catalog # PA1957) at 0.5 μg/mL overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:5000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for TJP2 at approximately 150 kDa. The expected band size for TJP2 is at 134 kDa.

Figure 2. IHC analysis of TJP2 using anti-TJP2 antibody (PA1957).
TJP2 was detected in a paraffin-embedded section of Human Intestinal Cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 μg/ml rabbit anti-TJP2 Antibody (PA1957) overnight at 4°C. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using HRP Conjugated Rabbit IgG Super Vision Assay Kit (Catalog # SV0002) with DAB as the chromogen.

Figure 3. IF analysis of TJP2 using anti-TJP2 antibody (PA1957).
TJP2 was detected in immunocytochemical section of A431 cells. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent (AR0022) for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 5μg/mL rabbit anti-TJP2 Antibody (PA1957) overnight at 4°C. DyLight®488 Conjugated Goat Anti-Rabbit IgG (BA1127) was used as secondary antibody at 1:100 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Figure 4. IF analysis of TJP2 using anti-TJP2 antibody (PA1957).
TJP2 was detected in immunocytochemical section of U2OS cells. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent (AR0022) for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 5μg/mL rabbit anti-TJP2 Antibody (PA1957) overnight at 4°C. DyLight®488 Conjugated Goat Anti-Rabbit IgG (BA1127) was used as secondary antibody at 1:500 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Figure 5. Flow Cytometry analysis of MCF-7 cells using anti-TJP2 antibody (PA1957).
Overlay histogram showing MCF-7 cells stained with PA1957 (Blue line). To facilitate intracellular staining, cells were fixed with 4% paraformaldehyde and permeabilized with permeabilization buffer. The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-TJP2 Antibody (PA1957, 1μg/1x10⁶ cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (BA1127, 5-10μg/1x10⁶ cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1μg/1x10⁶) used under the same conditions. Unlabelled sample without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.