Western blot (WB): | 1:500-2000 |
Immunohistochemistry (IHC): | 1:50-400 |
Immunocytochemistry/Immunofluorescence (ICC/IF): | 1:50-400 |
Flow Cytometry (Fixed): | 1:50-200 |
(Boiling the paraffin sections in 10mM citrate buffer,pH6.0,or PH8.0 EDTA repair liquid for 20 mins is required for the staining of formalin/paraffin sections.) Optimal working dilutions must be determined by end user. |
Figure 1. Western blot analysis of ADO using anti- ADO antibody (PB10032).
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 50ug of sample under reducing conditions.
Lane 1: rat brain tissue lysates,
Lane 2: rat testicular tissue lysates,
Lane 3: rat spleen tissue lysates,
Lane 4: human MCF-7 whole cell lysates,
Lane 5: human SW620 whole cell lysates,
Lane 6: mouse spleen tissue lysates,
Lane 7: mouse heart tissue lysates,
Lane 8: mouse HEPA1-6 whole cell lysates,
Lane 9: mouse SP2/0 whole cell lysates,
After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA 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- ADO antigen affinity purified polyclonal antibody (Catalog # PB10032) 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 1:10000 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 ADO at approximately 28,30KD. The expected band size for ADO is at 30KD.
Figure 2. Flow Cytometry analysis of A549 cells using anti-ADO antibody (PB10032).Overlay histogram showing A549 cells stained with PB10032 (Blue line).The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-ADO Antibody (PB10032, 1:100) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (BA1127, 1:100) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1:100) used under the same conditions. Unlabelled sample (Red line) was also used as a control.
Figure 3. Flow Cytometry analysis of U251 cells using anti-ADO antibody (PB10032).Overlay histogram showing U251 cells stained with PB10032 (Blue line).The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-ADO Antibody (PB10032, 1:100) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (BA1127, 1:100) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1:100) used under the same conditions. Unlabelled sample (Red line) was also used as a control.
Figure 4. Flow Cytometry analysis of Hela cells using anti-ADO antibody (PB10032).Overlay histogram showing Hela cells stained with PB10032 (Blue line).The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-ADO Antibody (PB10032, 1:100) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (BA1127, 1:100) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1:100) used under the same conditions. Unlabelled sample (Red line) was also used as a control.
Figure 5. ICC analysis using anti- ADO antibody (PB10032). was detected in immersion fixed A549 cell line. Cells were stained using the Dylight488-conjugated Anti-rabbit IgG Secondary Antibody (green)(Catalog # BA1127) and counterstained with DAPI (blue).
Figure 6. IHC analysis using anti- ADO antibody (PB10032). detected in paraffin-embedded section of rat brain tissue. Biotinylated goat anti-rabbit IgG was used as secondary antibody. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1022) with DAB as the chromogen.
Figure 7. IHC analysis using anti- ADO antibody (PB10032). detected in paraffin-embedded section of mouse brain tissue. Biotinylated goat anti-rabbit IgG was used as secondary antibody. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1022) with DAB as the chromogen.
Figure 1. Western blot analysis of ADO using anti- ADO antibody (PB10032).
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 50ug of sample under reducing conditions.
Lane 1: rat brain tissue lysates,
Lane 2: rat testicular tissue lysates,
Lane 3: rat spleen tissue lysates,
Lane 4: human MCF-7 whole cell lysates,
Lane 5: human SW620 whole cell lysates,
Lane 6: mouse spleen tissue lysates,
Lane 7: mouse heart tissue lysates,
Lane 8: mouse HEPA1-6 whole cell lysates,
Lane 9: mouse SP2/0 whole cell lysates,
After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA 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- ADO antigen affinity purified polyclonal antibody (Catalog # PB10032) 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 1:10000 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 ADO at approximately 28,30KD. The expected band size for ADO is at 30KD.
Figure 2. Flow Cytometry analysis of A549 cells using anti-ADO antibody (PB10032).Overlay histogram showing A549 cells stained with PB10032 (Blue line).The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-ADO Antibody (PB10032, 1:100) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (BA1127, 1:100) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1:100) used under the same conditions. Unlabelled sample (Red line) was also used as a control.
Figure 3. Flow Cytometry analysis of U251 cells using anti-ADO antibody (PB10032).Overlay histogram showing U251 cells stained with PB10032 (Blue line).The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-ADO Antibody (PB10032, 1:100) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (BA1127, 1:100) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1:100) used under the same conditions. Unlabelled sample (Red line) was also used as a control.
Figure 4. Flow Cytometry analysis of Hela cells using anti-ADO antibody (PB10032).Overlay histogram showing Hela cells stained with PB10032 (Blue line).The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-ADO Antibody (PB10032, 1:100) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (BA1127, 1:100) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1:100) used under the same conditions. Unlabelled sample (Red line) was also used as a control.
Figure 5. ICC analysis using anti- ADO antibody (PB10032). was detected in immersion fixed A549 cell line. Cells were stained using the Dylight488-conjugated Anti-rabbit IgG Secondary Antibody (green)(Catalog # BA1127) and counterstained with DAPI (blue).
Figure 6. IHC analysis using anti- ADO antibody (PB10032). detected in paraffin-embedded section of rat brain tissue. Biotinylated goat anti-rabbit IgG was used as secondary antibody. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1022) with DAB as the chromogen.
Figure 7. IHC analysis using anti- ADO antibody (PB10032). detected in paraffin-embedded section of mouse brain tissue. Biotinylated goat anti-rabbit IgG was used as secondary antibody. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1022) with DAB as the chromogen.