Figure 1. Western blot analysis of IDE using anti-IDE antibody (MBS1750623).
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: human Hela whole cell lysates,
Lane 2: human placenta tissue lysates,
Lane 3: human COLO-320 whole cell lysates,
Lane 4: human HepG2 whole cell lysates,
Lane 5: human SGC-7901 whole cell lysates,
Lane 6: human Jurkat whole cell lysates,
Lane 7: rat stomach tissue lysates,
Lane 8: mouse skeletal muscle tissue lysates,
Lane 9: mouse stomach tissue lysates,
Lane 10: mouse brain tissue 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-IDE antigen affinity purified polyclonal antibody at 0.5ug/mL overnight at 4 degree 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 with Tanon 5200 system. A specific band was detected for IDE at approximately 118KD. The expected band size for IDE is at 118KD.
Figure 2. IHC analysis of IDE using anti-IDE antibody (MBS1750623).
IDE was detected in paraffin-embedded section of human mammary cancer tissue. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1ug/ml rabbit anti-IDE Antibody (MBS1750623) overnight at 4 degree C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37 degree C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) with DAB as the chromogen.
Figure 3. IHC analysis of IDE using anti-IDE antibody (MBS1750623).
IDE was detected in paraffin-embedded section of rat pancreas tissue. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1ug/ml rabbit anti-IDE Antibody (MBS1750623) overnight at 4 degree C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37 degree C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) with DAB as the chromogen.
Figure 4. Flow Cytometry analysis of Hela cells using anti-IDE antibody (MBS1750623).
Overlay histogram showing Hela cells stained with MBS1750623 (Blue line).The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-IDE Antibody (MBS1750623,1ug/1x10^6 cells) for 30 min at 20 degree C. DyLight®488 conjugated goat anti-rabbit IgG (5-10ug/1x10^6 cells) was used as secondary antibody for 30 minutes at 20 degree C. Isotype control antibody (Green line) was rabbit IgG (1ug/1x106) used under the same conditions. Unlabelled sample (Red line) was also used as a control.
Figure 5. Flow Cytometry analysis of CACO-2 cells using anti-IDE antibody (MBS1750623).
Overlay histogram showing CACO-2 cells stained with MBS1750623 (Blue line).The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-IDE Antibody (MBS1750623,1ug/1x10^6 cells) for 30 min at 20 degree C. DyLight®488 conjugated goat anti-rabbit IgG (5-10ug/1x10^6 cells) was used as secondary antibody for 30 minutes at 20 degree C. Isotype control antibody (Green line) was rabbit IgG (1ug/1x106) used under the same conditions. Unlabelled sample (Red line) was also used as a control.