- 700186 - Invitrogen Antibodies SUMO3 antibody

Ahmed MM, Gardiner KJ
Journal of neuroscience methods 2011 Mar 15;196(1):99-106

No comments: Submit comment

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of SUMO-3 was performed by loading 20 µg of Mouse Esophagus (lane1), Mouse Skin (lane2), Mouse Skeletal muscle (lane3), Jurkat (lane4) and HepG2 (lane5) lysate using Novex® NuPAGE®4-12 % Bis-Tris gel (Product # NP0322BOX), XCell SureLock™ Electrophoresis System (Product # EI0002), Novex® Sharp Pre-Stained Protein Standard (Product # LC5800), and iBlot® Dry Blotting System (Product # IB21001). Proteins were transferred to a nitrocellulose membrane and blocked with 5 % skim milk for 1 hour at room temperature. SUMO-3 was detected at ~ 20 kDa using SUMO-3 Recombinant Rabbit Monoclonal Antibody (Product # 700186) at 1:1000 dilution in 5 % skim milk at 4ºC overnight on a rocking platform. Goat anti-Rabbit IgG - HRP Secondary Antibody (Product # G-21234) at 1:5000 dilution was used and chemiluminescent detection was performed using Pierce™ ECL Western blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of SUMO 3 in U2OS cells using a SUMO 3 recombinant rabbit monoclonal antibody (Product # 700186) at a dilution of 5.0 µg/mL followed by detection using an Alexa Fluor 488-conjugated goat anti-rabbit secondary antibody at a dilution of 1:1000 (left) and in the presence of the immunogenic phosphopeptide (right).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of SMT3/ SUMO3 was done on 70% confluent log phase MCF7 cells. The cells were fixed with 4% paraformaldehyde for 15 minutes, permeabilized with 0.25% Triton X-100 for 10 minutes, and blocked with 5% BSA for 1 hour at room temperature. The cells were labeled with SMT3/SUMO 3 (1H9L17), Recombinant Rabbit Monoclonal Antibody (Product # 700186) at 1 µg/mL in 1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Rabbit Secondary Antibody, Alexa Fluor® 488 conjugate at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d is a merged image showing Nuclear localization. Panel e is a no primary antibody control. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of SUMO 3 in U2OS cells using a SUMO 3 recombinant rabbit monoclonal antibody (Product # 700186) at a dilution of 5.0 µg/mL followed by detection using an Alexa Fluor 488-conjugated goat anti-rabbit secondary antibody at a dilution of 1:1000 (left) and in the presence of the immunogenic phosphopeptide (right).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of SUMO 3 in formalin-fixed, paraffin-embedded human colon carcimona using a SUMO 3 monoclonal antibody (Product # 700186) at a dilution of 5 µg/mL. Tissues were pretreated with EDTA and staining was visualized using DAB. Images were taken at a magnification of 20x. Results show cytoplasmic staining in tumor cells.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow cytometry analysis of SUMO3 was done on PC-3 cells. Cells were fixed with 70% ethanol for 10 minutes, permeabilized with 0.25% Triton™ X-100 for 20 minutes, and blocked with 5% BSA for 30 minutes at room temperature. Cells were labeled with ABfinity™ SUMO3 Recombinant Rabbit Monoclonal Antibody (700186, red histogram) or with rabbit isotype control (pink histogram) at 3-5 ug/million cells in 2.5% BSA. After incubation at room temperature for 2 hours, the cells were labeled with Alexa Fluor® 488 Goat Anti-Rabbit Secondary Antibody (A11008) at a dilution of 1:400 for 30 minutes at room temperature. The representative 10,000 cells were acquired and analyzed for each sample using an Attune® Acoustic Focusing Cytometer. The purple histogram represents unstained control cells and the green histogram represents no-primary-antibody control.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow cytometry analysis of SUMO 3 in Jurkat cells using a SUMO 3 recombinant rabbit monoclonal antibody (Product # 700186) at a dilution of 0.5 µg. Cells were fixed and permeabilized using FIX & PERM (Product # GAS-004) reagent, and detection was performed using an Alexa Fluor 488 goat anti-rabbit IgG (black) compared to a control without primary antibody (gray). Pre-incubation with the immunogenic peptide decreased the signal (red).

700186