- 720032 - Invitrogen Antibodies GLIS1 antibody

No submitted references: Submit reference

No comments: Submit comment

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on whole cell extracts (20 µg lysate) of Jurkat (Lane 1), Hep G2 (Lane 2), A431 (Lane 3), A549 (Lane 4) and MCF7 (Lane 5). The blots were probed with Anti-GLIS1 Rabbit Polyclonal Antibody (Product # 720032, 1-3 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:2500 dilution). A 80 kDa band corresponding to GLIS1 was observed across cell lines tested. The antibody also detects another protein at 120 kDa which could be GLIS3. Known quantity of protein samples were electrophoresed using Novex®NuPAGE®4-12% Bis-Tris gel (Product # NP0321BOX), XCell SureLock Electrophoresis System (Product # EI0002), and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary antibody following blocking with 5% skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on whole cell extracts (20 µg lysate) of Jurkat (Lane 1), Hep G2 (Lane 2), A431 (Lane 3), A549 (Lane 4) and MCF7 (Lane 5). The blots were probed with Anti-GLIS1 Rabbit Polyclonal Antibody (Product # 720032, 1-3 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:2500 dilution). A 80 kDa band corresponding to GLIS1 was observed across cell lines tested. The antibody also detects another protein at 120 kDa which could be GLIS3. Known quantity of protein samples were electrophoresed using Novex®NuPAGE®4-12% Bis-Tris gel (Product # NP0321BOX), XCell SureLock Electrophoresis System (Product # EI0002), and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary antibody following blocking with 5% skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on whole cell extracts (20 µg lysate) of Jurkat (Lane 1), Hep G2 (Lane 2), A431 (Lane 3), A549 (Lane 4) and MCF7 (Lane 5). The blots were probed with Anti-GLIS1 Rabbit Polyclonal Antibody (Product # 720032, 1-3 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:2500 dilution). A 80 kDa band corresponding to GLIS1 was observed across cell lines tested. The antibody also detects another protein at 120 kDa which could be GLIS3. Known quantity of protein samples were electrophoresed using Novex®NuPAGE®4-12% Bis-Tris gel (Product # NP0321BOX), XCell SureLock Electrophoresis System (Product # EI0002), and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary antibody following blocking with 5% skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence was performed on fixed and permeabilized SH-SY5Y cells for detection of Glis1 using Anti-Glis1 Rabbit Polyclonal Antibody (Product # 720032, 1 µg/mL) and labeled with Goat anti-Rabbit IgG (H+L) Superclonal Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034, 0.4 µg/mL, 1:2500). Panel a) shows representative cells that were stained for detection and localization of GLIS1 protein (green), Panel b) is stained for nuclei (blue) using SlowFade® Gold Antifade Mountant with DAPI (Product # S36938, 1:50). Panel c) represents cytoskeletal F-actin staining using Alexa Fluor® 594 Phalloidin (Product # A12381, 1:200). Panel d) is a composite image of Panels a, b and c clearly demonstrating nuclear localization of GLIS1 Panel e) represents control cells with no primary antibody to assess background.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of Glis1 showing staining in the nucleus and weak staining in the cytoplasm of paraffin-embedded human colon tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a Glis1 Rabbit Polyclonal Antibody (Product # 720032) diluted in 3% BSA-PBS at a dilution of 1:20 for 1 hour at 37ºC in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow Cytometry analysis of Glis1 was performed on SH-SY5Y cells labeled with Anti-Glis1 Rabbit Polyclonal Antibody (Product# 720032, 2-4 ug/ 1M cells) or with rabbit isotype control and detected with Goat anti-Rabbit IgG (H+L) Superclonalª Secondary Antibody, Alexa Fluor¨ 488 conjugate (Product # A27034, 0.4 ug/ml, 1:2500) as represented by the red and pink histograms respectively. The purple histogram represents unstained control cells and the green histogram represents no-primary-antibody control. A representative 10,000 cells were acquired and analyzed for each sample using an Attune¨ Acoustic Focusing Cytometer (4468770).

720032