- 33-8900 - Invitrogen Antibodies IQGAP1 antibody

Submitted references Purpuric drug eruptions induced by EGFR tyrosine kinase inhibitors are associated with IQGAP1-mediated increase in vascular permeability.

Sheen YS, Lin MH, Tzeng WC, Chu CY
The Journal of pathology 2020 Apr;250(4):452-463

AKAP220 manages apical actin networks that coordinate aquaporin-2 location and renal water reabsorption.

Whiting JL, Ogier L, Forbush KA, Bucko P, Gopalan J, Seternes OM, Langeberg LK, Scott JD
Proceedings of the National Academy of Sciences of the United States of America 2016 Jul 26;113(30):E4328-37

IQGAP1 is associated with nuclear envelope reformation and completion of abscission.

Lian AT, Hains PG, Sarcevic B, Robinson PJ, Chircop M
Cell cycle (Georgetown, Tex.) 2015;14(13):2058-74

Patient Mutation Directed shRNA Screen Uncovers Novel Bladder Tumor Growth Suppressors.

Hensel J, Duex JE, Owens C, Dancik GM, Edwards MG, Frierson HF, Theodorescu D
Molecular cancer research : MCR 2015 Sep;13(9):1306-15

IQGAP1 scaffold-kinase interaction blockade selectively targets RAS-MAP kinase-driven tumors.

Jameson KL, Mazur PK, Zehnder AM, Zhang J, Zarnegar B, Sage J, Khavari PA
Nature medicine 2013 May;19(5):626-630

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of HeLa cell lysates using IQGAP Monoclonal Antibody, Mouse (Product # 33-8900).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on membrane enriched extract (30 µg) of K562 (Lane 1), HEK 293 (Lane 2) and HeLa (Lane 3). The blots were probed with IQGAP1 Mouse Monoclonal Antibody (Product # 33-8900, 2 µg/mL) and detected by chemiluminescence using Goat anti-Mouse IgG (H+L) Superclonal Secondary Antibody, HRP conjugate (Product # A28177, 0.4 µg/mL, 1:2500 dilution). A 190 kDa band corresponding to IQGAP1 was observed across the cell lines tested. 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 by overnight transfer method. 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: Knockdown of IQGAP1 was achieved by transfecting THP-1 with IQGAP1 specific siRNAs (Silencer® select Product # s16837, s16839). Western Blot analysis (Fig. a) was performed using hole cell extracts (60 µg lysate) from the IQGAP1 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The Blot was probed with IQGAP1 Monoclonal Antibody (AF1) (Product # 33-8900, 1:1000 dilution) and Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:10000 dilution). Densitometric analysis of this western Blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knockdown confirms that antibody is specific to IQGAP1. The gel was equilibrated in 20% EtOH for 10 mins prior to transfer for improving detection levels of IQGAP1

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot was performed using Anti-IQGAP1 Monoclonal Antibody (AF1) (Product # 33-8900) and a 189 kDa band corresponding to IQGAP1 was observed in HaCaT, THP-1 and A431 but not in HEK-293 or K-562 which are reported low expressing for the protein. Whole cell extracts (60 µg lysate) of HaCaT (Lane 1), THP-1 (Lane 2), HEK-293 (Lane 3), A-431 (Lane 4) and K-562 (Lane 5) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The Blot was probed with the primary antibody (1 µg/mL dilution) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:8000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).The gel was equilibrated in 20% EtOH for 10 mins prior to transfer for improving detection levels of IQGAP1.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of IQGAP1 Monoclonal Antibody was performed using 70% confluent log phase HeLa cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with IQGAP1 (AF1) Mouse Monoclonal Antibody (Product # 33-8900) at 2 µg/mL in 0.1% BSA, incubated for 3 hours at room temperature and then labeled with Goat anti-Mouse IgG (H+L) Superclonal Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175) 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 represents the merged image showing cytoplasmic localization. Panel e shows the no primary antibody control. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: IQGAP1 is expressed and co-localized in CD-31-positive vascular endothelial cells in the lesional skin of purpuric eruptions. (A) Three-color immunofluorescence images were obtained for IQGAP1 (red), pEGFR (green), and nuclei (blue) in lesional skin and normal skin from healthy controls. The administration of erlotinib resulted in reduced pEGFR and increased IQGAP1 expression of the basal keratinocytes and vessels in the upper dermis. Images were acquired using confocal microscopy (10x objective). (B) Three-color immunofluorescence images were obtained for IQGAP1 (red), CD31 (green), and nuclei (blue) in lesional skin and normal skin from healthy controls. Double-positive cells are yellow in merged images. IQGAP1-positive cells were co-localized with the CD31-positive cells in the superficial dermis from the patient treated with erlotinib. The regions in red boxes are shown magnified (right). All cells were negative using an isotype control. The images were acquired by confocal microscopy using a 20x objective (bar = 100 mum).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Expression of IQGAP1, VE-cadherin/catenin complexes, and actin during erlotinib treatment in HMEC-1s. HMEC-1s were treated with erlotinib with indicated concentrations for 72 h. (A) Erlotinib treatment increased IQGAP1 expression but did not change VE-cadherin, E-cadherin, beta-catenin, or alpha-catenin protein expression significantly. (B) Increased IQGAP1 fluorescence at the cell borders and perinucleus area was detected after erlotinib treatment. The erlotinib-treated HMEC-1s revealed a punctate alpha-catenin fluorescence in the areas of cell-cell contacts compared with control cells. Erlotinib treatment did not affect the distribution of VE-cadherin, beta-catenin, and actin filament in HMEC-1s. The images were acquired by confocal microscopy using a 100x objective (bar = 10 mum).

33-8900

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