- 39-3700 - Invitrogen Antibodies NQO1 antibody

Submitted references Astrocytic Nrf2 expression protects spinal cord from oxidative stress following spinal cord injury in a male mouse model.

Zhao W, Gasterich N, Clarner T, Voelz C, Behrens V, Beyer C, Fragoulis A, Zendedel A
Journal of neuroinflammation 2022 Jun 6;19(1):134

The caspase-2 substrate p54nrb exhibits a multifaceted role in tumor cell death susceptibility via gene regulatory functions.

Eichler M, Distler U, Nasrullah U, Krishnan A, Kaulich M, Husnjak K, Eberhardt W, Rajalingam K, Tenzer S, Pfeilschifter J, Imre G
Cell death & disease 2022 Apr 20;13(4):386

NRF2 Dysregulation in Hepatocellular Carcinoma and Ischemia: A Cohort Study and Laboratory Investigation.

Ziv E, Zhang Y, Kelly L, Nikolovski I, Boas FE, Erinjeri JP, Cai L, Petre EN, Brody LA, Covey AM, Getrajdman G, Harding JJ, Sofocleous C, Abou-Alfa GK, Solomon SB, Brown KT, Yarmohammadi H
Radiology 2020 Oct;297(1):225-234

A ruthenium(II)-curcumin compound modulates NRF2 expression balancing the cancer cell death/survival outcome according to p53 status.

Garufi A, Baldari S, Pettinari R, Gilardini Montani MS, D'Orazi V, Pistritto G, Crispini A, Giorno E, Toietta G, Marchetti F, Cirone M, D'Orazi G
Journal of experimental & clinical cancer research : CR 2020 Jun 30;39(1):122

Novel high throughput pooled shRNA screening identifies NQO1 as a potential drug target for host directed therapy for tuberculosis.

Li Q, Karim AF, Ding X, Das B, Dobrowolski C, Gibson RM, Quiñones-Mateu ME, Karn J, Rojas RE
Scientific reports 2016 Jun 14;6:27566

NQO1 inhibits proteasome-mediated degradation of HIF-1α.

Oh ET, Kim JW, Kim JM, Kim SJ, Lee JS, Hong SS, Goodwin J, Ruthenborg RJ, Jung MG, Lee HJ, Lee CH, Park ES, Kim C, Park HJ
Nature communications 2016 Dec 14;7:13593

Cross-regulations among NRFs and KEAP1 and effects of their silencing on arsenic-induced antioxidant response and cytotoxicity in human keratinocytes.

Zhao R, Hou Y, Zhang Q, Woods CG, Xue P, Fu J, Yarborough K, Guan D, Andersen ME, Pi J
Environmental health perspectives 2012 Apr;120(4):583-9

Prolonged inorganic arsenite exposure suppresses insulin-stimulated AKT S473 phosphorylation and glucose uptake in 3T3-L1 adipocytes: involvement of the adaptive antioxidant response.

Xue P, Hou Y, Zhang Q, Woods CG, Yarborough K, Liu H, Sun G, Andersen ME, Pi J
Biochemical and biophysical research communications 2011 Apr 8;407(2):360-5

Interaction of human NAD(P)H:quinone oxidoreductase 1 (NQO1) with the tumor suppressor protein p53 in cells and cell-free systems.

Anwar A, Dehn D, Siegel D, Kepa JK, Tang LJ, Pietenpol JA, Ross D
The Journal of biological chemistry 2003 Mar 21;278(12):10368-73

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis was performed on membrane enriched extracts (30 µg lysate) of Hep G2 (Lane 1), HeLa (Lane 2), A549 (Lane 3), MCF7 (Lane 4), U-87 MG (Lane 5), SH-SY5Y (Lane 6). The blot was probed with Anti-NQO1 Mouse Monoclonal Antibody (Product # 39-3700, 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 ~30 kDa band corresponding to NQO1 was observed across the cell lines tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 12 % Bis-Tris gel (Product # NP0342BOX), 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® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody. Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using NQO1 Monoclonal Antibody (A180), (Product # 39-3700) and a 30 kDa band corresponding to NQO1 was observed across cell lines tested except in SH-SY5Y, MOLT-4 and HEK-293. Whole cell extracts (30 µg lysate) of HepG2 (Lane 1), HeLa (Lane 2), A549 (Lane 3), U-87 MG (Lane 4), SH-SY5Y (Lane 5), MOLT-4 (Lane 6) and HEK-293 (Lane 7) were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris 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 (2µg/mL) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L), Superclonal™ Recombinant Secondary Antibody, HRP conjugate (Product # A28177, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of NQO1 was performed using 70% confluent log phase Hep G2 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 NQO1 (A180) Mouse Monoclonal Antibody (Product # 39-3700) at 2µg/mL in 0.1% BSA and 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)
Main image
Experimental details: Flow cytometry analysis of NQO1 was done on Hep G2 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 NQO1 Mouse Monoclonal Antibody (39-3700, red histogram) or with mouse isotype control (pink histogram) at 3-5 µg/million cells in 2.5% BSA. After incubation at room temperature for 2 hours, the cells were labeled with Alexa Fluor® 488 Rabbit Anti-Mouse Secondary Antibody (A11059) 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)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 1 Stable knockdown of NRF2 and KEAP1 by lentiviral shRNAs in HaCaT cells. ( A and C ) Silencing effect of NRF2 ( A ) and KEAP1 ( C ) in HaCaT cells transduced with shRNA lentivirus targeted against human NRF2 , KEAP1 , or scrambled (SCR) nontarget negative control (sh-SCR) ( n = 3). sh-NRF2-1 through sh-NRF2-5 indicate shRNAs targeting NRF2 ; sh-KEAP1-1 through sh-KEAP1-5, shRNAs targeting KEAP1 . ( B and D ) Protein expression of NRF2 ( B ), KEAP1 ( D ), and NRF2 target genes NQO1 and GCLC in NRF2 -KD and KEAP1 -KD cells. Cells were treated with 50 muM tBHQ for the indicated time, and whole-cell lysates were separated on 4-12% Tris-glycine gels. beta-ACTIN was used as loading control. ( E ) Intracellular GSH content in NRF2 -KD and KEAP1 -KD cells ( n = 6). * p < 0.05 versus SCR ( A, C, E ).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 2 Effect of stable knockdown of NRF2 or KEAP1 in HaCaT cells on the induction of ARE-dependent genes in response to acute iAs 3+ exposure. Cells were exposed to iAs 3+ or vehicle (medium) for 6 hr. ( A , B ) Concentration response of iAs 3+ -induced mRNA ( A ) and protein ( B ) expression of NRF2 and ARE-dependent antioxidant genes in NRF2 -KD and SCR cells. ( C , D ) Concentration response of iAs 3+ -induced mRNA ( C ) and protein ( D ) expression of KEAP1 and ARE-dependent antioxidant genes in KEAP1 -KD and SCR cells. Integers in parentheses after gene names in A and C are cross-threshold PCR cycle numbers in SCR cells treated with vehicle. * p < 0.05 versus SCR with the same treatment.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig. 4 RuCUR compound induces NRF2 pathway in mutp53-carrying cancer cells. ( a ) Western blot analysis of the indicated protein was performed in T98 and SKBR3 cells untreated or treated with RuCUR (100 muM) for 24 h. Actin was used as protein loading control. Densitometry was performed using ImageJ software. Relative band intensities value were normalized to beta-actin (loading control) and finally quantified with respect to untreated control arbitrarily set to 1.0. ( b ) Total mRNA was extracted from T98 and SKBR3 cells untreated or treated with RuCUR (100 muM) for 24 h. The indicated gene expression was assayed by semi-quantitative polymerase chain reaction (PCR) of reverse-transcribed cDNA. Densitometric analysis was performed using ImageJ software to calculate the gene/28S ratio. Histograms represent the fold increase quantified with respect to controls set to 1.0, +- SD. * ( p

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Loss of p54nrb leads to an altered expressional pattern of carcinogenesis relevant genes. A Volcano plot diagram of LC/MS data derived from the analysis of HeLa shRNA-control versus shRNA-p54nrb cells. ( n = 3). Arrowheads depict significantly ( p < 0.01) upregulated (arrow up, red) and downregulated (arrow down, blue) proteins, and the total numbers of both categories are indicated on the diagram. B Up- and C downregulated proteins ( p < 0.01, Log 2 ratio either >0.5 or < -0.5) of HeLa-shRNA-p54nrb cells compared to shRNA control cells, which exert tumor regulatory and/or apoptosis regulatory properties. Differently colored oval diagrams represent various functional subcategories of these hits. D Immunoblot of HeLa shRNA-control and shRNA-p54nrb cells. Detection of p54nrb, gelsolin, cathepsin-Z, NQO1, and TPD52 levels. E Quantitative evaluation of immunoblots of p54nrb, gelsolin, cathepsin-Z, NQO1, TPD52, and CDKN2A from HeLa shRNA-control and shRNA-p54nrb#1 and #3 cells. The bands were normalized to the shRNA-control samples. Significance was calculated with one sample t test, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, n = 3.

39-3700

Antibody data