Product Search for n2: Novus Biologicals

AF-401-NA

IL-1 beta/IL-1F2 Antibody [Unconjugated]

(3 Reviews)

(538 Publications)

100 μg

AF-401-NA

Western blot shows lysates of THP-1 human acute monocytic leukemia cell line untreated (-) or treated (+) with 200 nM PMA for 24 hours and 10 µg/mL LPS for 4 hours and RAW 264.7 mouse monocyte/macrophage cell line untreated (-) or treated (+) with 10 µg/mL LPS for 24 hours. PVDF membrane was probed with 0.25 µg/mL of Goat Anti-Mouse IL-1 beta /IL-1F2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF-401-NA) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (HAF017). A specific band was detected for IL-1 beta /IL-1F2 at approximately 35 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

IL-1 beta /IL-1F2 was detected in immersion fixed MCF-7 human breast cancer cell line using Goat Anti-Mouse IL-1 beta /IL-1F2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF-401-NA) at 8 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Goat IgG Secondary Antibody (red; NL001) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

IL-1 beta /IL-1F2 was detected in perfusion fixed frozen sections of mouse thymus using Goat Anti-Mouse IL-1 beta /IL-1F2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF-401-NA) at 15 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Goat HRP-DAB Cell & Tissue Staining Kit (brown; CTS008) and counterstained with hematoxylin (blue). View our protocol for Chromogenic IHC Staining of Frozen Tissue Sections.

Simple Western lane view shows lysates of RAW 264.7 mouse monocyte/macrophage cell line untreated (-) or treated (+) with 10 µg/mL LPS for 24 hours, loaded at 0.5 mg/mL. A specific band was detected for IL‑1 beta /IL‑1F2 at approximately 40 kDa (as indicated) using 2.5 µg/mL of Goat Anti-Mouse IL‑1 beta /IL‑1F2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF-401-NA) followed by 1:50 dilution of HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF109). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.

Recombinant MouseIL‑1 beta /IL‑1F2 (Catalog # 401-ML) stimulates proliferation in the the D10.G4.1 mouse helper T cell line in a dose-dependent manner (orange line) as measured by Resazurin (Catalog # AR002). Proliferation elicited by Recombinant Mouse IL‑1 beta /IL‑1F2 (50 pg/mL) is neutralized (green line) by increasing concentrations of Goat Anti-Mouse IL-1 beta /IL-1F2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF-401-NA). The ND50 is typically ≤0.25 µg/mL.

Quercetin inhibits auto-reactive NLRP3 inflammasome.(A) IL-1 beta concentrations in Nlrp3A350V/A350V and WT BMDM culture supernatants primed with LPS (500 ng/ml) for 90 min and treated with quercetin 1 h after LPS. (B) Supernatants of BMDM were analyzed by anti-ASC, anti-Caspase-1 immunoblotting. Lysates of BMDM were analyzed by anti-IL-1 beta and anti-beta -actin immunoblotting. Data shown are representative of two or more experiments (means ± SD) ***p VC006). Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using VisUCyte Antigen Retrieval Reagent-Basic (Catalog # VCTS021). Tissue was stained using DAB (brown) and counterstained with hematoxylin (blue). Specific staining was localized to cytoplasm in epithelial cells. View our protocol for IHC Staining with VisUCyte HRP Polymer Detection Reagents.

GRA7-I-induced inflammasome activation was required for antimicrobial activity in MTB-infected macrophages.(A and B) BMDMs from PKC alpha +/+ and PKC alpha -/- mice (A) and BMDMs were transduced with lentivirus-shRNA-NS or lentivirus-shRNA-ASC for 2 days (B) infected with MTB (MOI = 1) for 4 h and then stimulated with rGRA7 (1, 5, 10 μg/ml) and its mutants for 18 h. IB analysis for IL-1 beta p17, IL-18 p18, or caspase-1 p10 in supernatants (SN), ASC, NLRP3, pro-IL-1 beta , pro-IL-18, or pro-caspase-1 in whole-cell lysates (WCL). Actin was used as a loading control. (C and D) Intracellular survival of MTB was assessed by CFU assay. BMDMs were infected with MTB for 4 h, followed by treatment with rGRA7, and then lysed to determine intracellular bacterial loads. The data are representative of five independent experiments with similar results (A and B). Data shown are the mean ± SD of five experiments (C and D). Significant differences (*P p>0.001,***p<0.001, ns = not significant ) as determined by one way ANOVA with Tukey's post test. Image collected and cropped by CiteAb from the following publication (//dx.plos.org/10.1371/journal.pone.0040722), licensed under a CC-BY license. Not internally tested by R&D Systems.

Effects of IL-1 beta neutralization on the accumulation of MDSCs in lungs.(A) Immunofluorescent photos showed that Gr-1+ (green)/CD11b+ (red) cells indicate MDSCs and increased in lungs of DJ-1 KO mice. Treatment with anti-IL-1 beta neutralizing antibody antagonized the accumulation of MDSCs. DAPI was used to stain nuclei. (B) Bar chart showed the summarized results of MDSCs in lungs of WT and DJ-1 KO mice. Note that the neutralizing antibody significantly reduced the number of MDSCs accumulated in lungs. n = 9 for each group, * p < 0.05 compared with control IgG-treated WT mice, # p<0.05 compared with respective control IgG in WT and DJ-1 KO mice. Image collected and cropped by CiteAb from the following publication (//dx.plos.org/10.1371/journal.pone.0115827), licensed under a CC-BY license. Not internally tested by R&D Systems.

Pro-IL-1 beta degradation is induced by the autophagy inhibitors chloroquine and bafilomycin A, and by the proteasome activator betulinic acid. (A) Chloroquine selectively attenuates IL-1 beta release from cardiac fibroblasts. The cells were incubated with LPS and/or 20 μM chloroquine phosphate as indicated for 18 h, then activated with 3 mM ATP for 60 min. IL-1 beta , TNF, MIP-2, and IL-6 were quantified in the conditioned media (n = 3). (B,C) Cardiac fibroblasts were incubated with LPS for 18 h with or without 20 μM chloroquine phosphate and pro-IL-1 beta quantified with western blot (n = 3). (D) Pro-IL-1 beta mRNA expression was quantified with qPCR (n = 3). (E,F) Cardiac fibroblasts were incubated with LPS for 20 h with or without 10% FCS or 20 μM chloroquine phosphate as indicated. The amount of total ubiquitinated protein was quantified with western blot (n = 4). (G,H) Cardiac fibroblasts were incubated with 10% FCS with or without 10 ng/mL LPS and/or 100 nM bafilomycin A1 for 20 h. Pro-IL-1 beta was quantified with western blot. (I,J) Cardiac fibroblasts were incubated with 10% FCS with or without 10 ng/mL LPS and/or 20 μM betulinic acid as indicated for 20 h. Pro-IL-1 beta was quantified with western blot. All columns are mean with SEM. *p < 0.05, **p < 0.01 (paired student t-test). Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/31244838), licensed under a CC-BY license. Not internally tested by R&D Systems.

Pro-IL-1 beta maturation is regulated by POH1-mediated deubiquitination. a, b HEK293T cells were transfected with plasmids encoding Flag-caspase-1, Vsv-ASC, His-pro-IL-1 beta (WT) or His-pro-IL-1 beta -K133R with different amounts (125, 250, 500 ng) of plasmid encoding Flag-POH1 and 500 ng of control plasmid, the secretion of a IL-1 beta in the supernatants was quantified by ELISA 36 h after transfection; b the cell lysates were IB with the indicated antibodies. c Mutants of murine pro-IL-1 beta with replacement of various lysine residues. d-f HEK293T cells were transfected with Flag-POH1, V5-caspase-1, V5-ASC, His-pro-IL-1 beta (WT) or its mutants, along d with or e-f without HA-tagged K63-only Ub, then d cells were subjected to IP with an anti-His antibody and IB with the indicated antibodies. e The IL-1 beta levels in supernatants were measured by ELISA; or f the cell lysates were collected and IB with the indicated antibodies. g Proposed function of POH1 as a negative regulator in mediating IL-1 beta processing. POH1, upregulated by TLR3/4 activation, interacts with and deubiquitinates pro-IL-1 beta , resulting in restriction of IL-1 beta cleavage and secretion. Similar results were obtained from three independent experiments. The results represent the mean ± s.d. of three independent sets of experiments. **p < 0.01, ***p < 0.001 (one-way ANOVA with Tukey’s post-hoc test) Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/30315153), licensed under a CC-BY license. Not internally tested by R&D Systems.

Serum starvation selectively induces pro-IL-1 beta degradation in cardiac fibroblasts. (A) Cells were incubated with 10 ng/mL LPS for 5 or 20 h with or without 10% FCS and western blot analysis of pro-IL-1 beta and NLRP3 performed. Blots are representative of 3 independent experiments. (B) Protein bands were quantified and the ratios to beta -actin normalized to mean control = 1 (n = 3). (C,D) Cardiac fibroblasts were incubated with or without 10% FCS and primed with LPS for 20 h. Western blot analysis of complex II (mitochondrial mass marker) and NLRP3- inflammasome protein components (NEK7, NLRP3, ASC, pro-caspase-1, and pro-IL-1 beta ) were performed. Bands were quantified and the ratios to beta -actin normalized to mean control = 1 (n = 4, for ASC n = 10). (E,F) Cardiac fibroblasts (120,000 cells per well seeded in 6 well plates) were incubated with 10 ng/mL LPS for 20 h, with or without 10% FCS, then stained with 500 nM MitoTracker Deep Red for 45 min before MitoTracker fluorescence intensity was quantified with flow cytometry analysis (633 nm laser). All cells were analyzed and all gated cells included in the analysis. The presented dot plots (E) show the gate in red and are representative for 9 biological repeats. Median MitoTracker intensity with and without 10% FCS are shown (F). All columns are mean with SEM. *p < 0.05,**p < 0.01 (paired student t-test). Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/31244838), licensed under a CC-BY license. Not internally tested by R&D Systems.

Differential effects of IL-1 beta neutralization on melanoma lung nodules in WT and DJ-1 KO mice.(A) IL-1 beta antibody (10μg) or control IgG was intravenously injected into mice once. The melanoma cells (6×104) were intravenously injected 30 minutes later. The mice were sacrificed three weeks later. Bar chart showed that the anti-IL-1 beta neutralizing antibody can effectively lower serum IL-1 beta levels as compared with control IgG in both WT and DJ-1 KO mice. Note that serum IL-1 beta levels were higher in DJ-1 KO mice than those in WT mice. (B) Inhibition of IL-1 beta neutralization on lung nodules formation in DJ-1 KO mice. Upper panels: melanoma nodules in lungs of WT (top) and KO (bottom) mice treated with control IgG (left) or neutralizing anti-IL-1 beta antibody (right). For each experimental condition, the gross and microscopic morphologies of tumor nodules are indicated by arrows, and are shown at left and right sites, respectively. Scale bars: 0.5 mm for lung photographs and 0.5 mm for H&E staining. Lower panel: Bar chart showed the summarized results of lung nodule numbers in WT and DJ-1 KO mice. Note that IL-1 beta neutralization enhanced and suppressed melanoma nodules formation in WT and DJ-1 KO mice, respectively. Data are presented as mean ± S.E.M. n = 5 for each group, * p < 0.05 compared with control IgG-treated WT mice, # p<0.05 compared with respective control IgG in WT and DJ-1 KO mice. Image collected and cropped by CiteAb from the following publication (//dx.plos.org/10.1371/journal.pone.0115827), licensed under a CC-BY license. Not internally tested by R&D Systems.

Inflammasome assembly and mature IL-1 beta release in the spinal cord after sciatic nerve axotomy. a Speck-like costaining of NLRP3 and ASC in injured neurons 3 days after the injury (arrows). b Representative WB images of mature IL-1 beta and beta -actin proteins 3 days after unilateral axotomy of sciatic nerve. c Quantification of active IL-1 beta protein expression. N = 3 animals/group. The graph shows values normalized to beta -actin levels and relative to contralateral side. Mean values are shown on bars. Bars represent average ± SEM. **p < 0.01 (ANOVA with Bonferroni post hoc, ax. + vehicle vs. contralateral side). #p < 0.05, ##p < 0.01 (ANOVA with Bonferroni post hoc, compared to ax. + vehicle). Ax. axotomy Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/35305649), licensed under a CC-BY license. Not internally tested by R&D Systems.

$GRA7-I activated inflammasomes in an ASC-binding dependent manner.(A) Bacterially purified 6xHis-GRA7-I and its mutants were analyzed by Coomassie blue staining (left) or IB with alpha His (right). (B) BMDMs from PKC alpha +/+ and PKC alpha -/- (left) or BMDMs were transduced with lentivirus-shRNA-NS or lentivirus-shRNA-ASC (MOI = 100) with polybrene (8 μg/mL) (right) for 2 days, the cells were stimulated with rGRA7 (5 μg/ml) and its mutants for the indicated times and culture supernatants were harvested and analyzed for cytokine ELISA for IL-1 beta and IL-18. Data shown are the means ± SD of five experiments. Significant differences (*P < 0.05; ***P < 0.001) compared with PKC alpha +/+ or shRNA-NS. (C) IB analysis for IL-1 beta p17, IL-18 p18, or caspase-1 p10 in supernatants (SN), ASC, NLRP3, pro-IL-1 beta , pro-IL-18, or pro-caspase-1 in whole-cell lysates (WCL). Actin was used as a loading control. (D) IB analysis of lysates of BMDMs as in B and C solubilized with Triton X-100–containing buffer, followed by cross-linkage of insoluble fractions with disuccinimidyl suberate to capture ASC oligomers and analysis of those fractions (I + DSS) and soluble fractions (S) with antibody to ASC. Actin was used as a loading control. (E) Fluorescence confocal images showing formation of speck-like ASC pyroptosomes in BMDMs from PKC alpha +/+ and PKC alpha -/- mice were stimulated with rGRA7 and its mutants for 18 h, fixed, immunostained with antibodies for ASC (Alexa 488). Scale bar, 10 μm. The data are representative of five independent experiments with similar results (A and C-E). Image collected and cropped by CiteAb from the following publication (//dx.plos.org/10.1371/journal.ppat.1006126), licensed under a CC-BY license. Not internally tested by R&D Systems.$

The B6 variant of NLRP1B is not cleaved by LF but can form an inflammasome in response to proteolysis.(A) The amino acid sequence of the first 244 residues of NLRP1BB6 was aligned to the homologous sequence of NLRP1B129. The arrow above the alignment indicates the LF-cleavage site in NLRP1B129. Asterisks indicate sites of amino acid identity. (B) For detection of NLRP1B expression, 293T cells were transfected with the indicated amounts of and empty vector (V) or plasmids encoding GFP-HA-NLRP1B129 or GFP-HA-NLRP1BB6 (construct schematics and the predicted molecular weight of each protein is depicted in the upper panel). Cells were treated overnight with anthrax lethal toxin protein (LeTx, 1μg/ml) 24h post-transfection, and then lysates were analyzed by immunoblotting (IB) with the indicated antibodies. For NLRP1 expression and cleavage, CASP1 and IL1B were omitted to prevent cell death and resulting apparent differences of expression. For blots probed with anti-HA (NLRP1B), the lysates were not boiled prior to loading to prevent aggregation and smearing of full-length and FIIND-processed NLRP1B on the immunoblot. To visualize the N-terminal processed form of NLRP1B, the lysates were boiled and resolved on a separate gel. (C) 293T Cells were transfected with the same amount of titrated NLRP1B encoding plasmid and treated as in B, but transfections also included plasmids encoding mouse CASP1 (200ng), IL-1 beta (200ng) and 200ng of empty vector. (D) For detection of NLRP1B expression, 293T cells were transfected for 36h with 250ng of plasmids encoding WT 129 or B6 NLRP1B or mutants engineered to express the TEV-protease site. Each plasmid was co-transfected with either 100ng of empty vector (V) or plasmids encoding TEV-protease (pTEV) or lethal factor protease (pLF), supplemented with 300ng empty vector (pMSCV). Cells were not treated with LeTx and lysates were analyzed by immunoblotting as in (B). (E) Cells were treated as in C, but with 8ng of plasmids encoding NLRP1B, along with 200ng of plasmids encoding mCASP-1 and mIL-1 beta , and 200ng empty vector to normalize plasmid quantities. Lower quantities of NLRP1B were transfected as compared to panel D so to avoid spontaneous NLPR1B activation. For panels B-E, data shown are representative of at least three similar experiments. Image collected and cropped by CiteAb from the following publication (//dx.plos.org/10.1371/journal.ppat.1006052), licensed under a CC-BY license. Not internally tested by R&D Systems.

SHP inhibits the signal 2 activation of NLRP3 inflammasome.(a,b) LPS-primed BMDMs were treated with fenofibrate (10, 20, 50 μM for 4 h), or (c,d) LPS-primed THP-1 cells were treated with recombinant macrophage-stimulating protein (MSP 10, 50, 100 ng ml−1 for 4 h), and then activated with ATP (5 mM) for 30 min, followed by IB analysis of IL-1 beta p17 or caspase-1 p10 in supernatants (SN), pro-IL-1 beta or pro-caspase-1 in whole-cell lysates (WCL), with actin as a loading control. Data are representative of three independent experiments (a,c). Supernatants were collected and subjected to ELISA for IL-1 beta , IL-18, TNF-alpha and IL-8. Data are the means±s.d. of values from four independent experiments (b,d). *P<0.05; **P<0.01; ***P<0.001, compared with control (two-tailed Student’s t-test). FF, fenofibrate. Image collected and cropped by CiteAb from the following publication (//www.nature.com/articles/ncomms7115), licensed under a CC-BY license. Not internally tested by R&D Systems.

NLRP1A is activated by N-terminal proteolysis.(A) The amino acid sequence of the first 244 residues of NLRP1AB6 was aligned to NLRP1B129 and NLRP1BB6. The arrow above the alignment indicates the LF-cleavage site in NLRP1B129. (B) 293T cells were transfected for 36h with 200ng empty vector (V) or plasmids pcDNA3.1-HA-NLRP1B129 or -NLRP1AB6-MYC (CMV promoter) along with 200ng mCASP1 and 200ng mIL-1 beta expression vectors. Cells were treated overnight with anthrax lethal toxin (LeTx, 1μg/ml) 24h post-transfection, and then lysates were analyzed by immunoblotting (IB) with the indicated antibodies. (C) 293T cells were transfected for 36h with plasmids encoding 400ng GFP-HA-NLRP1B129 or GFP-HA-NLRP1AB6 (under the control of the LTR promoter in pMSCV) or with mutants engineered to contain an N-terminal TEV protease site. Cells were also co-transfected with 100ng empty vector (V) or plasmids encoding TEV-protease (pTEV) or lethal factor protease (pLF), plus 300ng empty pMSCV. (D) To assess IL-1 beta cleavage, cells were transfected as in C, but with 8ng of GFP-HA-NLRP1B and co-transfected with vectors encoding 200ng mCASP1 and 200ng mIL-1 beta . For panels B-D, data shown are representative of at least three similar experiments. Image collected and cropped by CiteAb from the following publication (//dx.plos.org/10.1371/journal.ppat.1006052), licensed under a CC-BY license. Not internally tested by R&D Systems.

POH1 inhibits inflammasome-induced pro-IL-1 beta processing. a-c BMDMs from Poh1 delta /+ and Poh1 delta / delta mice were primed with LPS for 12–14 h and then stimulated with ATP (0.5 h), nigericin (Nig, 0.5 h), poly(dA:dT) (5 h) or flagellin (5 h) as indicated, a IL-1 beta , b IL-6 and c TNF alpha levels in supernatants were analysed by ELISA. d-f BMDMs lentivirally transduced with control (Vector) or POH1 were stimulated as in a, d IL-1 beta , e IL-6 and f TNF alpha levels in supernatants were analysed by ELISA. g, h BMDMs g derived from Poh1 delta /+ and Poh1 delta / delta mice or h infected by indicated lentiviruses were treated as in a, the cell lysates (CL) and supernatants (SN) were collected and immunoblotted with the indicated antibodies. Similar results were obtained from three independent experiments. The results represent the mean ± s.d. of three independent sets of experiments. **p < 0.01, ***p < 0.001 (two-tailed Student’s t-test) Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/30315153), licensed under a CC-BY license. Not internally tested by R&D Systems.

Quercetin inhibits NLRP3 and AIM2 Inflammasomes.LPS (500 ng/ml)-primed BMDM were treated with the indicated flavonoids and concentrations or vehicle (DMSO 0.01%), then stimulated with (A) 5 mM ATP (B) 10 μM Nigericin (C) 130 μg/ml Alum or (D) 400 ng/ml Poly (dA:dT) 30 minutes after treatment. (A–D) IL-1 beta and (E) TNF-alpha concentration in the culture supernatant were measured by ELISA. (B,F,G) LPS-primed BMDM were stimulated with Nigericin treated with quercetin 30 minutes before secondary stimulation. Supernatants and Lysate of BMDM were analyzed by immunoblotting. Cells were stained with FLICA and Caspase-1 activation was determined by FLICA positivity. Data shown are representative of two or more independent experiments (means ± SD) *p < 0.05, **p < 0.01, ***p < 0.001. Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28148962), licensed under a CC-BY license. Not internally tested by R&D Systems.

P. gingivalis and its OMVs differentially induce inflammasome signaling and pyroptosis in human macrophages. MDM were infected as before (2 h at MOI of 25:1, see Materials and Methods) with viable P. gingivalis (Pg), heat-killed-Pg (HK-Pg), OMVs, or heat-inactivated-OMVs (HI-OMVs) and (A) the activation of inflammasome components in the lysates was measured after 24 h by Western blot; beta -actin serves as a loading control throughout. Western blot data are representative of at least three independent experiments. (B) Production of IL-1 beta and IL-18 (by ELISA) from MDM was measured in the supernatant at 24 h. Data are mean ± SD from four independent experiments. Cell viability was determined by measuring extracellular LDH release and by 7-AAD exclusion (by flow-cytometry) at 24 h. 7-AAD is a membrane impermeant dye that is excluded from viable cells. The percent of 7-AAD positive (±SD) cells from three independent experiments was calculated relative to the untreated control. MDM treated with H2O2 (1 mM for 60 min) were included as positive control for 7-AAD. A representative histogram is shown. Cells were also treated where indicated with OMVs in the presence of KYT-1 and KYT-36 (10 μM of each). *p < 0.05: n.s, not significant. Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28824884), licensed under a CC-BY license. Not internally tested by R&D Systems.

P. gingivalis and its OMVs differentially induce inflammasome signaling and pyroptosis in murine macrophages. BMM were infected as before (2 h at MOI of 25:1, see Materials and Methods) with viable P. gingivalis (Pg), heat-killed-Pg (HK-Pg), OMVs, or heat-inactivated-OMVs (HI-OMVs) and (A) the activation of inflammasome components in the lysates [or supernatants (sup) where indicated] measured after 24 h by Western blot; beta -actin serves as a loading control throughout. Western blot data are representative of at least three independent experiments. (B) Production of IL-1 beta and IL-18 (by ELISA) from BMM was measured in the supernatant at 24 h. Data are mean ± SD from four independent experiments. Cell viability was determined by measuring extracellular LDH release and by 7-AAD exclusion (by flow-cytometry) at 24 h. 7-AAD is a membrane impermeant dye that is excluded from viable cells. The percent of 7-AAD positive (±SD) cells from three independent experiments was calculated relative to the untreated control. BMM treated with H2O2 (1 mM for 60 min) were included as positive control for 7-AAD. A representative histogram is shown. Cells were also treated where indicated with OMVs in the presence of KYT-1 and KYT-36 (10 μM of each). *p < 0.05. Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28824884), licensed under a CC-BY license. Not internally tested by R&D Systems.

Caspase-1 but not gasdermin-D processing depends on ASC oligomerization.(a) Western blot analysis for cleaved caspase-1 p20, IL-1 beta p17, and HMGB-1 in cell supernatants (SN) and pro-caspase-1, pro-IL-1 beta and HMGB-1 in cell lysates (lys) of LPS-primed immortalized Asc−/− BMDMs or Asc−/− BMDMs expressing ASCFL, ASCY59A or ASCE80R 3 h after poly(dA:dT) transfection (1 μg ml−1). (b) Release of LDH from LPS-primed immortalized Asc−/− BMDMs or Asc−/− BMDMs expressing ASCFL, ASCY59A or ASCE80R, or derived Casp1 knockouts 3 h after poly(dA:dT) transfection (1 μg ml−1). (c) Western blot analysis for processing of full-length gasdermin-D (GSDMDFL) into the active N-terminal fragment (GSDMDN-term) in combined lysates and supernatants (lys + SN) of LPS-primed immortalized Asc−/− BMDMs expressing ASCFL, ASCY59A or ASCE80R, or derived Casp1 knockouts 3 h after poly(dA:dT) transfection (1 μg ml−1). Arrowhead, gasdermin-DNterm p30; *a cross-reacting band. (d) Release of LDH from LPS-primed primary C57BL/6 WT (WT), Casp1−/−/Casp11−/− or Asc−/− BMDMs infected with S. Typhimurium (multiplicity of infection (MOI)=10, 1 h). (e) Western blot analysis for processing of full-length gasdermin-D (GSDMDFL) into the active N-terminal fragment (GSDMDN-term) in combined lysates and supernatants (lys+SN) of LPS-primed primary C57BL/6 WT (WT), Casp1−/−/Casp11−/− or Asc−/− BMDMs infected with S. Typhimurium (MOI=10, 1 h) or left uninfected. Arrowhead, gasdermin-DNterm p30; *a cross-reacting band; ** a S. Typhimurium-specific cross-reactive band. See also Supplementary Figs 8 and 9. Image collected and cropped by CiteAb from the following publication (//www.nature.com/articles/ncomms11929), licensed under a CC-BY license. Not internally tested by R&D Systems.

Nimbidiol treatment mitigated elevated pro-inflammatory cytokine and chemokine in the diabetic kidney. Representative images from (A) Semi-quantitative RT-PCR analyses showing gene expression and (B) western blot analyses showing protein expression of TNF-alpha , IL-1 beta and MCP-1 in the kidney. The bar diagrams represent the fold change from WT + Saline. Data are mean ± SD (n = 6/group). *p < 0.05 versus WT + Saline, WT + Nimbidiol and Akita + Nimbidiol, †p < 0.05 versus Akita + Saline. Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/36522378), licensed under a CC-BY license. Not internally tested by R&D Systems.

Reactivity:

Mu

Species Glossary

Reactivity: Mu

Applications:

, , , ,

Applications: , , , ,

Host:

Goat Polyclonal

Host: Goat Polyclonal

Sizes:

Sample Size Available

Sizes: Sample Size Available

Conjugate:

Unconjugated

Available Conjugates:

Conjugate	Catalog #	Availability	Size	Price
Biotin	BAF401

Available Formulations

Formulation	Catalog #	Availability	Price
Carrier Free	AF-401-SP

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DY401

Mouse IL-1 beta/IL-1F2 DuoSet ELISA, 15 Plate

(509 Publications)

1 Kit (for 5 Plates)

DY401

Exosomal release of IL-1 alpha by neutrophils(A) Representative confocal images of peritoneal neutrophils stimulated with LPS or curdlan for 6 h.(B) Quantification of IL-1 alpha and CD63 co-localization using ImageJ (each data point represents a single cell).(C) NTA of EV size distribution and concentration.(D–G) Neutrophils were stimulated in the presence of exosome inhibitor GW4869, and IL-1 alpha and IL-1 beta were quantified by ELISA in isolated EVs following lysis (D and F) and in total cell-free supernatants (E and G).(H) Inhibition of EV secretion shown by NTA.(I) Bioactive IL-1 signaling through IL-1R1 reporter cells was measured in isolated exosomes in the absence of detergent lysis (n = 4). Neutralizing antibodies (Abs) to IL-1 alpha , IL-1 beta , or both cytokines were included in the reporter assay, and bioactive cytokine concentration was calculated based on a standard curve using recombinant IL-1 alpha and IL-1 beta .Two-way ANOVA with Tukey’s multiple comparisons test. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. Experiments in (A) and (B) were repeated three times; (C)–(G) are biological replicates from repeat experiments. Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/34010648), licensed under a CC-BY license. Not internally tested by R&D Systems.

Reactivity:

Mu

Species Glossary

Reactivity: Mu

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KGE004B

Prostaglandin E2 Parameter Assay Kit

(418 Publications)

1 Kit

KGE004B

Reactivity:

All-Multi

Species Glossary

Reactivity: All-Multi

Applications:

Applications:

Conjugate	Catalog #	Availability	Size	Price
Biotin	SKGE004B
Biotin	PKGE004B

Available Formulations

Formulation	Catalog #	Availability	Price
Carrier Free	SKGE004B
Carrier Free	PKGE004B

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DY201

Human IL-1 beta/IL-1F2 DuoSet ELISA, 15 Plate

(364 Publications)

1 Kit (for 5 Plates)

DY201

Reactivity:

Hu

Species Glossary

Reactivity: Hu

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NBP2-81747

N-Deacetylase/N-Sulfotransferase 2/NDST2 Antibody - BSA Free

0.1 mg

NBP2-81747

Immunocytochemistry/ Immunofluorescence: N-Deacetylase/N-Sulfotransferase 2/NDST2 Antibody - BSA Free [NBP2-81747] - Immunofluorescence of N-Deacetylase/N-Sulfotransferase 2/NDST2 in Daudi cells with N-Deacetylase/N-Sulfotransferase 2/NDST2 antibody at 20 u/ml.

Reactivity:

Hu, Mu, Rt, Bv

Species Glossary

Reactivity: Hu, Mu, Rt, Bv

Applications:

, ,

Applications: , ,

Host:

Rabbit Polyclonal

Host: Rabbit Polyclonal

Conjugate:

Unconjugated

Available Conjugates:

Conjugate	Catalog #	Availability	Size	Price
Alexa Fluor 350	NBP2-81747AF350
Alexa Fluor 405	NBP2-81747AF405
Alexa Fluor 488	NBP2-81747AF488
Alexa Fluor 532	NBP2-81747AF532
Alexa Fluor 594	NBP2-81747AF594
Alexa Fluor 647	NBP2-81747AF647
Alexa Fluor 700	NBP2-81747AF700
Alexa Fluor 750	NBP2-81747AF750
Biotin	NBP2-81747B
CoraFluor 1	NBP2-81747CL1
DyLight 350	NBP2-81747UV
DyLight 405	NBP2-81747V
DyLight 488	NBP2-81747G
DyLight 550	NBP2-81747R
DyLight 594	NBP2-81747DL594
DyLight 650	NBP2-81747C
DyLight 680	NBP2-81747FR
DyLight 755	NBP2-81747IR
FITC	NBP2-81747F
HRP	NBP2-81747H
Janelia Fluor 525	NBP2-81747JF525
Janelia Fluor 549	NBP2-81747JF549
Janelia Fluor 585	NBP2-81747JF585
Janelia Fluor 635	NBP2-81747JF635
Janelia Fluor 646	NBP2-81747JF646
Janelia Fluor 669	NBP2-81747JF669
PerCP	NBP2-81747PCP
mFluor Violet 450 SE	NBP2-81747MFV450
mFluor Violet 500 SE	NBP2-81747MFV500
mFluor Violet 610 SE	NBP2-81747MFV610

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7689-ST

Recombinant Human NDST2 Protein, CF

10 μg

Reactivity:

Hu

Species Glossary

Reactivity: Hu

Applications:

Applications:

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NBP2-13645

N-Deacetylase/N-Sulfotransferase 2/NDST2 Antibody

0.1 ml

NBP2-13645

Reactivity:

Hu, Mu, Rt

Species Glossary

Reactivity: Hu, Mu, Rt

Applications:

,

Applications: ,

Host:

Rabbit Polyclonal

Host: Rabbit Polyclonal

Sizes:

Sample Size Available

Sizes: Sample Size Available

Conjugate:

Unconjugated

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NBP2-83260

N-Deacetylase/N-Sulfotransferase 2/NDST2 Antibody

0.1ml

NBP2-83260

Reactivity:

Hu

Species Glossary

Reactivity: Hu

Applications:

Applications:

Host:

Rabbit Polyclonal

Host: Rabbit Polyclonal

Conjugate:

Unconjugated

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NBP2-07972

N-Deacetylase/N-Sulfotransferase 2/NDST2 Overexpression Lysate

0.1 mg

NBP2-07972

Reactivity:

Hu

Species Glossary

Reactivity: Hu

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Applications:

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NBP2-13645PEP

N-Deacetylase/N-Sulfotransferase 2/NDST2 Recombinant Protein Antigen

0.1 ml

Reactivity:

Hu

Species Glossary

Reactivity: Hu

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NB100-417

Carbonic Anhydrase IX/CA9 Antibody - BSA Free

(7 Reviews)

(279 Publications)

0.1 ml

NB100-417

CA9 expressed in hypoxic regions as assessed by pimonidazole staining (A), but also observed in non-pimonidazole areas (B). Most of CA9 expressed in pimonidazole positive regions (C). Proliferation (BrdUrd) & apoptosis (caspase-3) in relation to CA9 expression shown in figure D & E. Red, CA9; Green, pimonidazole (A-B), BrdUrd (D) or caspase-3 (E); Yellow, overlap of CA9 (red) & pimonidazole (green); Light blue, vessels. Magnification 100x. Scale bars represent 100 um. Closed circles represent CA9 expression in pimonidazole positive regions; open circles represent CA9 expression in pimonidazole -ve regions. Image collected & cropped by CiteAb from the following publication (//dx.plos.org/10.1371/journal.pone.0108068), licensed under a CC-BY license.

<b>Orthogonal Strategies Validation. </b>Formalin-fixed paraffin-embedded tissue sections of human stomach were probed for Carbonic Anhydrase IX/CA9 mRNA (ACD RNAScope Probe, catalog # 559348; Fast Red chromogen, ACD catalog # 322750). Adjacent tissue section was processed for immunohistochemistry using Rabbit Polyclonal (Novus catalog # NB100-417) at 1:1000 dilution with overnight incubation at 4 degrees Celsius followed by incubation with anti-rabbit IgG VisUCyte HRP Polymer Antibody (Catalog # VC003) and DAB chromogen (yellow-brown). Tissue was counterstained with hematoxylin (blue). Specific staining was localized to glandular cells.

Analysis on various human cell lysates. Specific bands were detected for Carbonic Anhydrase IX/CA9 in HEK-293 and A498 cell lines at a molecular weight of approximately 50 kDa. WB image submitted by a verified customer review.

Immunohistochemical representative microphotographs representing the HIF-1alpha, GLUT-1, and CAIX expression in endometrial cancer according to FIGO classification (IA, IB, II, IIIA, IIIC, and IV). Primary objective magnification 20x. Image collected and cropped by CiteAb from the following publication (//www.hindawi.com/journals/bmri/2014/616850/) licensed under a CC-BY license.

<b>Biological Strategies Validation. </b>Hypoxias impact on proteins of the mitochondrial ISC assembly machinery. (A) Immunoblotting analyzed the total protein extracts from HeLa cells grown in normoxia (Nx, 21% O2) or hypoxia (Hx, 1% O2) conditions using VDACs poly antibody and anti-CAIX, -ISCU, -FXN, -NFS1, -HSC20 antibodies. Beta-Actin was used as loading control. Citation: Ferecatu I, Canal F, Fabbri L, Mazure NM, Bouton C, Golinelli-Cohen M-P (2018) Dysfunction in the mitochondrial Fe-S assembly machinery leads to formation of the chemoresistant truncated VDAC1 isoform without HIF-1 alpha activation. PLoS ONE 13(3): e0194782.

<b>Biological Strategies Validation. </b>Analysis using the DyLight 488 conjugate of NB100-417. Staining of Carbonic Anhydrase IX (red) in human glioma U87 cells. DAPI counterstains nuclei (blue). ICC/IF image submitted by a verified customer review.

Simple Western lane view shows a specific band for CAIX in 0.1 mg/mL of HeLa lysate. This experiment was performed under reducing conditions using the 12-230 kDa separation system.

An intracellular stain was performed on U-87 MG Cells with NB100-417 and a matched isotype control. Cells were fixed with 4% PFA and then permeablized with 0.1% saponin. Cells were incubated in an antibody dilution of 2.5 ug/mL for 30 minutes at room temperature, followed by Rabbit IgG APC-conjugated Secondary Antibody, (R&D Systems, F0111).

Western Blot analysis of Carbonic Anhydrase IX/CA9 Antibody on human breast cancer MCF7 cells. Image from verified customer review.

A431 cells were fixed in 4% paraformaldehyde for 10 minutes and permeabilized in 0.05% Triton X-100 in PBS for 5 minutes. The cells were incubated with anti-Carbonic Anhydrase IX/CA9 Antibody NB100-417 at 2 ug/ml overnight at 4C and detected with an anti-rabbit Dylight 488 (Green) at a 1:1000 dilution for 60 minutes. Nuclei were counterstained with DAPI (Blue). Cells were imaged using a 100X objective and digitally deconvolved.

An intracellular stain was performed on A431 cells with Carbonic Anhydrase IX/CA9 Antibody NB100-417 (blue) and a matched isotype control (orange). Cells were fixed with 4% PFA and then permeabilized with 0.1% saponin. Cells were incubated in an antibody dilution of 1.0 ug/mL for 30 minutes at room temperature, followed by Rabbit IgG (H+L) Cross-Adsorbed Secondary Antibody, Dylight 550 (SA5-10033, Thermo Fisher).

Analysis in 1) HeLa, 2) MDA-MB-231, and 3) A549 whole cell lysates. Specific bands were detected for Carbonic Anhydrase IX/CA9 at a molecular weight of 50 kDa.

Analysis on rat renal cortex. A specific band was detected at a molecular weight of approximately 50 kDa.

Immunocytochemical analysis of CNHCs with antibodies against the RCC marker CAIX. Clusters of CNHCs cytomorphologically classified as uncertain malignant (-UMF) with cytoplasmic positive staining with antibodies against the RCC marker CAIX. Image collected and cropped by CiteAb from the following publication (//translational-medicine.biomedcentral.com/articles/10.1186/1479-5876-11-214) licensed under a CC-BY license.

Analysis of human colon carcinoma, xenografted in mice. IHC-Fr image submitted by a verified customer review.

IHC analysis of a FFPE tissue section of human breast cancer using CAIX antibody at 1:1000 dilution. The primary antibody bound to CAIX antigens in the tissue section was detected using a HRP labeled secondary antibody and DAB reagent. Nuclei of the cells were counterstained with hematoxylin. This CAIX antibody generated an expected cytoplasmic staining of CAIX protein with an intense signal around the cellular membranes in tumor cores. The latter are more likely to be hypoxic in growing tumors which signifies that the observed CAIX staining is specific.

Strong cytoplasmic and membranous CA9 staining in glandular cells of human stomach tissue. Citrate buffer pH 6 antigen retrieval. Antibody at 1:250 dilution and detected with an Alexa Fluor 546 labeled goat-anti-rabbit secondary antibody. Nuclei detected with DAPI. IHC-P image submitted by a verified customer review.

Immunofluorescence of human RCC tumor cryosections using NB100-417 (Panel A). Panel B shows staining with normal rabbit serum.

Colorectal cancer xenograft growth was suppressed by CB-PIC(20 and 50 mg/kg body weight) in female athymic nude mice. Starting three days after SW620 cell inoculation, CB-PIC (20 and 50 mg/kg body weight) was injected in abdomen with 4% Tween 20 as vehicle once daily. Representative examples of immunohistochemical staining for CA IX in tumor section. Image collected and cropped by CiteAb from the following publication (//www.hindawi.com/journals/ecam/2013/974313/), licensed under a CC-BY license.

<b>Genetic Strategies Validation. </b>Iron depletion and nitric oxide stress and Iron depletion induce the truncated VDAC1 form accumulation. (D) Total protein extracts were analyzed by immunoblotting using VDACs poly antibody and anti-HIF-1 alpha and -CAIX antibodies. (E) Immunoblotting was used to analyze total protein extracts using antibodies against the three VDAC isoforms. (F) HeLa cells were grown in hypoxia (Hx, 1% O2) conditions and transfected with iscu- or NC-siRNA for 3 days, or grown in normoxia (Nx, 21% O2), some treated with DFO for 16 h. Western Blots analyzed total proteins using VDACs poly antibody and anti-HIF-1 alpha, -CAIX, -ISCU antibodies. Citation: Ferecatu I, Canal F, Fabbri L, Mazure NM, Bouton C, Golinelli-Cohen M-P (2018) Dysfunction in the mitochondrial Fe-S assembly machinery leads to formation of the chemoresistant truncated VDAC1 isoform without HIF-1 alpha activation. PLoS ONE 13(3): e0194782.

The images are representative of immunohistochemistry staining of nuclear staining of proliferation marker Ki67, membrane protein CD 44, and CA IX. The images of scaffold culture (left panel) are compared with cell pellet paraffin section (right panel). The scaffold dissolved in xylene presented as a clear region marked ∗. All scale bars are 100 μm, magnification 40x.

Chemosensitivity of the CTC SCLC lines (IC50, mean ± SD) and immunohistochemistry of sections of tumorospheres. All differences between single cells (SC) and tumorospheres (TS) are statistically significant. Immunohistochemical staining of sections of UHGc5, BHGc16 and BHGc26 CTCs was performed using antibodies directed to CD56, CHGA, CAIX and Ki67, respectively.

Western Blot: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - Analysis of protein levels of genes with increased transcription in hypoxia, IOX2 & VH032.HIF targets were increased in hypoxia, VH298 & FG-4592. 0.05% DMSO (vehicle control), 1% O2 (hypoxia), 100 µM VH298 & 50 µM FG-4592 were introduced to (A) HeLa or HFF for 24 hours & (B) HeLa for indicated time. Protein levels were analysed by immunoblotting using antibodies against indicated proteins, with beta -Actin as loading control. The blots shown are representative of three independent experiments. * indicates longer exposure. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/30801039), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunohistochemistry-Paraffin: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - IGF1R & CA9 expression in PDAC.(A) Representative staining of CA9 cells quantified on a scoring system of 0–3 according to staining intensity (original magnification ×200). CA9 was mainly expressed in the cell membrane of PDAC cells. (B) IGF1R was mainly expressed in the cytoplasm of PDAC cells. (C) Association between IGF1Rexpression & CA9 expression in 120 pancreas cancer. A significant correlation was found between IGF1R expression level was significantly correlated with CA9 expression in PDAC cells (p < 0.01, rs = 0.649; Spearman’s rank sum test). The size of circle indicates the number of PDAC patients. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/27487118), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunohistochemistry: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - Representative human colorectal cancer specimen. Immunohistochemistry of CA9 & GGT1 in mLNs. In these cases, both CA9 & GGT1 were expressed, suggesting that GGT1 expression is related to hypoxia in mLNs. Scale bar, 500 μm. Black arrowheads: GGT of cancer cells, white arrowheads: GGT surrounding cancer cells, arrow: accumulation of GGT. mLNs: metastatic lymph nodes, GGT: gamma-glutamyl transpeptidase. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/30542087), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunohistochemistry: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - Representative microscopy images of staining for hypoxia markers in prostate tissues (MO, 400×). A) HIF-1 alpha - notice the granular cytoplasmic immunoreactivity of the malignant epithelial cells. In this case, more than 50% of the glands stained. B) LOX - strong & diffuse nuclear immunoreactivity of the epithelial cells. C) CAIX - note a focal apical cytoplasmic immunoreactivity in epithelial cells. D) VEGFR2 - moderate nuclear & weak cytoplasmic expression of the epithelial cells Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28143503), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunohistochemistry: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - The images are representative of immunohistochemistry staining of nuclear staining of proliferation marker Ki67, membrane protein CD 44, & CA IX. The images of scaffold culture (left panel) are compared with cell pellet paraffin section (right panel). The scaffold dissolved in xylene presented as a clear region marked ∗. All scale bars are 100 μm, magnification 40x. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/24101930), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunohistochemistry: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - Hypoxia, blood vessel & proliferation staining. IHC staining in the Biopsy 23 & Biopsy 3 respectively for CA-9, EF5, CD31, Ki67 & LYVE1. 15× Magnification. Image collected & cropped by CiteAb from the following publication (//www.mdpi.com/2072-6694/4/3/821), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - VH298 elicits a HIF-dependent hypoxic response in various cell lines.1% DMSO, hypoxia (1% O2) & indicated compounds at 100 μM were introduced to (a) HFF, HeLa, U2OS & (b) RCC4 cells expressing HA-tagged VHL (with the exception of cis-VH298 & VH298 used at 150 μM) for 24 h & (c) CTLs for 8 h. Protein levels were analysed by immunoblotting using antibodies against the indicated proteins, with beta -actin (SMC1 for CTLs) as the loading control. The blots shown are representative of three independent experiments. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/27811928), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunohistochemistry: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - HE staining & immunohistochemistry (CA9 & GGT1) of fixed primary tumor & mLN specimens from orthotopic mouse model. CA9 & GGT1 were expressed inside the primary tumor & in mLNs. GGT1 was accumulated in the area of central necrosis. HE: hematoxylin-eosin. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/30542087), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunohistochemistry: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - Comparison between primary tumor & mLNs of the mouse model. Expression of CA9 & GGT1 in mLNs was higher than that in the primary tumor at 1week after injection of cells. (A) CA9 & GGT1 expression of HT29 mLNs compared to primary tumor at 6 weeks or 1 week after injection of cells. (B) Tissue oxygen concentration (%) of rectal submucosa (SM) & nLNs (n = 3). Error bars represent SD. mNs: metastatic lymph nodes, nLNs: non metastatic lymph nodes. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/30542087), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunohistochemistry: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - Chemosensitivity of the CTC SCLC lines (IC50, mean ± SD) & immunohistochemistry of sections of tumorospheres. All differences between single cells (SC) & tumorospheres (TS) are statistically significant. Immunohistochemical staining of sections of UHGc5, BHGc16 & BHGc26 CTCs was performed using antibodies directed to CD56, CHGA, CAIX & Ki67, respectively. Image collected & cropped by CiteAb from the following publication (//www.nature.com/articles/s41598-017-05562-z), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunocytochemistry/ Immunofluorescence: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - Immunocytochemical analysis of CNHCs with antibodies against the RCC marker CAIX. Clusters of CNHCs cytomorphologically classified as uncertain malignant (−UMF) with cytoplasmic positive staining with antibodies against the RCC marker CAIX (A). Clusters of CNHC-UMF & -BF without reactivity for CAIX antibodies (B & C, respectively). A single CNHC-MF with positive cytoplasmic (D) & without staining for CAIX (E). Single CAIX-negative CNHC-UMF & -BF (F & G, respectively). Image collected & cropped by CiteAb from the following publication (//translational-medicine.biomedcentral.com/articles/10.1186/1479-5876-11-214), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - GPRC5A promotes hypoxic cell survival via YAPAHypoxia induced increased total YAP expression in a panel of colorectal cancer cell lines. Actin confirmed equal loading; blots are representative of at least two independent experiments. Related to Fig 4.B–EqRT–PCR analysis revealed that hypoxia increased the expression of known YAP target genes AREG, BCL2L1 (BCL‐XL), CTGF & CYR61. Representative experiments are shown. Representative examples of n = 3 independent experiments are shown; data are presented as mean ± SD.FIncreased expression of YAP Ser397 in GPRC5A‐depleted cells was overridden by expression of constitutively active RhoA (G14V) in hypoxia. Expression of dominant negative RhoA (T19N) did not further increase YAP Ser397 phosphorylation in response to GPRC5A depletion.GDarker exposure of the GPRC5A blot in Fig 4I, confirming expression of 30–40 kDa & 80 kDa species. Note that YAP siRNA partially diminishes GPRC5A in line with the existence of positive HIF‐GPRC5A‐YAP feedback loop. Asterisk (*) indicates non‐specific band.Source data are available online for this figure. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/30143543), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunohistochemistry: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - CAIX expression in PDAC & association with outcomeA. Representative images of immunohistochemical staining showing: negative expression of CAIX in both tumor cell & stromal compartments (top left), positive expression of CAIX in tumor cells but not stromal compartment (top right), positive expression of CAIX in stromal compartment but not tumor cells (bottom left), positive expression of CAIX in both tumor cell & stromal compartments (bottom right). B. Kaplan-Meier analysis of CAIX expression in the distinct tumor cell & stromal compartments & the associated overall survival. C. Representative images of immunohistochemical staining showing low stromal volume & high stromal volume & low & high microvessel density D. Representative images of immunohistochemical staining showing: positive expression of MCT4 in tumor cell compartment & positive expression of MCT4 in tumor stromal compartment. Kaplan-Meier analysis of combinatorial MCT4 & CAIX expression in the stromal compartment & the associated overall survival. Hazard ratios & p-values for each comparison are summarized in the table. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/27623078), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunocytochemistry/ Immunofluorescence: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - Hypoxia & autophagy markers in breast cancer stem-cells(A) Co-expression of CD133 & BECLIN1 in pCR & non-pCR patients. Double IF X800. (B) Tumor cells (T) distributed around necrosis (N) express LC3B with typical cytoplasmic punctuated staining, ×1000. Double IF shows that some CD133-positive tumor cells co-express LC3B. ×400. (C) Tumor cells (T) distributed around necrosis (N) express CAIX, on their cytoplasmic membrane, ×1000. Double IF shows a co-expression of LC3B & CAIX in some tumor cells. ×400. (D) Laser microdissection of CD133-positive cells. (E) Laser-microdissected CD133-positive cells express PROMININ & not PTPRC (CD45). (F) mRNA relative quantification of HIF1A, CAIX, LC3B, BECN1, & BNIP3L show a significantly higher expression level in CD133-positive cells versus CD133-negative cells, in the 35 non-pCR biopsies. *p < 0.05. (G) Multivariate analysis of tumor cell characteristics & pCR shows an inverse correlation between pCR & cells bearing markers of stemness or autophagy. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28445132), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunocytochemistry/ Immunofluorescence: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - Immunofluorescent images showing CAIX expression, pimonidazole, apoptosis & proliferation in SCCNij202.CAIX is expressed in hypoxic regions as assessed by pimonidazole staining (A), but is also observed in non-pimonidazole areas (B). Yet, most of CAIX is expressed in pimonidazole positive regions (C). Proliferation (BrdUrd) & apoptosis (caspase-3) in relation to CAIX expression are shown in figure D & E respectively. Red, CAIX; Green, pimonidazole (A–B), BrdUrd (D) or caspase-3 (E); Yellow, overlap of CAIX (red) & pimonidazole (green); Light blue, vessels. Magnification 100×. Scale bars represent 100 µm. Closed circles represent CAIX expression in pimonidazole positive regions; open circles represent CAIX expression in pimonidazole negative regions. Abbrevations: 1× S4, 8 h, one i.p. injection S4, harvest after 8 hours; 1× S4, 24 h, one i.p. injection S4, harvest after 24 hours; 3× S4, one i.p. injection S4 a day for 3 days, harvest 8 hours after the last injection; 5× S4, one i.p. injection S4 a day for 5 days, harvest 8 hours after the last injection. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/25225880), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunohistochemistry: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - Colorectal cancer xenograft growth was suppressed by CB-PIC(20 & 50 mg/kg body weight) in female athymic nude mice. Starting three days after SW620 cell inoculation, CB-PIC (20 & 50 mg/kg body weight) was injected in abdomen with 4% Tween 20 as vehicle once daily. (a) Body weights of mice. (b) Tumor growth in a time course. (c) Final tumor weight at termination of experiment. (d) Representative examples of immunohistochemical staining for Ki-67, CD34, TUNEL, CA IX & pAMPK alpha in tumor sections. Graphs show the Ki67 index (proliferation), CD34 index (angiogenesis), TUNEL index (apoptosis), CA IX (hypoxia region), & pAMPK alpha index in tumor sections. Values are means ± SD, n = 6. *P < 0.05 & ***P < 0.001 compared with control mice. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/23589723), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - Hypoxia induces GPRC5A directly via HIFsASILAC‐based proteomics data identify known (red) & novel (green) hypoxia‐induced proteins in SW620 cells. One‐sample t‐test was performed.BWestern blotting confirmed GPRC5A as a hypoxia‐induced protein in SILAC lysates.CValidation of GPRC5A Western blot data using siRNA. *Non‐specific band of ˜60 kDa not depleted by GPRC5A siRNA.DConfocal microscopy showing plasma membrane GPRC5A expression in hypoxic SW620 cells (scale bars: 75 μm).EWestern blotting showing GPRC5A upregulation by hypoxia in a panel of colorectal tumour cell lines.FBasal & hypoxia‐induced GPRC5A protein expression was decreased by HIF‐1/2 alpha depletion.GDepletion of HIF‐1 beta decreased GPRC5A protein upregulation in hypoxia.HHypoxia mimetic DMOG induced HIF‐1/2 alpha , CA9 & GPRC5A protein expression. Dual HIF‐1/2 alpha depletion reduced GPRC5A induction by DMOG.IqRT–PCR demonstrating that GPRC5A mRNA was upregulated by hypoxia (n = 3). GPRC5A was normalised to HPRT (error bars ± SD).JqRT–PCR demonstrating that HIF‐1/2 alpha depletion decreased GPRC5A induction during hypoxia (n = 3). GPRC5A was normalised to HPRT (error bars ± SD).KChIP‐PCR analyses identify HIF‐1 alpha binding to the GPRC5A promoter region containing a putative optimal HRE (error bars ± SD, n = 3).Data information: Asterisks (*) indicate non‐specific band. Level adjustments were made to images in Adobe Photoshop post‐acquisition for clarity (equal changes applied to the entire image). Representative examples of n = 3 independent experiments are shown.Source data are available online for this figure. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/30143543), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: Carbonic Anhydrase IX/CA9 Antibody - BSA Free [NB100-417] - Establishment of a human protein replacement stable cell line using 47snoMEN.(a) Images of protein replacement stable cell lines (U2OSGFP–HIF-1 alpha -47PR). Expression of FP proteins was confirmed by fluorescence imaging with DFX treatment. Bar is 10 μm. (b) U2OS & U2OSGFP–HIF-1 alpha -47PR cells were exposed to 1% O2 for the indicated periods of time prior to lysis. Cell extracts were analysed by western blot using the indicated antibodies. (c) U2OS cells were transfected with 1 μg of control & HIF-1 alpha -47snoMEN vectors for 48 hours prior to lysis. DFX was added for the last 24 hours. Cell extracts were analysed by western blot using the indicated antibodies. (d) 3D culture analysis. HeLaGFP–HIF-1 alpha -47PR cells were cultured in lipidure coated plates to induce spheroid formation. HIF-1 alpha is visible in the core of the spheroid revealing hypoxia. Bar is 10 μm. Image collected & cropped by CiteAb from the following publication (//dx.plos.org/10.1371/journal.pone.0154759), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Reactivity:

Hu, Mu, Rt, Pl

Species Glossary

Reactivity: Hu, Mu, Rt, Pl

Applications:

, , , , , , , , , , , ,

Applications: , , , , , , , , , , , ,

Host:

Rabbit Polyclonal

Host: Rabbit Polyclonal

Sizes:

Sample Size Available

Sizes: Sample Size Available

Conjugate:

Unconjugated

Available Conjugates:

Validated by:

Validated by:

Conjugate	Catalog #	Availability	Size	Price
Alexa Fluor 350	NB100-417AF350
Alexa Fluor 405	NB100-417AF405
Alexa Fluor 488	NB100-417AF488
Alexa Fluor 532	NB100-417AF532
Alexa Fluor 594	NB100-417AF594
Alexa Fluor 647	NB100-417AF647
Alexa Fluor 700	NB100-417AF700
Alexa Fluor 750	NB100-417AF750
Allophycocyanin	NB100-417APC
Allophycocyanin/Cy7	NB100-417APCCY7
Biotin	NB100-417B
CoraFluor 1	NB100-417CL1
DyLight 350	NB100-417UV
DyLight 405	NB100-417V
DyLight 488	NB100-417G
DyLight 550	NB100-417R
DyLight 594	NB100-417DL594
DyLight 650	NB100-417C
DyLight 680	NB100-417FR
DyLight 755	NB100-417IR
FITC	NB100-417F
HRP	NB100-417H
Janelia Fluor 525	NB100-417JF525
Janelia Fluor 549	NB100-417JF549
Janelia Fluor 585	NB100-417JF585
Janelia Fluor 635	NB100-417JF635
Janelia Fluor 646	NB100-417JF646
Janelia Fluor 669	NB100-417JF669
PE	NB100-417PE
PE/Atto594	NB100-417PEATT594
PE/Cy5.5	NB100-417PECY55
PE/Cy7	NB100-417PECY7
PerCP	NB100-417PCP
mFluor Violet 450 SE	NB100-417MFV450
mFluor Violet 500 SE	NB100-417MFV500
mFluor Violet 610 SE	NB100-417MFV610

Available Formulations

Formulation	Catalog #	Availability	Price
BSA Free	NB100-417SS

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NBP3-16022

N4BP2 Antibody - Azide and BSA Free

100 ul

NBP3-16022

Reactivity:

Hu, Rt

Species Glossary

Reactivity: Hu, Rt

Applications:

,

Applications: ,

Host:

Rabbit Polyclonal

Host: Rabbit Polyclonal

Conjugate:

Unconjugated

Available Formulations

Formulation	Catalog #	Availability	Price
Azide and BSA Free	NBP3-16022-20ul

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At	=	A. thaliana	SyHa	=	Golden Syrian Hamster
All	=	All Species	Ha	=	Hamster
An	=	Animal	Hu	=	Human
ArHa	=	Armenian Hamster	I	=	Insect
Av	=	Avian	Ll	=	Llama
Bb	=	Baboon	Ma	=	Mammal
Ba	=	Bacteria	Mk	=	Monkey
Bv	=	Bovine	Mu	=	Mouse
Ca	=	Canine	Multi	=	Multi-species
Ce	=	C. Elegans	NA	=	Non-species specific
Ch	=	Chicken	Or	=	Orangutan
ChHa	=	Chinese Hamster	Pl	=	Plant
Cm	=	Cynomolgus Monkey	Pm	=	Primate
Do	=	Donkey	Po	=	Porcine
Dr	=	Drosophilia	Rb	=	Rabbit
Ec	=	E. Coli	Rt	=	Rat
Eq	=	Equine	RM	=	Rhesus Macaque
Fe	=	Feline	Sh	=	Sheep
Ft	=	Ferret	Vb	=	Vertebrate
Fi	=	Fish	Vi	=	Virus
Fu	=	Fungi	Xp	=	Xenopus
Ge	=	Gerbil	Ye	=	Yeast
GP	=	Guinea Pig	Ze	=	Zebrafish
Gt	=	Goat

Category

Protein/Target

Applications

Reactivity

Citation

Antibody Validation

Reviews

Host

Available Conjugates

Format

Marker

Modification

Clonality

Clone

Gene Symbol

Images Type

Isotype

Kit Type

Lysate Type

Preservative

Protein Or Peptide

Research Areas

Has Sample Size

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