Novex ecl chemi substrate

Manufactured by Thermo Fisher Scientific

Sourced in United States

The NOVEX ECL Chemi Substrate is a chemiluminescent substrate used for the detection of proteins in Western blotting applications. It provides a sensitive and reliable method for visualizing protein bands on a membrane.

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Lab products found in correlation

Ab5535, by Merck Group (4 mentions) β actin, by Merck Group (4 mentions) Ab5603, by Merck Group (4 mentions) Hrp linked antibodies, by Santa Cruz Biotechnology (2 mentions) A5441, by Merck Group (2 mentions) Phospho s6 ribosomal protein, by Cell Signaling Technology (1 mentions) Hoechst 33342, by Merck Group (1 mentions) E cadherin, by BD (1 mentions) N cadherin, by BD (1 mentions) Vimentin, by Agilent Technologies (1 mentions) Ab59252, by Abcam (1 mentions) P stat3, by Cell Signaling Technology (1 mentions) Horseradish peroxidase coupled secondary antibody, by GE Healthcare (1 mentions) P jak1, by Cell Signaling Technology (1 mentions) Erk2 sc 1647, by Santa Cruz Biotechnology (1 mentions) P stat1, by Cell Signaling Technology (1 mentions) Ab32064, by Abcam (1 mentions) Ab24610, by Abcam (1 mentions) Ab52866, by Abcam (1 mentions) Nb500 171, by Novus Biologicals (1 mentions)

Spelling variants of this product

ECL substrate (363 citations) Novex ECL (22 citations)

10 protocols using novex ecl chemi substrate

Protein Expression Analysis in Cells

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Immunoblot and immunofluorescence analysis were performed as previously described18 (link). Primary antibodies used were: SOX9 (AB5535, Millipore), phospho-SOX9 (ab59252, abcam), BMI1 (05-637, Millipore), E-cadherin (BD610181, BD Transduction Laboratories), Vimentin (M7020, DAKO), N-Cadherin (BD610920, BD Transduction Laboratories), phospho-S6 Ribosomal protein (Cell Signaling Technology^®, #4858) and β-actin (AC-15, Sigma). For Western blot detection of primary antibodies, we used HRP-linked antibodies (Santa Cruz Biotechnology) and detection was performed by chemiluminescence using NOVEX ECL Chemi Substrate (ThermoFisher). For immunofluorescence, secondary antibodies conjugated with fluorochromes were used and nuclear DNA was stained with Hoechst 33342 (Sigma). Images were obtained at a 40x magnification.

Carrasco-Garcia E., Lopez L., Aldaz P., Arevalo S., Aldaregia J., Egaña L., Bujanda L., Cheung M., Sampron N., Garcia I, & Matheu A. (2016). SOX9-regulated cell plasticity in colorectal metastasis is attenuated by rapamycin. Scientific Reports, 6, 32350.

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Western Blotting of JAK-STAT Signaling

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Cells were lysed using RIPA buffer and analysed by western blotting as described^{56 (link)}. Primary antibodies used were: pJAK1(Tyr1034/1035; 74129T, 1:500 dilution), pJAK1(Tyr1022/1023; 3331S, 1:500 dilution), pSTAT1(Tyr701; 7649T, 1:500 dilution), pSTAT3(Ser727; 9134T, 1:500 dilution) from Cell Signaling and ERK2 (sc-1647) from Santa Cruz Biotechnology. Detection was through enhanced chemiluminescence ECL using horseradish peroxidase-coupled secondary antibodies (1:2,000 dilution; GE Healthcare) and NOVEX ECL Chemi Substrate (Thermo Fisher).

Redondo-Muñoz M., Rodriguez-Baena F.J., Aldaz P., Caballé-Mestres A., Moncho-Amor V., Otaegi-Ugartemendia M., Carrasco-Garcia E., Olias-Arjona A., Lasheras-Otero I., Santamaria E., Bocanegra A., Chocarro L., Grier A., Dzieciatkowska M M., Bigas C., Martin J., Urdiroz-Urricelqui U., Marzo F., Santamaria E., Kochan G., Escors D., Larrayoz I.M., Heyn H., D’Alessandro A., Attolini C.S., Matheu A., Wellbrock C., Benitah S.A., Sanchez-Laorden B, & Arozarena I. (2023). Metabolic rewiring induced by ranolazine improves melanoma responses to targeted therapy and immunotherapy. Nature Metabolism, 5(9), 1544-1562.

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Immunoblot Analysis of Protein Expression

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Immunoblots were performed following standard procedures^{24 (link)}. Specific antibodies against HDAC6 (7558 S Cell Signalling), acetyl-α-tubulin (ab24610, Abcam), α-tubulin (ab52866, Abcam), acetyl-histone H3-lys9 (9649, Cell Signaling), histone H3 (NB500-171, Novus), PARP (ab32064, Abcam), BMI-1 (05-637, Millipore), SOX2 (AB5603, Millipore), SOX9 (AB5535, Millipore), and β-actin (A5441, Sigma) were used in the study. For secondary antibodies, horseradish peroxidase (HRP)-linked anti-rabbit (7074S, Cell Signalling) or anti-mouse (7076S, Cell Signalling) were used. Detection was performed by chemiluminiscence using NOVEX ECL Chemi Substrate (ThermoFisher).

Auzmendi-Iriarte J., Saenz-Antoñanzas A., Mikelez-Alonso I., Carrasco-Garcia E., Tellaetxe-Abete M., Lawrie C.H., Sampron N., Cortajarena A.L, & Matheu A. (2020). Characterization of a new small-molecule inhibitor of HDAC6 in glioblastoma. Cell Death & Disease, 11(6), 417.

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Protein Expression Analysis by Immunoblotting

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Immunoblots were performed following standard procedures. The antibodies for detection of SOX2 (AB5603 Millipore), SOX9 (AB5535 Millipore), p-p38 (9211 Cell Signalling), p38 (sc-7972 Santa Cruz), p-JNK (9251 Cell Signalling), JNK (sc-474 Santa Cruz), p-ERK1/2 (9101 Cell Signalling), ERK1/2 (9102 Cell Signalling), and ß-actin (AC-15 Sigma) were used in the study. For secondary antibodies, we used horseradish peroxidase (HRP)-linked anti-rabbit or anti-mouse (SantaCruz Biotechnology), both at a 1:2000 dilution. Detection was performed by chemiluminescence using NOVEX ECL Chemi Substrate (ThermoFisher).

Arrizabalaga O., Moreno-Cugnon L., Auzmendi-Iriarte J., Aldaz P., Ibanez de Caceres I., Garros-Regulez L., Moncho-Amor V., Torres-Bayona S., Pernía O., Pintado-Berninches L., Carrasco-Ramirez P., Cortes-Sempere M., Rosas R., Sanchez-Gomez P., Ruiz I., Caren H., Pollard S., Garcia I., Sacido A.A., Lovell-Badge R., Belda-Iniesta C., Sampron N., Perona R, & Matheu A. (2017). High expression of MKP1/DUSP1 counteracts glioma stem cell activity and mediates HDAC inhibitor response. Oncogenesis, 6, 401.

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Western Blot Analysis of SOX Proteins

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Immunoblots were performed following standard procedures. For SOX2 detection, AB5603 antibody (Millipore, Billerica, MA, USA) was used, for SOX9 AB5535 antibody (Millipore), and for β-actin AC-15 (Sigma). HRP-linked (Horseradish Peroxidase) anti-rabbit or anti-mouse (SantaCruz Biotechnology, Dallas, TX, USA) secondary antibodies, both at a 1:2000 dilution, were used. Detection was performed by chemiluminescence using NOVEX ECL Chemi Substrate (ThermoFisher, Waltham, MA, USA).

Garros-Regulez L., Aldaz P., Arrizabalaga O., Moncho-Amor V., Carrasco-Garcia E., Manterola L., Moreno-Cugnon L., Barrena C., Villanua J., Ruiz I., Pollard S., Lovell-Badge R., Sampron N., Garcia I, & Matheu A. (2016). mTOR inhibition decreases SOX2-SOX9 mediated glioma stem cell activity and temozolomide resistance. Expert Opinion on Therapeutic Targets, 20(4), 393-405.

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Immunoblot Analysis of Cellular Proteins

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Immunoblots were performed following standard procedures (25 (link)). Specific antibodies against LAMP2A (ab18528, Abcam, 1/500), SOX2 (AB5603, Millipore, 1/250), SOX9 (AB5535, Millipore, 1/1,000), TIM23 (11123–1-AP, Proteintech, 1/1,000), MRSP23 (SAB2701383, Sigma, 1/500), p-STAT1 (9167S, Cell Signaling, 1/1,000), STAT1 (14994S, Cell Signaling, 1/1,000), MX1 (ab95926, Abcam, 1/500), p-STAT3 (9145S, Cell Signaling, 1/1,000), STAT3 (9139, Cell Signaling, 1/1,000), ITGA6 (3750S, Cell Signaling, 1/250), ITGB4 (HPA036348, Sigma, 1/500), p-AKT (9271, Cell Signaling, 1/1,000), AKT (sc-8312, Santa Cruz Biotechnology, 1/200), and β-actin (A5441, Sigma-Aldrich, 1/100,000) were used in the study, performing overnight incubations at 4°C in orbital agitation. For secondary antibodies, horseradish peroxidase (HRP)-linked anti-rabbit (7074S, Cell Signaling, 1/2,000), or anti-mouse (7076S, Cell Signaling, 1/2,000) were used, performing incubations of 1 hour at room temperature in orbital agitation. Detection was performed by chemiluminiscence using NOVEX ECL Chemi Substrate and SuperSignal West Femto Maximum Sensitivity Substrate (Thermo Fisher Scientific). Same β-actin were represented when analyzed samples were the same and have been treated in same conditions.

Auzmendi-Iriarte J., Otaegi-Ugartemendia M., Carrasco-Garcia E., Azkargorta M., Diaz A., Saenz-Antoñanzas A., Andermatten J.A., Garcia-Puga M., Garcia I., Elua-Pinin A., Ruiz I., Sampron N., Elortza F., Cuervo A.M, & Matheu A. (2022). Chaperone-Mediated Autophagy Controls Proteomic and Transcriptomic Pathways to Maintain Glioma Stem Cell Activity. Cancer Research, 82(7), 1283-1297.

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Immunoblot Analysis of Stem Cell Markers

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Immunoblots were performed following standard procedures [26 (link)]. Specific antibodies against SOX2 (AB5603 Millipore, Burlington, MA, USA), p21^CIP1 (sc-397 Santa Cruz, Dallas, TX, USA), p27^KIP1 (sc-1641 Santa Cruz, Dallas, TX, USA), BMI1 (05-637 Millipore, Burlington, MA, USA) and β-actin (A5441, Sigma, St. Louis, MO, USA) were used in the study. For secondary antibodies, horseradish peroxidase (HRP)-linked anti-rabbit (7074S Cell Signaling, Danvers, MA, USA), anti-mouse (7076S Cell Signaling, Danvers, MA, USA) or anti-goat (sc-2020 Santa Cruz, Dallas, TX, USA) were used. Detection was performed by chemiluminescence using NOVEX ECL Chemi Substrate (Thermo Fisher, Waltham, MA, USA).

Saenz-Antoñanzas A., Moncho-Amor V., Auzmendi-Iriarte J., Elua-Pinin A., Rizzoti K., Lovell-Badge R, & Matheu A. (2021). CRISPR/Cas9 Deletion of SOX2 Regulatory Region 2 (SRR2) Decreases SOX2 Malignant Activity in Glioblastoma. Cancers, 13(7), 1574.

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Cytokine Profiling of Cell Supernatants

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Human Cytokine Antibody Array C5 Kit (Raybiotech) was performed, following the manufacturer's guidance. Before sample incubation, GNS166 pGK and shL2A supernatants were centrifuged at 1 × 10⁴ rpm for 4 minutes. Array membranes were incubated overnight with 1 mL of infected undiluted supernatant. After that, both biotin-conjugated antibodies and HRP-linked secondary antibody were incubated overnight. Detection was performed by chemiluminescence using Novex ECL Chemi Substrate (Thermo Fisher Scientific).

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Immunoblot and Immunofluorescence Assays

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Immunoblot and immunofluorescence assays were performed following standard procedures, as previously described [40 (link)]. Primary antibodies were: phospho Histone H3 Ser10 (ab14955, Abcam), TOMM20 (11802-1-AP, Proteintech), PGC1-α (NBP1-04676, Novus Biologicals), phospho p38MAPK Thr180/Tyr182 (9211, Cell Signaling), p38MAPK (sc-7972, Santa Cruz Biotechnology) AKT1/2/3 (sc-8312, Santa Cruz Biotechnology), phospho AKT Ser 473 (9271, Cell Signaling), DMPK (sc-134319, Santa Cruz Biotechnology), MBNL1 (ab45899, Abcam) and β-actin (AC-15, Sigma-Aldrich). For western blot detection of primary antibodies, we used HRP-linked antibodies (Santa Cruz Biotechnology) and the detection was performed by chemiluminescence using Novex ECL Chemi Substrate (Thermo Fisher). For immunofluorescence, nuclear DNA was stained with Hoechst (33342, Sigma-Aldrich).

García-Puga M., Saenz-Antoñanzas A., Fernández-Torrón R., de Munain A.L, & Matheu A. (2020). Myotonic Dystrophy type 1 cells display impaired metabolism and mitochondrial dysfunction that are reversed by metformin. Aging (Albany NY), 12(7), 6260-6275.

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Western Blot Analysis of PDGFR Signaling

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Cells were lysed and analysed by Western blotting as described previously [51 (link)]. Primary antibodies were: PDGFRα (cat# 3174S), PDGFRβ (cat# 3169S), p-PDGFRα (Y849)/β(Y857) (cat# 3170T), p-PDGFβ (Y740) (cat# 3168), p-SRC (Y416) (cat# 2101) and p-STAT5 (cat# 4322T) from Cell Signaling; p-PDGFRα (Tyr572)/PDGFRβ (Tyr579) (cat# bs-5554R) from Bioss Inc and β-Actin (cat# A5441) from Sigma. Detection was through enhanced chemiluminescence ECL using Horse Radish Peroxidase (HRP)-coupled secondary antibodies (GE Healthcare) and NOVEX ECL Chemi Substrate (ThermoFisher).

Aldaz P., Auzmendi-Iriarte J., Durántez M., Lasheras-Otero I., Carrasco-Garcia E., Zelaya M.V., Bragado L., Olías-Arjona A., Egaña L., Samprón N., Morilla I., Redondo-Muñoz M., Rico M., Squatrito M., Maria-Alonso M., Fernández-Irigoyen J., Santamaria E., Larráyoz I.M., Wellbrock C., Matheu A, & Arozarena I. (2021). Identification of a Dexamethasone Mediated Radioprotection Mechanism Reveals New Therapeutic Vulnerabilities in Glioblastoma. Cancers, 13(2), 361.

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