Supersignal west femto maximum sensitivity kit

Manufactured by Thermo Fisher Scientific

Sourced in United States

The SuperSignal West Femto Maximum Sensitivity kit is a chemiluminescent substrate used for the detection of proteins in Western blotting applications. It provides a high-sensitivity detection solution for low-abundance protein targets.

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Lab products found in correlation

Non fat dry milk, by Bio-Rad (1 mentions) Tris glycine gel, by Thermo Fisher Scientific (1 mentions) Secondary goat anti mouse and anti rabbit antibodies, by Santa Cruz Biotechnology (1 mentions) Nitrocellulose membrane, by Thermo Fisher Scientific (1 mentions) M per, by Thermo Fisher Scientific (1 mentions) Iblot transfer system, by Thermo Fisher Scientific (1 mentions) Las 3000 imaging system, by GE Healthcare (1 mentions) Chemidoc mp imaging system, by Bio-Rad (1 mentions) α αtubulin, by Merck Group (1 mentions) Multi gauge software, by GE Healthcare (1 mentions) α histone h3, by Abcam (1 mentions) Las 4000 mini bioimager, by GE Healthcare (1 mentions) α rabbit igg hrp, by Merck Group (1 mentions) Las 4000, by Cytiva (1 mentions) Avidin hrp, by BioLegend (1 mentions) Roti block, by Carl Roth (1 mentions) Pvdf membrane, by Merck Group (1 mentions) Protease inhibitor cocktail, by Cell Signaling Technology (1 mentions) Nitrocellulose membrane, by Bio-Rad (1 mentions) Cell lysis buffer, by Cell Signaling Technology (1 mentions)

Close spelling variants of this product

SuperSignal West Femto (427 citations) SuperSignal West Femto Maximum Sensitivity Substrate kit (129 citations) SuperSignal West Femto Maximum (12 citations) SuperSignal™ West Femto Maximum Sensitivity (10 citations) SuperSignal West Femto Maximum Sensitivity Substrate ECL kit (5 citations) SuperSignal West Femto Maximum Sensitivity Substrate detection kit (3 citations) SuperSignal West Femto maximum-sensitivity chemiluminescent substrate kit (2 citations)

6 protocols using supersignal west femto maximum sensitivity kit

Western Blot Analysis of STAT1 and Mx1

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Cell lysates were prepared using mammalian protein extraction reagent M-PER (Thermo Fisher Scientific, Waltham, MA) with Protease and Halt™ phosphatase inhibitor cocktails (Thermo Fisher Scientific) using an equal number of cells per sample. Samples were analyzed by SDS-PAGE using 10–20% Tris-Glycine gels (Thermo Fisher Scientific) under reducing conditions. As a molecular weight marker, protein ladder (cat# 7727S) from Cell Signaling Technology (Danvers, MA) was used. Nitrocellulose membranes and iBlot™ transfer system (Thermo Fisher Scientific) were used for Western Blot analysis. All other reagents for Western Blot analyses were purchased from Thermo Fisher Scientific. Membranes were blocked with nonfat dry milk (BIO-RAD, Hercules, CA) for 1 h followed by incubation with primary antibodies against STAT1, pSTAT1 (pY701, BD Transduction Lab, San Jose, CA), or Mx1 (gift from O. Haller, University of Freiburg, Freiburg, Germany) O/N at 4 °C. Secondary goat anti-mouse and anti-rabbit antibodies were purchased from Santa Cruz Biotechnology. SuperSignal West Femto Maximum Sensitivity Kit (Thermo Fisher Scientific) was used to develop membranes, and images were taken using LAS-3000 Imaging system (GE Healthcare Bio-Sciences, Pittsburgh, PA).

Chiang A.W., Li S., Kellman B.P., Chattopadhyay G., Zhang Y., Kuo C.C., Gutierrez J.M., Ghazi F., Schmeisser H., Ménard P., Bjørn S.P., Voldborg B.G., Rosenberg A.S., Puig M, & Lewis N.E. (2019). Combating viral contaminants in CHO cells by engineering innate immunity. Scientific Reports, 9, 8827.

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Western Blot Analysis of Bacterial Proteins

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Protein samples were separated on 10% SDS‐polyacrylamide gels and transferred to nitrocellulose membranes. The membranes were used in Western blot analyses with custom sheep anti‐SKA polyclonal antibodies (created for us by Pacific Immunology Inc), a commercial rabbit anti‐SLO/SPN polyclonal antibody (American Research Products Inc), a custom rabbit anti‐Spd3 polyclonal antibody (Pacific Immunology Inc) and a commercial horseradish peroxidase (HRP)‐conjugated rabbit anti‐SpeB polyclonal antibody (Toxin Technology) as primary antibodies. The blots were blocked with 5% non ‐fat milk in PBST buffer (2.7 mM potassium chloride, 137 mM sodium chloride pH 7.4 and 0.1% Tween 20) and incubated overnight at 4°C with specific primary antibodies. The proteins were detected using Alexa Fluor 680 donkey anti‐rabbit IgG (at a dilution 1:10,000) or HRP conjugated rabbit anti‐Sheep IgG (Abcam, at a dilution 1:20,000) secondary antibodies. The florescent signal was detected using a Li‐Cor Odyssey Near‐Infrared System or by using a ChemiDoc^TM MP Imaging System (BIO‐RAD) in association with the SuperSignal West Femto maximum sensitivity kit (ThermoFisher).

Jain I., Danger J.L., Burgess C., Uppal T, & Sumby P. (2019). The group A Streptococcus accessory protein RocA: regulatory activity, interacting partners and influence on disease potential. Molecular Microbiology, 113(1), 190-207.

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Separation of Nuclear Proteins by SDS-PAGE

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For separating nuclear-enriched protein extracts by SDS-PAGE, equal volumes of nuclear-enriched extracts were loaded. To separate total protein extracts, equal amounts of protein were resolved by SDS-PAGE. Protein samples were subsequently blotted onto PVDF membranes. After blotting, membranes were blocked with Rotiblock (Roth) reagent and incubated with the respective primary antibody followed by a horseradish peroxidase (HRP)-conjugated secondary antibody. HRP activity was detected using the SuperSignal West Femto Maximum Sensitivity kit (Thermo Scientific) and visualized by a LAS-4000 Mini bioimager (GE Healthcare Life Sciences). Signal intensities were quantified using Multi-Gauge software (GE Healthcare Life Sciences). Commercial antibodies used were HRP-conjugated α-HA (Roche), α-Histone H3 (Abcam), α-HSC70 (Stressgen), α-α-Tubulin (Sigma-Aldrich), α-rabbit IgG-HRP (Sigma-Aldrich) and α-mouse IgG-HRP (Sigma-Aldrich). α-SPA2 and α-COP1 antibodies were described previously in [42 (link)]. α-cry1 [67 ] and α-cry2 [68 (link)] antibodies were used to detect cry1 and cry2, respectively.

Chen S., Lory N., Stauber J, & Hoecker U. (2015). Photoreceptor Specificity in the Light-Induced and COP1-Mediated Rapid Degradation of the Repressor of Photomorphogenesis SPA2 in Arabidopsis. PLoS Genetics, 11(9), e1005516.

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Detection of HCMV IL-10 Protein

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RPE cells (1.2 × 10⁵) were infected with HCMV Merlin GFP or HCMV Merlin dUL11 GFP at a MOI of 1.0. After 6 days in culture, the cells were trypsinized and washed with ice-cold PBS before lysis in 200 μL 5× SDS-PAGE loading buffer (300 mM TRIS-HCl, 10%SDS, 0.1%bromophenol blue, 50% glycerol, 300 mM β-mecaptoethanol). The lysates were separated via SDS-PAGE before transfer onto a nitrocellulose membrane. The membranes were blocked with Roti block (Roth), and cmvIL-10 was detected using Viral HCMV IL-10 Biotinylated Antibody clone BAF117 (R&D Systems) and Avidin-HRP (Biolegend) prior to visualization using the Super Signal West Femto Maximum Sensitivity Kit (Thermo Scientific).

Osanyinlusi S.A., Zischke J., Jacobs R., Weissinger E.M., Schulz T.F, & Kay-Fedorov P.C. (2022). Human Cytomegalovirus pUL11, a CD45 Ligand, Disrupts CD4 T Cell Control of Viral Spread in Epithelial Cells. mBio, 13(6), e02946-22.

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Immunoblotting for NF-κB and AhR Signaling

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Cells were lysed with Cell Lysis buffer (1X) supplemented with protease inhibitor cocktail (Cell Signaling, USA). Total cell lysates (5–10 μg) were resolved on 4–12% Bis-Tris Nupage gels (Invitrogen, USA) and transferred onto PVDF membranes (Millipore) and developed with antibodies for GAPDH, NF-κB p65, Phospho-NF-κB p65, and anti-Rabbit IgG HRP-linked antibody using SuperSignal West Femto Maximum Sensitivity kit (Thermo Scientific), Data quantification was done using Image J software (NIH).
Alternatively, MOC1 and SUM149 cells were plated at 2 × 10 6 cells in T75 flasks and allowed to adhere overnight. Cells were pre-treated with 0.1% DMSO, 20 μM HP163 for 30 min and then treated with 0.1% DMSO, 100 μM Kyn or 1 nM TCDD for 1 hour. Nuclear and cytoplasmic cell extracts were prepared using a Nuclear Extract Kit (Thermo Scientific) as per the manufacturer’s instructions. Protein concentration was quantified using the Protein Assay Reagent (Bio-Rad). Protein (30 μg) was resolved on 10% SDS-polyacrylamide gels and transferred to nitrocellulose membranes (Bio-Rad). Membranes were probed with mouse anti-AHR (Pierce, cat #MA1-514), rabbit anti-Lamin-A/C (Cell Signaling, cat #2032) and mouse anti-α-tubulin (EMD Millipore, cat #CP06). Immuno-reactive bands were detected using HRP-conjugated secondary antibodies (goat anti-rabbit, Bio-Rad; goat anti-mouse Pierce) and ECL substrate.

Giovannoni F., Bosch I., Polonio C.M., Torti M.F., Wheeler M.A., Li Z., Romorini L., Rodriguez Varela M.S., Rothhammer V., Barroso A., Tjon E.C., Sanmarco L.M., Takenaka M.C., Modaresi S., Gutiérrez-Vázquez C., Zanluqui N.G., dos Santos N.B., Munhoz C.D., Wang Z., Damonte E.B., Sherr D., Gehrke L., Peron J.P., Garcia C.C, & Quintana F.J. (2020). AHR is a Zika virus host factor and a candidate target for antiviral therapy. Nature neuroscience, 23(8), 939-951.

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Native Protein Complex Characterization

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Protein samples were solubilized with 5% digitonin on ice for 15 min followed by centrifugation (20,000g, 30 min, 4°C). 0.25% of G-250 was added to the supernatant and complexes were separated via 4–16% Bis-Tris gels (NativePAGE, Thermo Fisher Scientific). The complexes were transferred to PVDF membranes via wet blotting without methanol. After fixation (8% acetic acid), destaining (50% methanol and 25% acetic acid), and blocking (3% milk powder in TBST, 1 h, RT), the membranes were incubated with the respective primary antibodies (overnight, 4°C, rotating). The used antibodies are listed above. The staining of HRP-coupled secondary antibodies was detected with the Clarity Western ECL Kit (Bio-Rad)/SuperSignal West Femto Maximum sensitivity Kit (Thermo Fisher Scientific) using a Bio-Rad detection system.

Zhang L., Dietsche F., Seitaj B., Rojas-Charry L., Latchman N., Tomar D., Wüst R.C., Nickel A., Frauenknecht K.B., Schoser B., Schumann S., Schmeisser M.J., vom Berg J., Buch T., Finger S., Wenzel P., Maack C., Elrod J.W., Parys J.B., Bultynck G, & Methner A. (2022). TMBIM5 loss of function alters mitochondrial matrix ion homeostasis and causes a skeletal myopathy. Life Science Alliance, 5(10), e202201478.

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