Supersignal west femto chemiluminescent reagent

Manufactured by Thermo Fisher Scientific

Sourced in United States

SuperSignal West Femto is a chemiluminescent reagent designed for the detection of proteins in Western blot analysis. It provides a high-intensity signal for the detection of low-abundance proteins.

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Lab products found in correlation

Protan ba85, by Cytiva (2 mentions) Chemidoc touch imaging system, by Bio-Rad (2 mentions) Hrp conjugated anti rabbit igg, by Thermo Fisher Scientific (1 mentions) Ipvh00010, by Merck Group (1 mentions) Protease inhibitor cocktail, by Roche (1 mentions) Chemidoc xrs system, by Bio-Rad (1 mentions) Peroxidase labelled rabbit anti mouse immunoglobulins, by Agilent Technologies (1 mentions) Nw04125box, by Thermo Fisher Scientific (1 mentions) P0260, by Agilent Technologies (1 mentions) Supersignal west pico plus chemiluminescent reagent, by Thermo Fisher Scientific (1 mentions)

4 protocols using supersignal west femto chemiluminescent reagent

Western Blotting of Macrophage Phosphoproteins

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Western blotting of macrophage lysates was performed with rabbit
anti-phospho antibodies (PAN) (Thermo Fisher Scientific). Briefly, 50µg
proteins were resolved on 10% denatured polyacrylamide gel followed by transfer
on nitrocellulose membrane by using semi-dray transfer approach. The membrane
was blocked with blocking buffer (TBS containing 0.05% Tween 20 and 3% bovine
serum albumin) for 1hr at 25°C followed by overnight incubation at
4°C with 1:500 dilution of anti-phospho antibodies in blocking buffer.
Membrane was subsequently washed thrice in TBST (TBS containing 0.05% Tween 20)
followed by incubation with HRP-conjugated anti-rabbit IgG (Thermo Fisher
Scientific) for 1 hour at 25°C. Finally, the immunoblot was washed with
TBST for three times and signals were obtained by incubating with SuperSignal
West Femto chemiluminescent reagent (Thermo Fisher Scientific) and captured on
X-ray films.

Choudhary E., Bullen C.K., Goel R., Singh A.K., Praharaj M., Thakur P., Dhiman R., Bishai W.R, & Agarwal N. (2020). Relative and Quantitative Phosphoproteome Analysis of Macrophages in Response to Infection by Virulent and Avirulent Mycobacteria Reveals a Distinct Role of the Cytosolic RNA Sensor RIG-I in Mycobacterium tuberculosis Pathogenesis. Journal of proteome research, 19(6), 2316-2336.

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Western Blot Protein Analysis

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Cells were lysed in Sodium Dodecyl Sulfate (SDS) lysis buffer (10% glycerol, 2% SDS in 62.5 mM Tris–HCl), including Protease Inhibitor Cocktail (Roche). After the determination of protein concentration, an equal amount of protein from each lysate was resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to a polyvinylidene difluoride membrane (Millipore, #IPVH00010). Membranes were incubated with primary antibodies at 4°C overnight, then incubated with the secondary antibody for 2 hours at room temperature. Finally, these membranes were immersed in SuperSignal West Femto chemiluminescent reagent (Thermo, #34095), and the protein bands were visualized with the ChemiDocTMXRS system (Bio-Rad, Calif, USA).

Balçık-Erçin P., Aysan A., Salık N, & Erciyas E. (2023). SIX1 Downregulation Suppresses Self-renewal Capacity and THY1 Expression in Hepatocellular Carcinoma and SIX1 Dominate the Survival in Liver Cancer. The Turkish Journal of Gastroenterology, 34(8), 881-889.

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Cell and Tissue Protein Extraction and Western Blot

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Cell lysis buffer (125 mM Tris pH 6.8, 2 % SDS, 5 % 2-beta mercaptoethanol, 5 % glycerol with protease inhibitors: Sigma P8340 and 1mM PMSF) was used for the extraction of cultured cells and tissue lysis buffer (50mM Tris pH 6.8, 1 % EDTA, 10 % SDS, 5 % beta mercaptoethanol, 10 % glycerol with protease inhibitors) was used for the extraction of tissue samples (250 mg/ml). Bromophenol blue was added to the samples which were then boiled and separated by SDS-PAGE using 4 to 12 % polyacrylamide gels (Ref: NW04125BOX; ThermoFisher Scientific) and then electroblotted onto nitrocellulose membranes (Protan BA85, Whatman). Non-specific sites were blocked with 5 % skimmed milk protein and the membranes then incubated with monoclonal antibody supernatants (diluted: 1/10, except MANNES1E: 1/50 and MANDRA1: 1/100), followed by washing and incubation with secondary antibody (peroxidase-labelled rabbit anti-mouse immunoglobulins; 1/1000, Dako, Denmark). Antibody reacting bands were detected with SuperSignal West Femto chemiluminescent reagent (Cat No: 34094; ThermoFisher Scientific) and visualized with a ChemiDoc Touch imaging system (BioRad Ltd.).

Holt I., Duong N.T., Zhang Q., Lam L.T., Sewry C.A., Mamchaoui K., Shanahan C.M, & Morris G.E. (2016). Specific localization of nesprin-1-α2, the short isoform of nesprin-1 with a KASH domain, in developing, fetal and regenerating muscle, using a new monoclonal antibody. BMC Cell Biology, 17, 26.

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Western Blot Analysis of XIRP1 and Annexin A5

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Samples were mixed with SDS buffer (125 mM Tris pH 6.8; 2% SDS; 5% 2-beta mercaptoethanol; 5% glycerol; with bromophenol blue), boiled and subjected to SDS-PAGE using 10% polyacrylamide gels and transferred to nitrocellulose membranes (Protan BA85, Whatman). Non-specific sites were blocked with 5% skimmed milk protein, membranes washed with PBS and then incubated with primary monoclonal antibodies against: XIRP1, (Xin-alpha (D-8); sc-166,658; 1/100; Santa Cruz Biotechnology, Insight Biotechnology Ltd., Wembly, UK); or GST, mAb 17A10 (1/100). This was followed by washing in PBS and incubation with secondary antibody (peroxidase-labelled rabbit anti-mouse immunoglobulins, P0260; 1/1000; Dako, Denmark). Alternatively, for the detection of annexin A5, pAb ANXA5 (Abcam; ab14196; 1.4 μg/mL) primary antibody followed by goat anti-rabbit Ig HRP (P0448; Dako; 1/1000) secondary antibody. All antibodies were diluted in PBS containing 0.05% Triton X, 0.1% BSA, 1% horse serum and 1% fetal bovine serum. XIRP1 and annexin A5 antibody reacting bands were detected with SuperSignal West Femto chemiluminescent reagent (Cat No: 34094; ThermoFisher Scientific) and GST antibody reacting bands were detected with SuperSignal West Pico Plus chemiluminescent reagent (Cat No: 34580; ThermoFisher Scientific) and visualized with a ChemiDoc Touch imaging system (BioRad Ltd.).

Holt I., Fuller H.R., Schindler R.F., Shirran S.L., Brand T, & Morris G.E. (2020). An interaction of heart disease-associated proteins POPDC1/2 with XIRP1 in transverse tubules and intercalated discs. BMC Molecular and Cell Biology, 21, 88.

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