Femto chemiluminescence detection system

Manufactured by Thermo Fisher Scientific

Sourced in United States

The Femto chemiluminescence detection system is a lab equipment designed to measure and analyze low-level light emissions, known as chemiluminescence, from biological samples. The system provides sensitive detection and quantification of chemiluminescent signals.

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Lab products found in correlation

Nitrocellulose membrane, by Bio-Rad (2 mentions) Mops buffer, by Bio-Rad (2 mentions) Protease inhibitor cocktail, by Roche (2 mentions) Nonfat milk powder, by Bio-Rad (1 mentions) 5 lox, by Abcam (1 mentions) Cox 2, by Abcam (1 mentions) Protein assay kit, by Bio-Rad (1 mentions) Pvdf membrane, by Merck Group (1 mentions) β actin, by ICN Biomedicals (1 mentions) Myecl imager, by Thermo Fisher Scientific (1 mentions) Ripa lysis buffer, by Merck Group (1 mentions) Gapdh, by Abcam (1 mentions)

Spelling variants of this product

Chemiluminescence detection system (58 citations) Chemiluminescence system (42 citations) Femto system (4 citations) Femto detection system (2 citations) SuperSignal West‐Femto Chemiluminescence detection system (2 citations)

6 protocols using femto chemiluminescence detection system

Western Blotting of Inflammatory Markers

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Protein was extracted from the LVI or the remote LV, as previously described.26 After electrophoresis of 5 to 10 μg of LVI or remote LV protein, primary antibodies ALOX‐5, HO‐1, COX‐2, ALX/FPR2, TNF‐α, or GAPDH were allowed to incubate overnight at 4°C. After the incubation, respective secondary antibody (1:10 000 dilution in a 5% blocking solution) was added, as previously described.26 Femto chemiluminescence detection system was used to detect protein expressions (Pierce Chemical, Rockford, IL). To normalize the total protein/lane, densitometry was performed using Image J software (NIH, Bethesda, MD).26

Pullen A.B., Kain V., Serhan C.N, & Halade G.V. (2020). Molecular and Cellular Differences in Cardiac Repair of Male and Female Mice. Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease, 9(8), e015672.

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Quantifying Protein Levels in Macrophages and Fibroblasts

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Immunoblotting was used to quantify the protein levels in peritoneal macrophages and cardiac fibroblast. The samples were resolved on criterion XT bis-tris 4–12% 18 well (Bio-Rad Inc. Hercules, California) gel and MOPs Buffer (Bio-Rad, Hercules, California). The kaleidoscope precision plus standard (Bio-Rad) was used to determine the molecular weight of the protein for immunoblotting, 10–15 μg of protein lysate per sample was denatured and resolved by using criterion XT bis-tris 4–12% 18-gel (Bio-Rad I, Hercules, California.) gel in MOPs Buffer (Bio-Rad)., then transferred to a nitrocellulose membrane (Bio-Rad, Hercules, California), and blocked with 5% nonfat milk. The membrane was probed with COX-2 (1:1000) and 5LOX (1:200) overnight at 4˚C, followed by secondary antibody (Bio-Rad, Hercules, California). The proteins were detected using the Femto chemiluminescence detection system (Pierce Chemical, Rockford, IL). Densitometry was performed using Image J software (NIH, USA).

Kain V, & Halade G.V. (2018). Immune responsive Resolvin D1 programs peritoneal macrophages and cardiac fibroblat phenotypes in diversified metabolic microenvironment. Journal of cellular physiology, 234(4), 3910-3920.

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Quantitative Immunoblotting of LVI Proteins

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Electrophoresis of 10 µg of LVI protein as previously described^{15 (link)} and probed with primary antibody (FPR2; 1:1000 and GPR40; 1:1000) overnight at 4 °C followed by secondary antibody (Biorad). The proteins were detected using femto chemiluminescence detection system (Pierce Chemical, Rockford, IL, USA). Densitometry was performed using Image J software (NIH, USA).

Kain V., Liu F., Kozlovskaya V., Ingle K.A., Bolisetty S., Agarwal A., Khedkar S., Prabhu S.D., Kharlampieva E, & Halade G.V. (2017). Resolution Agonist 15-epi-Lipoxin A4 Programs Early Activation of Resolving Phase in Post-Myocardial Infarction Healing. Scientific Reports, 7, 9999.

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Western Blot Analysis of Inflammatory Mediators

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Electrophoresis of 10 µg of LVI protein were performed using criterion XT bis-tris 4–12 % gel (Bio-Rad Inc.) in MOPs Buffer (Bio-Rad) and transferred on nitrocellulose membrane (BioRad Inc.). The total protein stain was acquired using pierce reversible protein stain, nitrocellulose membranes kit (Thermo Scientific Inc.). The membrane was blocked for 1 hr at room temperature, using 5% non-fat milk powder (Bio-Rad) dissolved in TPBS and probed with primary antibody [COX-2 1:1000, COX-1 1:1000, 5-LOX 1:200 (abcam) and ALX/FPR2 1:500 (Santa cruz)] overnight at 4°C followed by secondary antibody (Biorad). The proteins were detected using the femto chemiluminescence detection system (Pierce Chemical, Rockford, IL, USA). Densitometry was performed using Image J software (NIH, USA).

Kain V., Ingle K.A., Colas R.A., Dalli J., Prabhu S.D., Serhan C.N., Joshi M, & Halade G.V. (2015). Resolvin D1 activates the inflammation resolving response at splenic and ventricular site following myocardial infarction leading to improved ventricular function. Journal of molecular and cellular cardiology, 84, 24-35.

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PARP Cleavage and Mn-SOD Expression Assay

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To analyze PARP cleavage [29] (link), and Mn-SOD expression, cells (3.5×10⁴) were treated with 5.5 and 33 mM glucose with or without various concentrations of ML1 and 100U Mn-SOD for the indicated time. Cells were scraped into radio-immunoprecipitation assay lysis buffer (120 mM NaCl, 1.0% Triton X-100, 20 mM Tris–HCl, pH 7.5, 10% glycerol, 2 mM EDTA, protease inhibitor cocktail (Roche GmbH, Germany) and the protein concentration in each sample was determined using a Bio-Rad protein assay kit with BSA as the standard. For immunoblotting, 40 μg of protein lysate per sample was denatured in 4×SDS–PAGE sample buffers and resolved by 8–12% SDS–PAGE, transferred to a PVDF membrane (Millipore, Germany), blocked with nonfat milk, and probed for PARP, Mn-SOD using HRP-conjugated appropriate secondary antibody. The enhanced chemiluminescence was detected using the Femto chemiluminescence detection system (Pierce Chemical, Rockford, IL, USA). Membranes were stripped and re-probed with β-actin (ICN Biomedicals, USA) primary antibody (1:10,000) as a protein loading control. Densitometry was performed using Image J software.

Kain V., Sawant M.A., Dasgupta A., Jaiswal G., Vyas A., Padhye S, & Sitasawad S.L. (2016). A novel SOD mimic with a redox-modulating mn (II) complex, ML1 attenuates high glucose-induced abnormalities in intracellular Ca2+ transients and prevents cardiac cell death through restoration of mitochondrial function. Biochemistry and Biophysics Reports, 5, 296-304.

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Immunoblotting of Cardiovascular Markers

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Immunoblotting was done using 10–15 μg of LV tissues from 4 months old WT and ALX^−/− mice. Briefly, LV lysates were prepared using radio-immunoprecipitation assay (RIPA) lysis buffer (Sigma) and protease inhibitor cocktail (Roche GmbH, Germany). The proteins were electrophoresed. The blots were probed with primary CD31 antibody (Cat# ab124432, Abcam, 1/1000), eNOS (Cat# ab76198, Abcam, 1/1000), peNOS (Cat# ab184154, Abcam, 1/1000), COX-2 (Cat# ab15191, Abcam, 1/1000), iNOS (Cat#ab15323, Abcam),1:1000 or GAPDH (cat# ab9485, Abcam; 1/5000) kept overnight at 4 °C followed by respective secondary antibody (Biorad) as described previously. The proteins were detected using the Femto chemiluminescence detection system (Pierce Chemical, Rockford, IL, USA). After exposure, blot membrane image were acquired using myECL^™ Imager (Thermo Scientific, USA). Densitometry data were normalized to GAPDH or total eNOS expression using Image J software (NIH, USA) (21 (link)).

Tourki B., Kain V., Shaikh S.R., Leroy X., Serhan C.N, & Halade G.V. (2020). Deficit of Resolution Receptor Magnifies Inflammatory Leukocyte Directed Cardiorenal and Endothelial Dysfunction with Signs of Cardiomyopathy of Obesity. FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 34(8), 10560-10573.

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