Snap-frozen EDL muscles were homogenized in RIPA lysis buffer containing protease inhibitor cocktail (Sigma-Aldrich). 30 ug of protein homogenates were electrophoretically separated on 4/15% gels (Bio-Rad) and transferred to polyvinylidene difluoride membranes (Bio-Rad), blocked in 5% skim milk, and incubated overnight at 4°C with anti-utrophin primary antibody 1/500 (Santa Cruz Biotechnology). The membranes were washed and incubated with horseradish peroxidase-conjugated secondary antibody (Santa Cruz Biotechnology). Protein bands were revealed using the ECL-Plus chemiluminescent detection system (Perkin-Elmer) and were normalized for loading using Gelcode Blue Stain reagent (Thermo Scientific). ImageQuant LAS 4000 biomolecular imager was used to detect a chemiluminescent signal and analyzed using Quantity One software (version 4.6.6; Bio-Rad). Representative utrophin blot is presented in S1 Fig .
Ecl plus chemiluminescent detection system
Manufactured by PerkinElmer
The ECL-Plus chemiluminescent detection system is a laboratory equipment product designed for the detection of protein and nucleic acid samples. It utilizes a chemiluminescent substrate to generate a luminescent signal that can be detected and quantified.
Automatically generated - may contain errors
Lab products found in correlation
Gelcode blue stain reagent, by Thermo Fisher Scientific (1 mentions)
Horseradish peroxidase conjugated secondary antibody, by Santa Cruz Biotechnology (1 mentions)
Quantity one software, by Bio-Rad (1 mentions)
Polyvinylidene difluoride membrane, by Bio-Rad (1 mentions)
Protease inhibitor cocktail, by Merck Group (1 mentions)
Quantity one software version 4, by Bio-Rad (1 mentions)
Glyceraldehyde 3 phosphate dehydrogenase, by Santa Cruz Biotechnology (1 mentions)
Horseradish peroxidaseeconjugated secondary antibody, by Santa Cruz Biotechnology (1 mentions)
2 protocols using ecl plus chemiluminescent detection system
1
Utrophin Protein Expression Analysis in Muscle Tissue
2
Western Blotting Analysis of β2-AR in Skeletal Muscle
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For Western blotting, Sol and EDL muscles from mice treated for 10 days with OPG-Fc (1 mg/kg per day) or formoterol (10 or 100 mg/kg per day) alone or in combination were homogenized in lysis buffer, separated on 9% SDS-polyacrylamide gels, and transferred to polyvinylidene difluoride membranes. The protein bands were immunoblotted with an antieb 2 -AR primary antibody overnight at 4 C (1:500; Santa Cruz Biotechnology, Dallas, TX), followed by a horseradish peroxidaseeconjugated secondary antibody (1:1000; Santa Cruz Biotechnology). Bands were revealed using the ECL-Plus chemiluminescent detection system (PerkinElmer, Waltham, MA). Images of the membranes were acquired, scanned, and analyzed using Quantity One software version 4.6.6 (Bio-Rad, Mississauga, ON, Canada). Glyceraldehyde-3-phosphate dehydrogenase (1:5000; Santa Cruz Biotechnology) was used as a loading control. Images of the membranes were acquired, scanned, and analyzed using Quantity One software version 4.6.6.
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