SOD-2, PPARα and γ expression markers were assayed through western blotting analyses. The proteins have been extracted using Radio-Immunoprecipitation Assay (RIPA) buffer R0278 (Sigma Aldrich, Milan, Italy) and the concentrations were determined using the Bio-Rad protein assay reagent (Bio-Rad Laboratories, Milan Italy). Equal amounts of protein (30 μg) were loaded on a SDS-PAGE and transferred to a nitrocellulose membrane [19 –22 (link)]. The filters were incubated with a monoclonal antibody against SOD-2 (A-2, sc-133,134) diluted 1:250 v/v, polyclonal antibody against PPARα (H-98, sc-9000) diluted 1:200 v/v, monoclonal antibody against PPARγ (E-8, sc-7273) diluted 1:250 v/v, polyclonal antibody against Actin (I-19, sc-1616) diluted 1:500 v/v, at room temperature for 2 h. All reagents were from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Membranes were washed three times for 10 min and incubated with a 1:10,000 v/v dilution of horseradish peroxidase-conjugated anti-mouse, anti-rabbit and anti-goat antibodies for 1 h respectively. Blots were developed using ECL system (Amersham Biosciences, UK) according to the manufacturer’s protocols.
Ripa buffer r0278
Manufactured by Merck Group
Sourced in Italy
RIPA buffer (R0278) is a laboratory reagent used for the extraction and solubilization of proteins from cells and tissues. It is a commonly used lysis buffer that contains a combination of ionic and non-ionic detergents, as well as other components, to ensure efficient protein extraction.
Automatically generated - may contain errors
Lab products found in correlation
Tfeb a303 673a, by Fortis Life Sciences (2 mentions)
Lps derived from e coli 0111 b4 l2630, by Merck Group (2 mentions)
Luria bertani broth l8050 luria bertani agar, by Teknova (2 mentions)
Ampicillin l1002, by Teknova (2 mentions)
Pbs 10010 023, by Thermo Fisher Scientific (2 mentions)
Zkscan3, by Santa Cruz Biotechnology (2 mentions)
β actin a5441, by Merck Group (2 mentions)
Lamp1, by Abcam (2 mentions)
Ecl system, by Cytiva (1 mentions)
Protein assay reagent, by Bio-Rad (1 mentions)
3 protocols using ripa buffer r0278
1
Protein Expression Analysis of Antioxidant and Metabolic Regulators
2
Immunoblotting and Immunostaining Analysis of Lysosomal Regulators
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LPS derived from E. coli 0111:B4 (L2630) and RIPA buffer (R0278) were purchased from Sigma (St. Louis, MO), Luria-Bertani broth (L8050) Luria-Bertani Agar+50μg/ml Ampicillin (L1002) were from Teknova (Hollister, CA). PBS (10010–023) was obtained from Thermo Fisher Scientific (Waltham, MA), whereas saline (MSDW-1000) was from Growcells (Irvine, CA). Western blot analysis was conducted using specific antibodies to LAMP1 (Ab25245; Abcam Cambridge, MA), TFEB (A303–673A; Bethyl, Montgomery, TX), and ZKSCAN3 (SC515285; Santa Cruz, Dallas TX). Antibodies to TFE3 (HPA023881), LC3 (L8918) and β-actin (A5441) were from Sigma (St. Louis, MO). Antibodies for immunohistochemistry are as the follows: Rubicon: Proteintech 21444–1-AP.
3
Immunoblotting and Immunostaining Analysis of Lysosomal Regulators
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