SDS-PAGE was as described (Ausubel et al., 1995 ; Towbin et al., 1979), loading equal amounts (5 μg/lane) of protein along with pre-stained Low Range SDS-PAGE Standards (BioRad). The gel was electroblotted to Immobilon-P PVDF transfer membrane (Millipore) using the MiniBlot Module (Thermo EC). Immunoblotting was done according to the manufacturer’s instructions using ECL Advance Western Blotting Detection Kit (Amersham Pharmacia). The primary mouse anti-HA monoclonal antibody (clone HA-7, Sigma, St. Louis, MO) was used at 1:5000 dilution. The secondary was goat anti-mouse IgG1 conjugated horseradish peroxidase (sc-2060; 1:5000 dilution; Santa Cruz Biotechnology, Santa Cruz, CA).
Low range sds page standards
Manufactured by Bio-Rad
Sourced in United States, Italy
The Low-range SDS-PAGE Standards is a set of pre-stained protein standards used for estimating the molecular weights of proteins in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) experiments. The standards cover a range of low-molecular-weight proteins, typically from 2.5 to 45 kDa.
Automatically generated - may contain errors
Lab products found in correlation
Coomassie blue g 250, by Bio-Rad (3 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
Ecl advance western blotting detection kit, by Cytiva (1 mentions)
Immobilon p pvdf transfer membrane, by Merck Group (1 mentions)
Mini protean 2 system, by Bio-Rad (1 mentions)
Ppiczαa, by Thermo Fisher Scientific (1 mentions)
Glycerol, by Vetec (1 mentions)
Sorbitol, by Merck Group (1 mentions)
Pierce glycoprotein staining kit, by Thermo Fisher Scientific (1 mentions)
Pichia pastoris x 33, by Thermo Fisher Scientific (1 mentions)
Zeocin, by Thermo Fisher Scientific (1 mentions)
Ethylene diamine tetraacetic acid (edta), by Merck Group (1 mentions)
Trypsin, by Promega (1 mentions)
Restriction enzyme, by New England Biolabs (1 mentions)
Pngase f, by New England Biolabs (1 mentions)
Methanol, by Vetec (1 mentions)
Electroporation cuvette, by Bio-Rad (1 mentions)
6 protocols using low range sds page standards
1
Western Blot Analysis of HA-tagged Proteins
2
SDS-PAGE Protein Separation and Visualization
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SDS-PAGE was carried out in non-reducing conditions according to Laemmli [25 (link)], on 12% polyacrylamide gel. Gels were stained either by Coomassie Blue G-250 (BioRad, Milan, Italy), or silver staining as described by Merril et al. 1981 [26 (link)]. Low-range SDS-PAGE Standards (Bio-Rad, Hercules, CA, USA) were used for estimation of Mr.
3
SDS-PAGE Protein Separation Technique
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SDS–PAGE was carried out according to Laemmli [35 (link)] on 12% polyacrylamide gel. For peptides separations, 15% polyacrylamide gels were used. Gels were stained by Coomassie Blue G-250 (BioRad, Milan, Italy.) Low-range SDS-PAGE Standards (Bio-Rad, Hercules, CA, USA) were used for the Mr calculations.
4
SDS-PAGE Analysis of Xylanase and Amylase
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Protein profiles were analysed by denaturing polyacrylamide gel electrophoresis (SDS-PAGE) in a MiniProtean II system (Bio-Rad). Proteins were detected by 0.5 % Coomassie brilliant blue R staining and Low-Range SDS-PAGE Standards (Bio-Rad) were used as markers. Xylanase and amylase band intensities were analysed with ImageJ software [44 (link)].
5
SDS-PAGE Protein Separation Protocol
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SDS-PAGE was carried out according to [81 (link)], on 12% polyacrylamide gel. For peptides separations, 16% polyacrylamide gels were used. Gels were stained by Coomassie Blue G-250 (BioRad, Milan, Italy.) Low-range SDS-PAGE Standards (Bio-Rad, Hercules, CA, USA) were used for Mr calculations
6
Heterologous Protein Expression in Pichia pastoris
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Yeast extract, peptone, biotin, dextrose, agar, peroxidase-conjugated anti-rabbit secondary antibody, Pefabloc SC (4-(2-aminoethyl)-benzenesulfonyl fluoride), sorbitol, EDTA (ethylenediaminetetraacetic acid) and DAB (3,3’-diaminobenzidine) substrate kit were purchased from Sigma (Missouri, USA). Glycerol and methanol were supplied by VETEC (Rio de Janeiro, Brazil). Pichia pastoris X-33, pPICZαA, and Zeocin were purchased from Invitrogen (California, USA). Restriction enzymes and PNGase F were furnished by New England Biolabs (Massachusetts, USA). Electroporation cuvettes, TMB (3,3’,5,5’-tetramethylbenzidine) EIA substrate kit and low-range SDS-PAGE standards were obtained from Bio-Rad Laboratories (California, USA). Trypsin was purchased from Promega (California, USA). Pierce Glycoprotein Staining kit, DMEM (Dubelcco’s Modified Eagle Medium), and fetal bovine serum were purchased from Thermo Fisher Scientific (Massachusetts, USA).
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