Mini protean tgx gradient gel

Manufactured by Bio-Rad

The Mini-PROTEAN TGX gradient gels are pre-cast polyacrylamide gels designed for fast, efficient protein separation. The gels feature a continuous polyacrylamide gradient that allows for the separation of a wide range of protein molecular weights in a single gel.

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Lab products found in correlation

Supersignal west pico chemiluminescent substrate, by Thermo Fisher Scientific (3 mentions) 40150 t62 cov2, by Sino Biological (3 mentions) Chemiluminescent substrate, by Thermo Fisher Scientific (3 mentions) Las 4000, by Cytiva (3 mentions) Horseradish peroxidase, by Merck Group (2 mentions) Chemidoc touch imagining system, by Bio-Rad (2 mentions) Hrp conjugated secondary antibody, by Cell Signaling Technology (2 mentions) Chemidoc imaging system, by Bio-Rad (2 mentions) Odyssey clx, by LI COR (2 mentions) Trans blot turbo, by Bio-Rad (2 mentions) Nitrocellulose membrane, by Bio-Rad (2 mentions) Protease inhibitor cocktail, by Roche (2 mentions) Amersham ecl prime western blotting detection reagent, by GE Healthcare (2 mentions) Amersham ecl prime, by Cytiva (2 mentions) Western lightning plus ecl substrate, by PerkinElmer (1 mentions) Coomassie brilliant blue r 250, by Bio-Rad (1 mentions) Immobilon p pvdf membrane, by Merck Group (1 mentions) Chemidoc mp system, by Bio-Rad (1 mentions) Chemidoc touch imaging system, by Bio-Rad (1 mentions) Anti rabbit and anti mouse hrp conjugated secondary antibodies, by Cell Signaling Technology (1 mentions)

Spelling variants of this product

4–20% gradient gels (350 citations) TGX gels (272 citations) Mini-PROTEAN TGX (242 citations) Mini-PROTEAN (158 citations) Mini-PROTEAN gels (77 citations) 4–20% TGX gradient gel (44 citations) Mini-PROTEAN® TGX™ precast 4–15% gradient gels (3 citations) TGX mini-gel (2 citations) Gradient Mini-PROTEAN TGX precast protein gels (2 citations) Mini-PROTEAN TGX 5–20% gradient gels (2 citations)

28 protocols using mini protean tgx gradient gel

Protein Analysis of Muscle Extracts

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Solubilised muscle extracts were separated by SDS-PAGE on a 7.5% gel, using 10 μg of collagen I as a standard. Gels were stained with Coomassie Brilliant Blue-R 250 (Bio-Rad, Hercules, CA), and destained in 40% methanol/10% glacial acetic acid/50% ddH₂O. Gels were imaged using a ChemiDoc MP System (BioRad).
For Western blots, solubilised muscle extracts (40 μg) were separated by SDS-PAGE on a 4–15% Mini-PROTEAN TGX gradient gel (Bio-Rad) and proteins transferred to Immobilon-P PVDF membrane (Millipore, Billerica, MA). The membrane was blocked in 5% skimmed milk/0.1% Tween-20/PBS overnight at 4°C and incubated in primary antibodies. Membranes were washed in 0.1% Tween-20 in PBS (PBST) and incubated in PBST containing anti-rabbit, anti-mouse or anti-rat HRP-conjugated secondary antibodies (dilution, 1:2000 for anti-rabbit and 1:1000 for anti-mouse and rat). Membranes were washed, incubated in Western Lightning Plus-ECL substrate (Perkin Elmer, Waltham, MA) for 5 min, then imaged using ChemiDoc MP System.

Chaturvedi V., Dye D.E., Kinnear B.F., van Kuppevelt T.H., Grounds M.D, & Coombe D.R. (2015). Interactions between Skeletal Muscle Myoblasts and their Extracellular Matrix Revealed by a Serum Free Culture System. PLoS ONE, 10(6), e0127675.

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Western Blot Analysis of Protein Lysates

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Protein lysates were prepared from cell lines by mechanical homogenization in 1.5% dodecyl maltoside (DDM, Sigma) under ice-cold conditions. Protein concentration was measured using the BCA protein assay kit. About 35–50 μg of protein was loaded on 10–12% mini-protean TGX gradient gel (BioRad) at 100 V for 2 h at room temperature and transferred on to nitrocellulose membrane at 35 V overnight at 4°C. The membranes were blocked using AdvanBlock-chemi blocking solution (Advansta) for 1 h at room temperature, incubated with primary antibody overnight at 4°C followed by incubation with HRP-conjugated secondary antibodies for 1 h at room temperature. The signal was developed using SuperSignal West Pico Chemiluminescent Substrate (ThermoFisher Scientific) and digitally imaged using the ChemiDoc Touch Imaging System (BioRad). The primary antibodies for CPT1A (Proteintech #15184-1-AP), PARP (Cell Signaling #9542), cleaved PARP (Cell Signaling #5625) and β-actin (Cell Signaling #4970) were purchased from the noted manufacturer. HRP-conjugated anti-mouse and anti-rabbit secondary antibodies were purchased from Cell Signaling.

Jariwala N., Mehta G.A., Bhatt V., Hussein S., Parker K.A., Yunus N., Parker J.S., Guo J.Y, & Gatza M.L. (2021). CPT1A and fatty acid β-oxidation are essential for tumor cell growth and survival in hormone receptor-positive breast cancer. NAR Cancer, 3(3), zcab035.

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SARS-CoV-2 Spike and Nucleocapsid Proteins Detection

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BHK cells infected at 5 MOI were harvested 24-post infection. Proteins were solubilized in PBS with 0.1% Triton X-100, supplemented with protease inhibitor, then reduced and denatured in Laemmli buffer containing DTT and boiled at 95°C for ~10 minutes. Proteins were resolved on a 4–20% Mini Protean TGX gradient gel (BioRad), and transferred onto PVDF membrane. S protein was probed with anti-SARS-CoV-1 S1 subunit rabbit polyclonal antibody (40150-T62-COV2, Sino Biological); N protein was probed with anti-SARS-CoV1 NP rabbit polyclonal antibody (40413-T62, Sino Biological). Vaccinia BR5 protein was probed as a loading control. Anti-rabbit polyclonal antibody conjugated with horseradish peroxidase (Sigma-Aldrich) was used as a secondary antibody and protein bands were visualized with chemiluminescent substrate (ThermoFisher).

Chiuppesi F., Salazar M.D., Contreras H., Nguyen V.H., Martinez J., Park S., Nguyen J., Kha M., Iniguez A., Zhou Q., Kaltcheva T., Levytskyy R., Ebelt N.D., Kang T.H., Wu X., Rogers T., Manuel E.R., Shostak Y., Diamond D.J, & Wussow F. (2020). Development of a Synthetic Poxvirus-Based SARS-CoV-2 Vaccine. bioRxiv.

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Quantitative Western Blot Analysis

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Ten microgram total protein was electrophoresed on a 4–15% Mini-PROTEAN TGX Gradient gel (Bio-Rad), transferred to nitrocellulose, and probed with anti-β-actin (C4, sc-47778), anti-HSC-70 (sc-1059), or anti-CUX1 (B-10, sc-514008) from Santa Cruz. Secondary antibodies were anti-mouse-HRP (A9044, Sigma) or anti-goat HRP (sc-2020, Santa Cruz), detected with Chemiluminescence Supersignal West Pico kit (Thermo Scientific), and quantified with ImageJ (23 (link)).

Arthur R.K., An N., Khan S, & McNerney M.E. (2017). The haploinsufficient tumor suppressor, CUX1, acts as an analog transcriptional regulator that controls target genes through distal enhancers that loop to target promoters. Nucleic Acids Research, 45(11), 6350-6361.

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SARS-CoV-2 Spike and Nucleocapsid Protein Detection

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BHK cells infected at 5 MOI were harvested 24-post infection. Proteins were solubilized in PBS with 0.1 % Triton X-100, supplemented with protease inhibitor, then reduced and denatured in Laemmli buffer containing DTT and boiled at 95 °C for ~ 10 minutes. Proteins were resolved on a 4–20% Mini Protean TGX gradient gel (BioRad), and transferred onto PVDF membrane. S protein was probed with anti-SARS-CoV-1 S1 subunit rabbit polyclonal antibody (40150-T62-COV2, Sino Biological); N protein was probed with anti-SARS-CoV1 NP rabbit polyclonal antibody (40413-T62, Sino Biological). Vaccinia BR5 protein was probed as a loading control. Anti-rabbit polyclonal antibody conjugated with horseradish peroxidase (Sigma-Aldrich) was used as a secondary antibody and protein bands were visualized with chemiluminescent substrate (ThermoFisher).

Chiuppesi F., Salazar M.D., Contreras H., Nguyen V., Martinez J., Park S., Nguyen J., Kha M., Iniguez A., Zhou Q., Kaltcheva T., Levytskyy R., Ebelt N., Kang T., Wu X., Rogers T., Manuel E., Shostak Y., Diamond D, & Wussow F. (2020). Development of a Multi-Antigenic SARS-CoV-2 Vaccine Using a Synthetic Poxvirus Platform. Research Square.

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Characterization of Biotinylated SPIONs by SDS-PAGE

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SDS-Gel electrophoresis according to the protocol of Laemmli^{40 (link)} was performed to show the binding of streptavidin
and avidin to biotinylated SPION. All SPION samples that contained
avidin or streptavidin were purified with Amicon 100 kDa centrifugation
units three times in order to remove excess unbound streptavidin or
avidin. To prove that avidin and streptavidin selectively bind to
biotinylated SPION and not to PEGylated SPION without biotin, we incubated
PEGylated SPION with avidin/streptavidin and subjected them
to the same purification process (3 × Amicon 100 kDa) as the
biotinylated SPION. All SPION were lyophilized and completely redispersed
at ∼1.5 mg SPION/mL in Laemmli buffer. Clear dispersions were
obtained for all SPION. All samples were split in two; one sample
was denatured at 100 °C for 3 min and the other sample remained
untreated. All samples were centrifuged (no pellet was observed upon
centrifugation), and the supernatant was applied to a BioRad Mini-Protean
TGX gradient gel (4–20% acrylamide). Separation was performed
at 200 V for 45 min. Gels were stained with 0.1% Coomassie Brilliant
Blue G in 40% methanol and 10% acetic acid for 15 min at 60 °C
and destained with 10% acetic acid until the background appeared colorless.
Gels were scanned using a Licor Odyssey Infrared Reader.

Lassenberger A., Scheberl A., Stadlbauer A., Stiglbauer A., Helbich T, & Reimhult E. (2017). Individually Stabilized, Superparamagnetic Nanoparticles with Controlled Shell and Size Leading to Exceptional Stealth Properties and High Relaxivities. ACS Applied Materials & Interfaces, 9(4), 3343-3353.

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Targeted Proteomics of Bacterial Lysates

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Bacterial lysates for targeted proteomics were prepared as described (Prigozhin et al.,2016 (link)). Lysates corresponding to 30 □g total protein were subjected to electrophoresis through a 4–20% mini PROTEAN TGX gradient gel (Bio-Rad) under denaturing conditions for approx. 2 cm into the gel. After staining with Novex Colloidal Blue staining kit (ThermoFisher Scientific), gel containing almost all of the proteins was excised, subjected to in-gel digestion with trypsin, and analyzed by LC/MS/MS as described (Shin et al., 2017 (link)). The PRM assay utilized three tryptic peptides of Mdh (residues 25–40 LASGSLLGPDRPIELR, 205–226 NAAEVVNDQAWIEDEFIPTVAK, and 236–252 GASSAASAASATIDAAR), and three tryptic peptides of SigA (238–251 VALLNAEEEVELAK, 343–352 FSTYATWWIR, and 488–500 TLDEIGQVYGVTR; all annotations as per http://tuberculist.epfl.ch). Three fragment ion intensities that generated the most robust signals were extracted from each peptide and summed to determine the peptide abundance. Quantification of targeted peptides was accomplished through the Skyline software (University of Washington). The average of peptide abundance from two technical replicates were determined and the relative level of Mdh was calculated by normalizing the total transition area of each of the 3 Mdh peptides to that of each of the 3 SigA peptides.

Rittershaus E.S., Baek S.H., Krieger I.V., Nelson S.J., Cheng Y.S., Nambi S., Baker R.E., Leszyk J.D., Shaffer S.A., Sacchettini J.C, & Sassetti C.M. (2018). A lysine acetyltransferase contributes to the metabolic adaptation to hypoxia in Mycobacterium tuberculosis. Cell chemical biology, 25(12), 1495-1505.e3.

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Native PAGE Immunoblotting Protocol

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For native polyacrylamide gel electrophoresis (PAGE), cells were cultured as described for conventional immunoblotting. Lysates were prepared in P100E2 buffer and sonicated for 10 seconds on ice, and after addition of native sample loading buffer (final concentrations: 31.25 mM Tris-HCl (pH 6.8), 12.5% glycerol, 0.5% bromophenol blue), 25 µg was separated on a 4% to 15% precast mini-PROTEAN TGX gradient gel (BIO-RAD) under native conditions (1× PAGE buffer: 25 mM Tris-base, 192 mM glycine, ~pH 8.3), and further processed as described for conventional immunoblots.

Groeneweg S., van den Berge A., Lima de Souza E.C., Meima M.E., Peeters R.P, & Visser W.E. (2020). Insights Into the Mechanism of MCT8 Oligomerization. Journal of the Endocrine Society, 4(8), bvaa080.

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Western Blot Analysis of p-Akt and Akt

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HCC38 cells were harvested using Triton lysis buffer containing 25mM HEPES, 100mM NaCl, 1mM EDTA, 10% glycerol and 1% TritonX-100 with 1X protease and phosphatase inhibitor added fresh prior to use. 50 μg of protein was loaded on 4–20% Mini-protean TGX gradient gel (Biorad) at 100V for 2 hours at room temperature and transferred onto nitrocellulose membrane at 100V for 1 hour at 4°C. The membranes were blocked using 5% milk solution, incubated with primary antibody against p-Akt, total Akt and beta-actin (Cell Signaling Technology) overnight at 4°C followed by incubation with HRP-conjugated secondary antibodies (Cell Signaling Technology) for 1 hour at room temperature. The signal was developed using SuperSignal West Pico Chemiluminescent Substrate (ThermoFisher Scientific), digitally imagined using the ChemiDoc Touch Imagining System (BioRad) and signaling intensity quantified by Image J software (https://imagej.nih.gov/ij/).

Mehta G.A., Parker J.S., Silva G.O., Hoadley K.A., Perou C.M, & Gatza M.L. (2017). Amplification of SOX4 promotes PI3K/Akt signaling in human breast cancer. Breast cancer research and treatment, 162(3), 439-450.

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SARS-CoV-1 Spike and Nucleocapsid Protein Detection

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BHK cells infected at 5 MOI were harvested 24 h post infection. Proteins were solubilized in PBS with 0.1% Triton X-100, supplemented with protease inhibitor, then reduced and denatured in Laemmli buffer containing DTT and boiled at 95 °C for ~10 min. Proteins were resolved on a 4–20% Mini Protean TGX gradient gel (BioRad) and transferred onto PVDF membrane. S protein was probed with anti-SARS-CoV-1 S1 subunit rabbit polyclonal antibody at a dilution of 1:1300 (40150-T62-COV2, Sino Biological); N protein was probed with anti-SARS-CoV1 NP rabbit polyclonal antibody (40413-T62, Sino Biological) was used at dilution of 1:10,000. Vaccinia B5R protein was probed as a loading control using 19C2 (ref. ^{58 (link)}) rat monoclonal antibody (hybridoma supernatant) at a dilution of 1:20. Anti-rabbit IgG polyclonal antibody and anti-rat IgG polyclonal antibody conjugated with horseradish peroxidase (A6154 and A5795, Sigma-Aldrich) were used as a secondary antibody at a dilution of 1:5000 and 1:3300, respectively. Protein bands were visualized with a chemiluminescent substrate (Thermo Fisher Scientific).

Chiuppesi F., Salazar M.D., Contreras H., Nguyen V.H., Martinez J., Park Y., Nguyen J., Kha M., Iniguez A., Zhou Q., Kaltcheva T., Levytskyy R., Ebelt N.D., Kang T.H., Wu X., Rogers T.F., Manuel E.R., Shostak Y., Diamond D.J, & Wussow F. (2020). Development of a multi-antigenic SARS-CoV-2 vaccine candidate using a synthetic poxvirus platform. Nature Communications, 11, 6121.

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