Tgx acrylamide stain free gels

Manufactured by Bio-Rad

The TGX Acrylamide Stain-Free gels are a line of pre-cast polyacrylamide gels designed for protein electrophoresis. These gels contain a proprietary stain-free technology that allows protein visualization without the need for traditional staining methods.

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Lab products found in correlation

Clarity western ecl substrate, by Bio-Rad (6 mentions) Horseradish peroxidase conjugated secondary antibody, by Jackson ImmunoResearch (5 mentions) Pvdf membrane, by Merck Group (4 mentions) Image lab software, by Bio-Rad (3 mentions) Chemidoc imaging system, by Bio-Rad (3 mentions) Gapdh, by Fortis Life Sciences (3 mentions) Pkr thr 446 p, by Abcam (3 mentions) Adar1, by Santa Cruz Biotechnology (3 mentions) Chemidoc touch imaging system, by Bio-Rad (2 mentions) Dc protein assay, by Bio-Rad (2 mentions) Polyvinylidene difluoride membrane, by Merck Group (1 mentions) Goat anti mouse igg, by Bio-Rad (1 mentions) Hrp conjugated goat anti rabbit igg, by Bio-Rad (1 mentions) Geldoc go imaging system, by Bio-Rad (1 mentions) Blocking one, by Nacalai Tesque (1 mentions) Protoglow ecl, by National Diagnostics (1 mentions)

Spelling variants of this product

TGX gels (272 citations) Acrylamide (183 citations) 12% acrylamide gel (109 citations) TGX stain-free gels (84 citations) Stain-free gels (51 citations) 4–20% stain-free TGX gels (2 citations) TGX Stain-Free™ FastCast™ 12% Acrylamide Gels (2 citations)

8 protocols using tgx acrylamide stain free gels

Western Blot Analysis of Signaling Proteins

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Cottrell K.A., Chiou R.C., & Weber J.D. (2020). ARF suppresses 5’-terminal oligopyrimidine mRNA translation.

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Immunoblotting Quantification of ADAR1, PKR, and pPKR

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Cell pellets were lysed and sonicated in RIPA Buffer (50 mM Tris pH 7.4, 150 mM NaCl, 1% Triton X-100, 0.1% sodium dodecyl sulfate and 0.5% sodium deoxycholate) with 1x HALT Protease Inhibitor (Pierce).
Forty micrograms of protein lysate were resolved on 4-12% TGX Acrylamide Stain-Free gels (Bio-Rad).
Proteins were transferred to PVDF membrane (Millipore). The membrane was cut into strips corresponding to the molecular weight of proteins of interest. The blots were blocked and then probed with the appropriate primary antibodies: Primary antibodies: ADAR1 (Santa Cruz, sc-73408), PKR (Cell Signaling, #3072), PKR Thr-446-P (Abcam, ab32036), GAPDH (Bethyl, A300-641A). Primary antibodies were detected with horseradish-peroxidase conjugated secondary antibodies (Jackson ImmunoResearch) and detection was carried out with Clarity Western ECL Substrate (Bio-Rad). Chemiluminescence was imaged using a ChemiDoc imaging system (Bio-Rad). Quantification of immunoblots was performed using Image Lab software (Bio-Rad).
All proteins were normalized to GAPDH abundance. For PKR and pPKR, two separate gels were resolved, transferred, and probed for either PKR or pPKR in addition to GAPDH for both. PKR and pPKR abundance were normalized to GAPDH prior to normalizing pPKR to PKR. Uncropped immunoblot images are available in Supplemental Figures 12345.

Cottrell K.A., Torres L.S., & Weber J.D. (2021). 8-azaadenosine and 8-chloroadenosine are not selective inhibitors of ADAR.

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Western Blot Quantification of ADAR1, PKR, and pPKR

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Cell pellets were lysed and sonicated in RIPA Buffer (50 mmol/L Tris pH 7.4, 150 mmol/L NaCl, 1% Triton X-100, 0.1% SDS and 0.5% sodium deoxycholate) with 1× HALT Protease Inhibitor (Pierce). Forty micrograms of protein lysate were resolved on 4%–12% TGX Acrylamide Stain-Free gels (Bio-Rad). Proteins were transferred to polyvinylidene difluoride membrane (Millipore). The membrane was cut into strips corresponding to the molecular weight of proteins of interest. The blots were blocked and then probed with the appropriate primary antibodies: ADAR1 (Santa Cruz Biotechnology, sc-73408), PKR (Cell Signaling Technology, #3072), PKR Thr-446-P (Abcam, ab32036), GAPDH (Bethyl Laboratories, A300–641A). Primary antibodies were detected with horseradish peroxidase–conjugated secondary antibodies (Jackson ImmunoResearch) and detection was carried out with Clarity Western ECL Substrate (Bio-Rad). Chemiluminescence was imaged using a ChemiDoc imaging system (Bio-Rad). Quantification of immunoblots was performed using Image Lab software (Bio-Rad). The abundance of each protein was normalized to GAPDH abundance. For PKR and pPKR, two separate gels were resolved, transferred, and probed for either PKR or pPKR in addition to GAPDH for both. PKR and pPKR abundance were normalized to GAPDH prior to normalizing pPKR to PKR. Uncropped immunoblot images are available in Supplementary Figs. S1–S5.

Cottrell K.A., Soto-Torres L., Dizon M.G, & Weber J.D. (2021). 8-Azaadenosine and 8-Chloroadenosine are not Selective Inhibitors of ADAR. Cancer Research Communications, 1(2), 56-64.

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Western Blot Analysis of Cellular Proteins

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Cell pellets were lysed and sonicated in RIPA Buffer (50 mM Tris pH 7.4, 150 mM NaCl, 1% Triton X-100, 0.1% sodium dodecyl sulfate and 0.5% sodium deoxycholate) with 1 × HALT Protease Inhibitor (Pierce). Seventy-five micrograms of protein lysate were resolved on 4–12% TGX Acrylamide Stain-Free gels (Bio-Rad). Stain-Free gels were imaged prior to transfer to PVDF membrane (Millipore). The blots were then probed with the appropriate primary antibodies: Abcam PABP (ab21060); Bethyl—GAPDH (A300-641A), LARP1 (A302-087A), EEF2 (A301-688A), RPL23A (A303-932A-T); Cell Signaling Technologies—TPT1(5128), p-MTOR (2971), p-S6 (2215S), p-S6K (9205S), p-4EBP1 (2855S), eIF4G1 (2858), p53 (2524S), eIF4E (9742S), 4EBP1 (9452S); Santa Cruz Biotechnology—ARF (sc-32748), RPL22 (sc-136413), University of Iowa, DSHB—puromycin. Primary antibodies were detected with horseradish-peroxidase conjugated secondary antibodies (Jackson ImmunoResearch) and detection was carried out with Clarity Western ECL Substrate (Bio-Rad).

Cottrell K.A., Chiou R.C, & Weber J.D. (2020). Upregulation of 5′-terminal oligopyrimidine mRNA translation upon loss of the ARF tumor suppressor. Scientific Reports, 10, 22276.

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Immunoblot Quantification of ADAR1, PKR, and pPKR

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Cell pellets were lysed and sonicated in RIPA Buffer (50 mM Tris pH 7.4, 150 mM NaCl, 1% Triton X-100, 0.1% sodium dodecyl sulfate and 0.5% sodium deoxycholate) with 1x HALT Protease Inhibitor (Pierce). Forty micrograms of protein lysate were resolved on 4–12% TGX Acrylamide Stain-Free gels (Bio-Rad). Proteins were transferred to PVDF membrane (Millipore). The membrane was cut into strips corresponding to the molecular weight of proteins of interest. The blots were blocked and then probed with the appropriate primary antibodies: Primary antibodies: ADAR1 (Santa Cruz, sc-73408), PKR (Cell Signaling, #3072), PKR Thr-446-P (Abcam, ab32036), GAPDH (Bethyl, A300–641A). Primary antibodies were detected with horseradish-peroxidase conjugated secondary antibodies (Jackson ImmunoResearch) and detection was carried out with Clarity Western ECL Substrate (Bio-Rad). Chemiluminescence was imaged using a ChemiDoc imaging system (Bio-Rad). Quantification of immunoblots was performed using Image Lab software (Bio-Rad). The abundance of each protein was normalized to GAPDH abundance. For PKR and pPKR, two separate gels were resolved, transferred, and probed for either PKR or pPKR in addition to GAPDH for both. PKR and pPKR abundance were normalized to GAPDH prior to normalizing pPKR to PKR. Uncropped immunoblot images are available in Supplemental Figures 1–5.

Cottrell K.A., Soto-Torres L., Dizon M.G, & Weber J.D. (2021). 8-Azaadenosine and 8-Chloroadenosine are not Selective Inhibitors of ADAR. Cancer Research Communications, 1(2), 56-64.

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Western Blot Analysis of MFSD2A and FABP5

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Equivalent amounts of protein from each sample were electrophoretically separated on 4–15% TGX Stain-Free gradient acrylamide gels (Bio-Rad, Hercules, CA; 4568084) or 12% TGX Stain-Free acrylamide gels (Bio-Rad; 161–0185) and transferred to low fluorescent polyvinylidene difluoride membranes (Bio-Rad; 1704274). Stain-Free technology using GelDoc go imaging system (Bio-Rad) was used for total protein normalization. Membranes were then blocked using Blocking One (Nacalai Tesque; 03953–95). MFSD2A, FABP5 and β-actin were detected using antibodies against MFSD2A (1:1,000; Sigma; SAB3500576), FABP5 (1:1,000; Sigma; SAB1401130) and β-actin (1:8,000; Sigma; A1978). After washing, the membranes were incubated in HRP–conjugated goat anti-rabbit IgG (Bio-Rad; 170–6515) or goat anti-mouse IgG (Bio-Rad; 170–6516), as appropriate. Immunoreactive bands were detected using Clarity Western ECL Substrate (Bio-Rad; 1705061). Images of the bands were digitally captured using a MultiImager II ChemiBOX (BioTools, Gunma, Japan), and band intensities were quantified using ImageJ software (National Institutes of Health Image, Bethesda, MD, USA). The relative intensity of each individual protein was expressed as the ratio of the corresponding protein to the total protein loading or β-actin.

Iwao T., Takata F., Matsumoto J., Aridome H., Yasunaga M., Yokoya M., Kataoka Y, & Dohgu S. (2023). Aging decreases docosahexaenoic acid transport across the blood-brain barrier in C57BL/6J mice. PLOS ONE, 18(2), e0281946.

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Quantitative Western Blot Analysis

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Cells were lysed in 2X Laemmli buffer and stored at -20°C until use. The DC Protein Assay (Bio-Rad) was used to quantify protein concentration as per the manufacturer’s instructions. 10–15 μg of protein lysate was resolved by SDS-PAGE on TGX Stain-Free acrylamide gels (BioRad). Total protein images were acquired from the PVDF membranes after transfer on the ChemiDoc Touch Imaging system (BioRad). Membranes were blocked in 5% BSA in TBS-T (Tris-buffered saline 0.1% Tween-20). Primary and secondary antibodies were diluted in 2.5% BSA, dilutions can be found in Table 2. Membranes were visualized using Clarity Western ECL substrate and the ChemiDoc Touch Imaging system (BioRad). Densitometry was conducted using ImageJ, area under the curve for each band was calculated, normalized to the respective beta actin loading control band, and presented as fold change.

Kleer M., Mulloy R.P., Robinson C.A., Evseev D., Bui-Marinos M.P., Castle E.L., Banerjee A., Mubareka S., Mossman K, & Corcoran J.A. (2022). Human coronaviruses disassemble processing bodies. PLoS Pathogens, 18(8), e1010724.

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Quantitative Protein Analysis Protocol

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Cells were lysed in 1× Laemmli buffer (4% SDS, 20% glycerol, 120 mM Tris-Cl [pH 6.8] and ddH₂O) and stored at −20°C until use. The DC protein assay (Bio-Rad) was used to quantify total protein concentration as per the manufacturer’s instructions.10 to 15 μg of protein lysate was resolved by SDS-PAGE on TGX Stain-Free acrylamide gels (Bio-Rad). Membranes were blocked in 5% bovine serum albumin (BSA) or 5% skim milk in Tris-buffered saline-Tween 20 (TBS-T). Primary and secondary antibodies were diluted in 2.5% BSA or 2.5% skim milk, dilutions can be found in Table 1. Membranes were visualized using ProtoGlow ECL (National Diagnostics) and the ChemiDoc Touch Imaging system (Bio-Rad).

Kleer M., MacNeil G., Adam N., Pringle E.S, & Corcoran J.A. (2022). A Panel of Kaposi’s Sarcoma-Associated Herpesvirus Mutants in the Polycistronic Kaposin Locus for Precise Analysis of Individual Protein Products. Journal of Virology, 96(5), e01560-21.

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