Memcode reversible protein stain

Manufactured by Thermo Fisher Scientific

MemCode Reversible Protein Stain is a laboratory reagent used to detect and visualize proteins in polyacrylamide gels. It provides a reversible staining method that allows for subsequent protein analysis or recovery.

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Lab products found in correlation

Odyssey infrared scanner, by LI COR (1 mentions) Chemidoc mp gel imager, by Bio-Rad (1 mentions) Any kd mini protean tgx precast gel, by Bio-Rad (1 mentions) S adenosyl l methyl 3h methionine, by PerkinElmer (1 mentions) Amicon ultra centrifugal filter, by Merck Group (1 mentions) En3hance spray surface autoradiography enhancer, by PerkinElmer (1 mentions) Image lab v4, by Bio-Rad (1 mentions) Chemidoc xrs imaging system, by Bio-Rad (1 mentions) Dc protein assay kit, by Bio-Rad (1 mentions) Schneider s drosophila medium, by Thermo Fisher Scientific (1 mentions) Immobilon fl, by Merck Group (1 mentions) Imagej, by LI COR (1 mentions) Odyssey clx, by LI COR (1 mentions) Bolt bis tris gel, by Thermo Fisher Scientific (1 mentions)

Spelling variants of this product

MemCode Reversible Protein Stain Kit (13 citations) MemCode Reversible Stain (11 citations) MemCode (11 citations) Pierce MemCode Reversible Protein Stain Kit (2 citations) Reversible protein stain-MemCode (MemC) (2 citations)

5 protocols using memcode reversible protein stain

Protein Staining and Imaging Protocols

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PVDF membranes were stained for total protein with Memcode Reversible Protein Stain (Thermo Scientific) according to the manufacturer’s recommendations. Erasure of the Memcode stain was performed with the Eraser/Methanol solution for 20 minutes. For total-protein staining with Ponceau S, membranes were incubated with 0.1% (w/v) Ponceau S in 5% acetic acid for 5 minutes and washed 2 × 5 minutes in 10% acetic acid, followed by washes of 5 minutes in 100% methanol and 5 minutes in a 70/30/4 volume ratio of methanol, acetic acid, and PEG-400. For erasing, Ponceau S-stained membranes were treated with 0.1 N NaOH for 30 seconds and washed with running deionized water for two minutes.
For both total-protein stains, digital images were captured on a ChemiDoc MP gel imager (Bio-Rad) with “Colorimetric” settings (2×2 camera binning). Fluorescence images were obtained on an Odyssey infrared scanner (LI-COR) at 169 µm resolution and 0 mm focus offset, with a 700-channel intensity of 5.0 and an 800-channel intensity of 8.0.

, & Janes K.A. (2015). An Analysis of Critical Factors for Quantitative Immunoblotting. Science signaling, 8(371), rs2.

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Characterizing ALK Kinase Domain Methylation

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A C‐terminal portion of ALK (a.a. 1058–1620) including a TKD was subcloned and ligated with pCAGGSn‐C‐3FLAG. The 293T cells were transfected with WT ALK‐TKD expression vector (pCAGGSn‐3FLAG–ALK‐TKD) or substituted types of ALK‐TKD expression vector (pCAGGSn‐3FLAG–ALK‐TKD K1451A, K1455A, or K1610A). After 48 h of the culture, the protein was immunoprecipitated with anti‐FLAG antibody and purified with Amicon Ultra Centrifugal Filters (Millipore, Billerica, MA, USA). For the in vitro methyltransferase assay, recombinant ALK‐TKD‐WT, ALK‐TKD‐K1451A, ALK‐TKD‐K1455A, or ALK‐TKD‐K1610A was separately incubated with SMYD2 enzyme using 2 μCi S‐adenosyl‐l‐[methyl‐³H]‐methionine (PerkinElmer, Waltham, MA, USA) as the methyl donor in a mixture of 10 μL methylase activity buffer (50 mM Tris‐HCl at pH 8.8, 10 mM DTT, and 10 mM MgCl₂) for 2 h at 30°C. Proteins were resolved on a Mini PROTEAN TGX Precast gel (Any kD; Bio‐Rad, Hercules, CA, USA) and visualized by fluorography using EN3HANCE Spray Surface Autoradiography Enhancer (PerkinElmer). Loading proteins were visualized by MemCode Reversible Protein Stain (Thermo Fisher Scientific).

Wang R., Deng X., Yoshioka Y., Vougiouklakis T., Park J., Suzuki T., Dohmae N., Ueda K., Hamamoto R, & Nakamura Y. (2017). Effects of SMYD2‐mediated EML4‐ALK methylation on the signaling pathway and growth in non‐small‐cell lung cancer cells. Cancer Science, 108(6), 1203-1209.

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Quantification of Lethal Factor in Anthrax

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Relative amounts of LF were quantified using previously described methods [33] . Briefly, the exotoxin-containing supernatants used in the cytotoxicity assays were quantified using the D_C Protein Assay Kit (Bio-Rad, Hercules CA), normalized to 40 µg in a final volume of 30 µL of 1x Laemmli buffer, and heated at 95°C for 4 minutes. Total protein amounts were measured by staining with MemCode Reversible Protein stain (Thermo Scientific, Rockford IL) and relative amounts quantified by densitometry using the ChemiDoc XRS Imaging system and Image lab v4.0 (Bio-Rad, Hercules CA). The membrane was blocked in a 4% Non-fat milk/TBS +0.05% Tween-20 (TBS-T) solution at 4°C overnight and then incubated for 1 hour with 1∶2500 α-LF (DD-6; BEI, Manassas, VA). After washing with TBS-T, the membranes were incubated for 1 hour with 1∶1000 goat α-mouse IgG Alkaline phosphatase. Bands were developed by incubating with Immuno BCIP/NBT substrate solution (MP Biomedicals, Solon OH) until bands were clearly visible. The amount of LF was then quantified using densitometry via the ChemiDoc XRS Imaging system and Image lab v4.0 (Bio-Rad, Hercules CA).
Relative quantitation was calculated using the following formula:

Lowe D.E., Ya J, & Glomski I.J. (2014). In Trans Complementation of Lethal Factor Reveal Roles in Colonization and Dissemination in a Murine Mouse Model. PLoS ONE, 9(4), e95950.

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Quantitative Western Blot Analysis of MMP-1

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VNCs were dissected in Schneider’s Drosophila Medium (ThermoFisher, #21720001) and homogenized in 3 μL 1x Loading Buffer per VNC. Protein lysate of 5–6 VNCs were loaded onto 4–20% Tris-Glycine gels (Lonza, #59517) and transferred to Immobilon-FL (Millipore, #IPFL00010). After transfer, total protein density per lane was measured using MemCode Reversible Protein Stain (ThermoFisher, #24585). Blots were probed with mouse anti-MMP-1 (1:100 at a 1:1:1:1 ratio) and incubated overnight at 4°C, washed several times with 1xPBS/0.01% Tween 20, and probed with appropriate fluorophore-conjugated antibodies secondary antibody at 1:2000 (Jackson Immunoresearch, #715-625-150 RRID:AB_2340868) for 2 hr at room temperature. Additional washes were performed with 1xPBS/0.01% Tween 20 and a final wash in 1xPBS. Total protein stain blots were imaged on G:BOX F3 Imaging System and analyzed with ImageJ (RRID:SCR_003070); fluorescent blots were imaged on Li-cor Odyssey CLx (RRID:SCR_014579) quantitative western blot imaging system and data was quantified using LiCor Image Studio software (RRID:SCR_014211). Images in Figure 5 has been cropped for presentation. Full size image is presented in Figure 5—figure supplement 1.

Purice M.D., Ray A., Münzel E.J., Pope B.J., Park D.J., Speese S.D, & Logan M.A. (2017). A novel Drosophila injury model reveals severed axons are cleared through a Draper/MMP-1 signaling cascade. eLife, 6, e23611.

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Western Blot Analysis of Hippocampal Protein

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All western blotting was carried out as previously described^{84 (link)}. Briefly, hippocampal tissue was homogenized in lysis buffer protease and phosphatase inhibitors. Protein concentration was measured using BCA assay (Pierce). Aliquots of protein (20–40 μg) were separated using Bolt Bis-Tris gels (4–12%; Thermo Scientific) and transferred to a nitrocellulose membrane. Membranes were washed and probed for total protein using MemCode Reversible Protein Stain (Thermo Scientific), before the expression of the protein of interest was probed using appropriate primary and secondary antibody pairs. All primary antibodies were incubated with the membranes overnight at 4 °C, rocking gently. All secondary antibodies were incubated with the membranes for 1 h at room temperature. Antibody signal was detected using chemiluminescence (GE Healthcare). Density of bands corresponding to the specified molecular weight for each target protein were normalized to the density of individual lanes of total protein as detected using MemCode or Ponceau staining.

Elder M.K., Erdjument-Bromage H., Oliveira M.M., Mamcarz M., Neubert T.A, & Klann E. (2021). Age-dependent shift in the de novo proteome accompanies pathogenesis in an Alzheimer’s disease mouse model. Communications Biology, 4, 823.

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