Mg2 lysis buffer

Manufactured by Merck Group

Mg2+ lysis buffer is a reagent used in laboratory protocols to facilitate the disruption and lysis of cells. It contains magnesium ions (Mg2+) which are essential for the activation of cellular enzymes involved in the lysis process. The buffer is designed to maintain the stability of cellular components during lysis, allowing for the efficient extraction and purification of biomolecules from the lysed material.

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Lab products found in correlation

Chemidoc xr, by Bio-Rad (2 mentions) Tris glycine gel, by Bio-Rad (2 mentions) Phosphatase inhibitor, by Merck Group (1 mentions) Horse anti mouse igg, by Cell Signaling Technology (1 mentions) Ecl detection reagent, by GE Healthcare (1 mentions) Na934, by GE Healthcare (1 mentions) Rabbit anti nox4, by Santa Cruz Biotechnology (1 mentions) Mouse anti β actin, by Merck Group (1 mentions) Protease inhibitor, by Merck Group (1 mentions) Nitrocellulose membrane, by Bio-Rad (1 mentions) Bradford reagent, by Bio-Rad (1 mentions) Protease inhibitor cocktail, by Merck Group (1 mentions) Phosphatase inhibitor cocktail, by Merck Group (1 mentions)

Spelling variants of this product

Lysis buffer (324 citations) Mg2+ Lysis/Wash Buffer (2 citations) Mg2+ Lysis/Wash buffer (MLB) (2 citations)

4 protocols using mg2 lysis buffer

Rac/Cdc42 Activation Assay Protocol

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At 3 d after reperfusion, brains were isolated and lysed with Mg²⁺ lysis buffer according to the manufacturer's instruction (Millipore). Briefly, brian tissue lysates (1 mg/500 μl) were incubated with a Rac/cdc42 assay reagent (PAK1-PBD, agarose) at 4°C for 1 h. The pellets were collected by centrifugation at 14,000 g for 5 s, washed in lysis buffer, and resuspended in 40 μl of 2× Laemmli buffer. Samples were incubated for 15 min at 30°C with GTPγS as a positive control, while guanosine diphosphate as a negative control. Finally, co-IP was performed with Rac/cdc42 assay reagent (PAK1-PBD, agarose).

Zhou J., Zhang Y.H., Song H.Z., Ji H., Wang X.L., Wang L., Qian J., Ling J.J, & Ping F.F. (2016). 5d, a novel analogue of 3-n-butylphthalide, decreases NADPH oxidase activity through the positive regulation of CK2 after ischemia/reperfusion injury. Oncotarget, 7(26), 39444-39457.

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Rac1 Activation Assay in Cortex

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Total somatosensory cortex lysates from wild-type and KO animals were homogenized with Mg²⁺ lysis buffer (Millipore) with a complete protease inhibitor cocktail. The expression level of Rac1-GTP was then assessed using a Rac/Cdc42 pull-down kit (Millipore). Samples (100 mg) were incubated and rocked with 10 mg of glutathione S-transferase (GST)-tagged PAK-PBD agarose beads (Millipore) for 2 hours. Beads were pelleted by centrifugation (14,000g for 15 s at 4 °C), and the supernatant was discarded. The pelleted beads were washed with lysis buffer three times, resuspended in 50 ml of 2× Laemmli buffer, boiled for 5 min, and subjected to Western blot analysis. GTP and guanosine diphosphate (GDP) loading controls were incubated with 100 μM GTP-gS or 1 mM GDP for 30 min at 30°C.

Pyronneau A., He Q., Hwang J.Y., Porch M., Contractor A, & Zukin R.S. (2017). Aberrant Rac1-cofilin signaling mediates defects in dendritic spines, synaptic function, and sensory perception in fragile X syndrome. Science signaling, 10(504), eaan0852.

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Western Blotting for NOX4 Protein Detection

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Western blotting was performed as previously described [26 ]. Immediately after enucleation, retinas were dissected and lysed in 1X Mg2+ lysis buffer (EMD Millipore) with a protease inhibitor (Sigma Aldrich, St. Louis, MO) and a phosphatase inhibitor (EMD Millipore). Samples were quantified with Bradford Reagent (Bio-Rad) [27 (link)]. 40 μg of total protein were separated on 4–20% Tris-glycine gel (Bio-Rad), transferred to nitrocellulose membrane (Bio-Rad), and blocked with 5% BSA diluted in Tris-buffered saline supplemented with Tween-20 (TBST) for one hour. Membranes were incubated with rabbit anti-NOX4 (1:200 dilution, sc-30141, Santa Cruz, Dallas, TX) for 18 hours or mouse anti-β-actin (1:5000 dilution, A5316, Sigma-Aldrich) diluted in 5% BSA in TBST for one hour. After washing, membranes were incubated with donkey anti-rabbit (1:5000 dilution, NA934, GE Healthcare, Piscataway, NJ) or horse anti-mouse IgG (1:5000 dilution, 7076, Cell Signaling, Boston, MA) and exposed with ECL Detection Reagents (GE Healthcare). Bands were analyzed using ChemiDoc XRS (Bio-Rad) and normalized to β-actin. Data are fold change ± SEM.

Lajko M., Cardona H.J., Taylor J.M., Farrow K.N, & Fawzi A.A. (2017). Photoreceptor oxidative stress in hyperoxia-induced proliferative retinopathy accelerates rd8 degeneration. PLoS ONE, 12(7), e0180384.

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Retinal Protein Extraction and Western Blot

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Retinas were processed as described previously [34 ]. Briefly, immediately after enucleation, retinas were dissected and lysed in 1X Mg2+ lysis buffer (EMD Millipore, Billerica, MA) with a protease inhibitor cocktail (Sigma Aldrich, St. Louis, MO) and a phosphatase inhibitor cocktail (EMD Millipore). Protein concentration of the supernatant was quantified using the Bradford method [35 (link)]. 40 μg of protein were separated on 4–20% Tris-glycine gel (Bio-Rad, Hercules, CA), transferred to nitrocellulose membrane, and blocked at room temperature with 5% BSA diluted in Tris-buffered saline supplemented with Tween-20 (TBST) for 1 hour. Membranes were incubated with primary antibodies (Table 1) diluted in 5% BSA in TBST for 18 hours at 4°C. After washing, membranes were incubated with secondary antibodies (Table 1) for 2 hours and exposed using chemiluminescence (GE Healthcare, Little Chalfont, UK). Bands were analyzed using ChemiDoc XRS (Bio-Rad) and normalized to β-actin. Data are fold change relative to room air controls ± standard error of mean (SEM).

Lajko M., Cardona H.J., Taylor J.M., Shah R.S., Farrow K.N, & Fawzi A.A. (2016). Hyperoxia-Induced Proliferative Retinopathy: Early Interruption of Retinal Vascular Development with Severe and Irreversible Neurovascular Disruption. PLoS ONE, 11(11), e0166886.

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