Complete edta free cocktail

Manufactured by Roche

Sourced in United States

COmplete EDTA-free cocktail is a laboratory reagent designed for protein extraction and stabilization. It is a ready-to-use solution that does not contain EDTA, a common chelating agent. The product is formulated to efficiently extract and preserve proteins from various biological samples while maintaining their native structure and functionality.

Automatically generated - may contain errors

Lab products found in correlation

Tissueruptor, by Qiagen (3 mentions) Protein g sepharose, by GE Healthcare (2 mentions) Tnt coupled transcription translation system, by Promega (1 mentions) Glutathione sepharose 4b beads, by GE Healthcare (1 mentions) 0.2 μm nitrocellulose membrane, by Bio-Rad (1 mentions) Anti mecp2, by Merck Group (1 mentions) Image studio lite software, by LI COR (1 mentions) Irdye 800cw donkey anti mouse igg, by LI COR (1 mentions) Anti γ tubulin, by Merck Group (1 mentions) Odyssey infrared imager, by LI COR (1 mentions) Nab protein a g spin column, by Thermo Fisher Scientific (1 mentions) Dynabeads protein a, by Thermo Fisher Scientific (1 mentions) Nupage gel, by Thermo Fisher Scientific (1 mentions) Nupage lds sample buffer, by Thermo Fisher Scientific (1 mentions) Mitochondria isolation kit, by Thermo Fisher Scientific (1 mentions) Chaps, by Merck Group (1 mentions) Gfp trap, by Proteintech (1 mentions) Phosphor screen, by GE Healthcare (1 mentions) Doxycycline, by Merck Group (1 mentions) Ab113685, by Abcam (1 mentions)

Close spelling variants of this product

Complete EDTA-free (281 citations) Cocktail Complete (7 citations) COmplete mini EDTA-free protease and PhosSTOP phosphatase inhibitor cocktails (2 citations)

11 protocols using complete edta free cocktail

Cutaneous Lesion Sampling in Infected Mice

Check if the same lab product or an alternative is used in the 5 most similar protocols

Skin samples from infected mice with visible lesions (displaying redness, edema, ulcer, necrosis, or healing) were excised (we removed 1 cm of cutaneous tissue from around the lesion) and crushed in phosphate-buffered saline (PBS) supplemented with cOmplete EDTA-free cocktail (Roche), with a TissueRuptor (QIAGEN). The resulting suspensions were centrifuged at 3500g for 10 min at 4°C. The supernatants were stored at −80°C. Blood samples were collected by retro-orbital puncture at each clinical stage of infection before tail excision. The blood was centrifuged at 1500g for 10 min to isolate serum, which was recovered and stored at −80°C.

Foulon M., Pouchin A., Manry J., Khater F., Robbe-Saule M., Durand A., Esnault L., Delneste Y., Jeannin P., Saint-André J.P., Croué A., Altare F., Abel L., Alcaïs A, & Marion E. (2020). Skin-specific antibodies neutralizing mycolactone toxin during the spontaneous healing of Mycobacterium ulcerans infection. Science Advances, 6(9), eaax7781.

+ Open protocol

+ Expand

In vitro Protein Interaction Assay

Check if the same lab product or an alternative is used in the 5 most similar protocols

Full-length HA-tagged wt-HDAC6, HA-HDAC6-ΔBUZ and BUZ domain mutants, or HA-tagged HIV-1 Vif proteins were expressed in vitro using the rabbit-reticulocytes system, whereas GST, GST-A3G (wt or its mutants) and GST-Vif proteins were expressed in BL21-DE3 E. coli cells. HA-tagged proteins were resuspended in RIPA 1X buffer (PBS 1X, 1% NP40, 0.5% sodium deoxycholate, 0.05% SDS) supplemented with protease inhibitors (complete EDTA Free cocktail, Roche), and then purified using anti-HA Agarose beads following the manufacturer’s recommendations. For extraction of GST proteins, bacteria cells were lysed in Lysis Buffer (50 mM Tris–HCl pH 7.4, 2 mM EDTA, 150 mM NaCl, 1 mM DTT, 1% Triton X-100, 30 μg/mL Lysozyme, and protease inhibitors) and then purified by affinity using GSH-Sepharose beads. 2 μg of purified HA-tagged protein were incubated overnight in RIPA buffer at 4°C with 3 μg of GST, GST-A3G (wt, C97A or D128K mutants), or GST-Vif proteins immobilized on glutathione-Sepharose beads. Then, beads were washed three times in RIPA buffer and the bound cellular proteins were analyzed by Western blotting using the appropriate antibodies. Hence, in vitro expressed, purified and pulled-down HA-tagged wt, ΔBUZ or BUZ constructs of HDAC6, and HIV-1 Vif proteins were analyzed using an anti-HA monoclonal antibody. Loading of GST proteins was analyzed using an anti-GST antibody.

Valera M.S., de Armas-Rillo L., Barroso-González J., Ziglio S., Batisse J., Dubois N., Marrero-Hernández S., Borel S., García-Expósito L., Biard-Piechaczyk M., Paillart J.C, & Valenzuela-Fernández A. (2015). The HDAC6/APOBEC3G complex regulates HIV-1 infectiveness by inducing Vif autophagic degradation. Retrovirology, 12, 53.

+ Open protocol

+ Expand

GST-Fusion Protein Purification and Interaction Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

Glutathione S-Transferase (GST) and GST-fusion proteins were purified from Escherichia coli BL21 (DE3) codon plus strain extracts, with glutathione-Sepharose 4B beads (GE Healthcare). Beads carrying the GST or the GST-fusion proteins were equilibrated in the following binding buffer (10 mM Tris-HCL pH 8, 250 mM NaCl, 0.1% NP40 and 2 mg/ml bovine serum albumin (BSA)) in the presence of protease inhibitors (complete EDTA-free cocktail from Roche Molecular Biochemicals) and incubated with radiolabeled proteins synthesized in vitro in the presence of [³⁵S]-methionine using the TnT Coupled Transcription/Translation system (Promega), in binding buffer for 4 h at 4°C. Beads were washed 5× in binding buffer without BSA and bound proteins were fractionated by sodium dodecylsulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and revealed by autoradiography.

Bazot Q., Deschamps T., Tafforeau L., Siouda M., Leblanc P., Harth-Hertle M.L., Rabourdin-Combe C., Lotteau V., Kempkes B., Tommasino M., Gruffat H, & Manet E. (2014). Epstein–Barr virus nuclear antigen 3A protein regulates CDKN2B transcription via interaction with MIZ-1. Nucleic Acids Research, 42(15), 9700-9716.

+ Open protocol

+ Expand

Co-Immunoprecipitation of FBXL8 Interactors

Check if the same lab product or an alternative is used in the 5 most similar protocols

48 h after treatment of the MCF7 and MDA-MB231 cells with FBXL8-siRNA or control siRNAs, Co-IP assay was performed. For Co-IP study with MCF7 or MDA-MB231 total cell lysates, the cells were lysed in CHAPS lysis buffer containing 20 mM Tris-HCl, pH 7.5, 5 mM MgCl2, 137 mM KCl, 1 mM EDTA, 1 mM EGTA, 1% CHAPS and 1 protease inhibitors (complete EDTA-free cocktail, Roche, St. Louis, MO, USA). Then, the cell lysate was precleared by incubating with protein G Sepharose (GE Healthcare, Little Chalfont, UK) at 4 °C for 2 h. The supernatant was incubated overnight at 4 °C with FBXL8 or control IgG_2b antibody, followed by a 3-h incubation with protein G Sepharose. The washed immunoprecipitates, resuspended in Laemmli buffer, was boiled at 95 °C for 5 min before immunodetection of FBXL8, CCND2 or IRF5.

Chang S.C., Hsu W., Su E.C., Hung C.S, & Ding J.L. (2020). Human FBXL8 Is a Novel E3 Ligase Which Promotes BRCA Metastasis by Stimulating Pro-Tumorigenic Cytokines and Inhibiting Tumor Suppressors. Cancers, 12(8), 2210.

+ Open protocol

+ Expand

Skin Biopsy Specimen Preparation

Check if the same lab product or an alternative is used in the 5 most similar protocols

Skin biopsy specimens were provided by the Centre de Diagnostic et de Traitement de la Lèpre et de l'Ulcère de Buruli (CDTLUB) of Pobé (Benin). The biopsy specimens were crushed in PBS supplemented with protease inhibitors (cOmplete EDTA-free cocktail, Roche), with a TissueRuptor (QIAGEN), as described for mouse tissues. The resulting suspensions were centrifuged at 3500g for 10 min at 4°C. The resulting supernatants were stored at −80°C.

+ Open protocol

+ Expand

Western Blot Analysis of MeCP2 Protein

Check if the same lab product or an alternative is used in the 5 most similar protocols

One frozen half brain was homogenized in ice-cold NE1 buffer [20 mm Hepes pH 7.9, 10 mm KCl, 1 mm MgCl₂, 0.1% Triton X-100, 20% glycerol, 0.5 mm DTT, protease inhibitor (complete EDTA free cocktail, Roche)] before adding 750 units of benzonase for 15 min at room temperature then an equal volume of 2× SDS loading buffer (0.125 M Tris pH 6.8, 20% glycerol, 4% SDS, 0.25% bromophenol blue, 20 mm DTT, 0.3 M β-mercaptoethanol). Samples were boiled for 3 min before loading equal volumes on a 4–12% Run Blue SDS precast gel (Expedeon). Gels were transferred to a 0.2-μm nitrocellulose membrane (Bio-Rad) and then blocked for 1 h (5% milk, 1× TBS, 0.1% Tween) before applying anti-Mecp2 1:1000 (Sigma M6818) or anti-γ-tubulin 1:3000 (Sigma T5326) overnight at 4°C, followed by IRDye 800CW donkey anti-mouse IgG 1:10 000 (LiCor) for 2 h at RT after washing. Membranes were imaged using the Odyssey Infrared Imager (LiCor) and quantified using Image Studio Lite Software (LiCor).

Brown K., Selfridge J., Lagger S., Connelly J., De Sousa D., Kerr A., Webb S., Guy J., Merusi C., Koerner M.V, & Bird A. (2015). The molecular basis of variable phenotypic severity among common missense mutations causing Rett syndrome. Human Molecular Genetics, 25(3), 558-570.

+ Open protocol

+ Expand

Purification of Ig from Infected Mouse Tails

Check if the same lab product or an alternative is used in the 5 most similar protocols

Tail skins from infected mice were excised and crushed with metallic beads and a TissueRuptor (QIAGEN) in an equal volume of PBS supplemented with proteases inhibitor (cOmplete EDTA-free cocktail, Roche) and protein A/G IgG Binding Buffer (Thermo Fisher Scientific). The resulting suspensions were centrifuged at 8000g for 45 min at 4°C. Ig were then purified with a NAb protein A/G spin column (Thermo Fisher Scientific), followed by a NAb protein L spin column (Thermo Fisher Scientific), according to the manufacturer’s instructions.

+ Open protocol

+ Expand

Co-immunoprecipitation of Vif Protein

Check if the same lab product or an alternative is used in the 5 most similar protocols

HEK 293T cells were co-transfected with pA3G-HA and plasmids expressing wild type or mutant Vif, Twenty-four h post-transfection, cells were washed in PBS (140 mM NaCl, 8 mM NaH₂PO₄, 2 mM Na₂HPO₄) and lysed in RIPA 1X (PBS, 1% NP40, 0.5% sodium deoxycholate, 0.05% SDS) supplemented with protease inhibitors (cOmplete EDTA Free cocktail, Roche). After centrifugation, an aliquot fraction (50 μl) was used for determination of the protein expression level, and the remaining was incubated 2 h at 4 °C with 1 μg of HA antibody (Santa Cruz, California, USA) on a rotating wheel. After equilibration, protein A Dynabeads (Life Technologies) were added and incubated for 1.5 h at 4 °C. Beads were washed 5 times with cold RIPA, and eluted in glycine pH 2.8, NuPAGE LDS sample buffer (Life Technologies). After 10 min at 70 °C, supernatant was loaded on NuPAGE gel (Life Technologies) and analyzed by western blot.

Guerrero S., Libre C., Batisse J., Mercenne G., Richer D., Laumond G., Decoville T., Moog C., Marquet R, & Paillart J.C. (2016). Translational regulation of APOBEC3G mRNA by Vif requires its 5′UTR and contributes to restoring HIV-1 infectivity. Scientific Reports, 6, 39507.

+ Open protocol

+ Expand

Cytosolic and Mitochondrial Protein Interactions

Check if the same lab product or an alternative is used in the 5 most similar protocols

To prepare cytosolic and mitochondrial lysates, BMDM cells were homogenized and mitochondria were pelleted by a mitochondria isolation kit (Thermo Fisher Scientific, Rockford, IL, USA). The cytosolic fractions were subjected to Co-IP with anti-Bax or anti-SARM antibody. The mitochondrial fraction products were immunoblotted and probed sequentially with antibodies to Bax, SARM and VDAC. For Co-IP study with J774 or RAW264.7 total cell lysates, the cells were lysed in CHAPS lysis buffer containing 20 mM Tris-HCl, pH 7.5, 5 mM MgCl₂, 137 mM KCl, 1 mM EDTA, 1 mM EGTA, 1% CHAPS and 1 × protease inhibitors (complete EDTA-free cocktail, Roche). Then, the cell lysate was precleared by incubating with protein G Sepharose (GE Healthcare, Little Chalfont, UK) at 4 °C for 2 h. The supernatant was incubated overnight at 4 °C with Bax or SARM or Bcl-2 antibody, followed by a 3-h incubation with protein G Sepharose. The washed immunoprecipitates, resuspended in Laemmli buffer, was boiled at 95 °C for 5 min before immunodetection of SAG, Bax, SARM, Bcl-2 or ubiquitin.

Chang S.C, & Ding J.L. (2014). Ubiquitination by SAG regulates macrophage survival/death and immune response during infection. Cell Death and Differentiation, 21(9), 1388-1398.

+ Open protocol

+ Expand

Immunoprecipitation from Yeast Cell Lysates

Check if the same lab product or an alternative is used in the 5 most similar protocols

Lysates for immunoprecipitations were prepared from indicated strains in mid-logarithmic growth grown in YPD reach media. Cells were harvested, washed in distilled water, and resuspended in lysis buffer (50 mM Tris HCl pH 7, 150 mM NaCl) supplemented with protease inhibitors (Complete EDTA-free cocktail; Roche) and frozen in a drop-by-drop fashion in liquid nitrogen. Frozen cells were then pulverized in a ball mill (1 min at 30 Hz; Retsch), thawed with nutation. Samples were thawed in 1ml lysis buffer supplemented with protease inhibitors and 1% CHAPS (Sigma Aldrich) at 4°C for 1 hour. All samples were then clarified by centrifugation at 14000 × g at 4°C for 15 minutes. The remaining supernatant was added to GFP-trap (Chromotek) for 1 hour followed by centrifugation at 1000 × g at 4°C for 3 minutes, and the supernatant was set aside as the flow through. Beads were washed three times with lysis buffer supplemented with protease inhibitors, and bound proteins were released from the beads by a 5 minute incubation in 95°C in sample buffer. The total protein lysate, the flow through and the immunoprecipitation (IP) fraction were analyzed by western blot.

Aviram N., Ast T., Costa E.A., Arakel E.C., Chuartzman S.G., Jan C.H., Haßdenteufel S., Dudek J., Jung M., Schorr S., Zimmermann R., Schwappach B., Weissman J.S, & Schuldiner M. (2016). The SND proteins constitute an alternative targeting route to the endoplasmic reticulum. Nature, 540(7631), 134-138.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!