A6782

Manufactured by Merck Group

Sourced in United States

The A6782 is a laboratory equipment product. It is a multi-purpose device that can be used for various tasks in a research or scientific setting. The core function of the A6782 is to perform specific laboratory operations, but a detailed description cannot be provided while maintaining an unbiased and factual approach.

Automatically generated - may contain errors

Lab products found in correlation

7 protocols using a6782

Detecting S1PT Antigen Expression in rBCG Strains

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

To confirm the expression of the S1PT antigen in rBCG-S1PT, rBCG-S1i, and rBCG-S1+S1i, the rBCG strains were grown in MB7H9 until mid-log phase when cells where centrifuged and washed twice with PBS. Cells were resuspended in PBS and lysed by sonication. Total protein extracts were separated by centrifugation into soluble and insoluble fractions and separated by SDS-PAGE. Protein extracts were electrotransferred to a PVDF membrane using a semidry electroblotter (Owl Separation Systems) and blocked with 5% nonfat dry milk solution for 16 h at 4°C. Mouse polyclonal anti-S1PT generated in-house (1:1,000) was used for antigen detection incubating the membrane for 2 h. The secondary antibody, goat anti-IgG HRP was incubated at 1:2,000 for 1 h (A6782, Sigma). Peroxidase reaction was detected using the ECL Prime Detection Reagent (GE) and a LAS4000 photoimaging equipment (GE).

Kanno A.I., Goulart C., Leite L.C., Pagliarone A.C, & Nascimento I.P. (2019). A Bivalent Recombinant Mycobacterium bovis BCG Expressing the S1 Subunit of the Pertussis Toxin Induces a Polyfunctional CD4+ T Cell Immune Response. BioMed Research International, 2019, 9630793.

+ Open protocol

+ Expand

Western Blot Analysis of Cellular Proteins

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

Whole cell lysates were prepared by direct lysis of cells in 1X RIPA buffer (150 mM NaCl, 1.0% IGEPAL® CA-630, 0.5% sodium deoxycholate, 0.1% SDS, 50 mM Tris, pH 8.0) (Sigma-Aldrich, R0278) supplemented with 1X Protease Inhibitor Cocktail (Sigma-Aldrich, P8340) and the phosphatase inhibitors sodium fluoride (10 mM) (Sigma-Aldrich, S1504) and sodium orthovanadate (1 mM) (Sigma-Aldrich, 450243). Lysates were then subjected to SDS-polyacrylamide gel electrophoresis (PAGE) and wet transfer (BioRad) to a nitrocellulose membrane (BioRad), prior to detection with primary and secondary antibodies diluted in 5% milk/PBS-Tween 20 (Sigma-Aldrich, P7949). Western blots were developed using an ECL detection kit (BioRad Clarity^TM 1705061). Primary antibodies used were: mouse anti-β-actin (Sigma-Aldrich, A1978), rabbit anti-CUL1 (Abcam, ab75812), and rabbit anti-NEDD8 (Abcam, ab81264). Secondary antibodies purchased from Sigma-Aldrich were sheep anti-mouse (A6782) and goat anti-rabbit (A6154) horse-radish peroxidase.

Chung D., Salsman J, & Dellaire G. (2019). Inhibition of neddylation induces mitotic defects and alters MKLP1 accumulation at the midbody during cytokinesis. Cell Cycle, 18(10), 1135-1153.

+ Open protocol

+ Expand

Western Blot Analysis of Microglia and Cerebellar Proteins

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

Total protein was extracted from microglia by scraping the cells in SDS/sample buffer. Tissue from cerebellar organotypic slices was directly heated at 100°C in sample buffer (7 min). Samples were loaded and size‐separated by electrophoresis using Criterion TGX Precast 12% gels and transferred to Trans‐Blot Turbo Midi PVDF Transfer Packs (Bio‐Rad, Hercules, USA). Membranes were blocked in 5% skimmed milk and 5% serum in Tris‐buffered saline/0.05% Tween‐20 (TBS‐T) and proteins detected by specific primary antibodies to BDNF (#sc‐547, 1:200; Santa Cruz), to MBP (#SMI‐99P, 1:2,000, Covance), to GAPDH (#MAB374, 1:2,000; Millipore), and to β‐actin (#A2066, 1:1,000; Sigma), followed by secondary peroxidase‐coupled goat anti‐rabbit antibodies (#A6154, 1:2,000; Sigma) or sheep anti‐mouse antibodies (#A6782, 1:2,000; Sigma). After washing, blots were developed using an enhanced chemiluminescence detection kit according to the manufacturer's instructions (SuperSignal West Dura or Femto, Pierce). Images were acquired with a ChemiDoc MP system (Bio‐Rad) and quantified using ImageJ software. Values of BDNF and MBP were normalized to corresponding β‐actin and GAPDH signal, respectively.

Zabala A., Vazquez‐Villoldo N., Rissiek B., Gejo J., Martin A., Palomino A., Perez‐Samartín A., Pulagam K.R., Lukowiak M., Capetillo‐Zarate E., Llop J., Magnus T., Koch‐Nolte F., Rassendren F., Matute C, & Domercq M. (2018). P2X4 receptor controls microglia activation and favors remyelination in autoimmune encephalitis. EMBO Molecular Medicine, 10(8), e8743.

+ Open protocol

+ Expand

Recombinant BCG Expression of LTAK63

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

Competent BCGΔlysA were prepared as previously described and transformed with pAN71-ltak63-lysA(t) and pAN71-ltak63-lysA(c), generating rBCGΔ-LTAK63(t or c), respectively. Selected clones were grown in MB7H9-OADC until an OD₆₀₀ 1.0, when bacteria were recovered, and protein extracts used to detect the expression of LTAK63 by Western blot (20 (link)). Detection of LTAK63 was performed using anti-serum of mice previously immunized with rLTK63 (1:1,000) incubated for 60 min and an anti-mouse IgG antibody conjugated with peroxidase (A6782, 1:3,000 Sigma-Aldrich^®) incubated for 60 min. Additionally, growth curves of complemented auxotrophs were compared to wild-type BCG to determine whether rBCGΔ-LTAK63(t or c) would show altered in vitro growth.

Moraes L., Trentini M.M., Fousteris D., Eto S.F., Chudzinski-Tavassi A.M., Leite L.C, & Kanno A.I. (2022). CRISPR/Cas9 Approach to Generate an Auxotrophic BCG Strain for Unmarked Expression of LTAK63 Adjuvant: A Tuberculosis Vaccine Candidate. Frontiers in Immunology, 13, 867195.

+ Open protocol

+ Expand

Quantification of Synaptic Proteins in AD Models

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

Protein lysates from cortical neurons, 3xTg-AD mice and human brains were separated by SDS-PAGE using 7.5% Tris-Glycine polyacrylamide gels (Bio-Rad). Electrophoresis was conducted in a Tris-Glycine buffer (25 mM Tris, pH 8.3, 192 mM glycine, 0.1% SDS in dH₂O) by using the Criterion cell system (Bio-Rad). Blots were developed with rabbit anti-NR2B (1:1000, Millipore, #AB1557P), rabbit anti-pSer¹³⁰³NR2B (1:1000, Millipore, #07-398), rabbit anti-NR2A (1:1000, Millipore, #AB1555P), rabbit anti-PSD95 (1:500, abcam, #ab18258), mouse anti-synaptophysin (1:500, Millipore, #MAB329), rabbit antiphosho PKC (1:1000, Cell Signaling, #9371), rabbit anti-PKC (1:1000, abcam, #ab179521) and rabbit anti-β-actin (1:5000; Sigma-Aldrich, #A2066). Secondary antibodies conjugated with horseradish peroxidase (HRP) were purchased from Sigma (1:5000, sheep antimouse-HRP #A6782, goat antirabbit-#HRP A6154).

Ortiz-Sanz C., Balantzategi U., Quintela-López T., Ruiz A., Luchena C., Zuazo-Ibarra J., Capetillo-Zarate E., Matute C., Zugaza J.L, & Alberdi E. (2022). Amyloid β / PKC-dependent alterations in NMDA receptor composition are detected in early stages of Alzheimer´s disease. Cell Death & Disease, 13(3), 253.

+ Open protocol

+ Expand

Western Blot Protein Analysis

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

Tissue samples (20 μg/lane) were run in 7.5% Tris-Glycine gels and transferred onto nitrocellulose membranes (sc-3724, Santa Cruz Biotechnology). The nitrocellulose membranes were blocked in 10% (w/v) semi fat dry milk in TBS (Tris 0.01 M, NaCl 0.15 M, pH 7.4) for 1 h at room temperature and were incubated with primary antibodies overnight followed by rinses and an incubation with anti-rabbit (#7074, Cell Signalling Technology, Bioké) or anti-mouse (A6782, Sigma) immunoglobulin conjugated to horseradish peroxidase. After several rinses, the membranes were incubated with SuperSignal West Pico Substrate (Pierce) and were exposed to an X-ray film (Pierce) or to a DARQ-7 CCD cooled camera (Vilber-Lourmat) in a SOLO 4S WL system. Levels of optical density (OD) of protein signals were estimated by densitometry analysis using the NIH ImageJ program. Anti-β-actin immunoblots were used to normalize protein loading.

Ando K., Ndjim M., Turbant S., Fontaine G., Pregoni G., Dauphinot L., Yilmaz Z., Suain V., Mansour S., Authelet M., De Dekker R., Leroy K., Delatour B., Duyckaerts C., Potier M.C, & Brion J.P. (2020). The lipid phosphatase Synaptojanin 1 undergoes a significant alteration in expression and solubility and is associated with brain lesions in Alzheimer’s disease. Acta Neuropathologica Communications, 8, 79.

+ Open protocol

+ Expand

Immunoblotting of Nuclear and Cytoplasmic Extracts

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

Nuclear and cytoplasmic protein extracts from mIMCD3 Flp-in myc-HNF1β and mIMCD3 Flp-in wild-type (WT) cells were denatured in Laemmli sample buffer (2% (v/v) SDS, 0.01% (w/v) bromophenol blue, 6% (v/v) glycerol, and 60 mM Tris-HCl/pH 6.8) containing 100 mM of DTT for 30 minutes at 37°C. Subsequently, samples were subjected to SDS-PAGE and used for immunoblotting. Rabbitanti-c-Myc (9E10, 1:500, Santa Cruz Biotechnology, CA, USA), Rabbit anti-HNF1β (SC-22840, 1:500, Santa Cruz Biotechnology), mouse anti-HA (1:5,000, 6E2, Cell signaling) or mouse anti-Flag M2 (F1804, 1:5,000, Sigma-Aldrich) primary antibodies and peroxidase-conjugated sheep anti-mouse (A6782, 1:10,000; Sigma-Aldrich) and peroxidase-conjugated goat anti-rabbit (A4914, 1:10,000; Sigma-Aldrich) secondary antibodies were used to visualize proteins.

Tholen L.E., Bos C., Jansen P.W.T.C., Venselaar H., Vermeulen M., Hoenderop J.G.J, & de Baaij J.H.F. (2021). Bifunctional protein PCBD2 operates as a co-factor for hepatocyte nuclear factor 1β and modulates gene transcription. FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 35(4).

+ 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!