SDS-PAGE Analysis of Cell Signaling

BMSC or OCL precursor cell lysates were loaded onto 7.5% sodium dodecyl sulfate polyacrylamide gels (SDS-PAGE) (for V-CAM1 expression) or 10% SDS-PAGE (for cell signaling analyses) and electrophoresed under reducing conditions using precast gels (Bio-Rad Laboratories, Hercules, CA), as described (16 (link)). Antibodies used for immunoblotting included: anti-phospho PKCζ (Thr410/403), anti-phospho IκBα (Ser32), anti-phospho p38 MAPK (Thr180/Tyr182) and anti-p38 MAPK, anti-PKCξ, anti-IκBα, cleaved and uncleaved caspases 3, 7, and 9 (all from Cell Signaling, Boston, MA, catalog numbers 9378, 2859, 9211, 9212, 9368, 9242, respectively), anti-c-Fos, anti-NFATc1 and anti-V-CAM1 (Santa Cruz Biotechnology, Dallas, TX, catalog numbers sc-52, sc-7294, sc-1504, respectively), and anti-TNF-α (R&D Systems, Minneapolis, MN, catalog number AF-410-NA). β-actin was used for a control (Abcam, Cambridge, MA, catalog ab49900).

Partial Protocol Preview
This section provides a glimpse into the protocol.
The remaining content is hidden due to licensing restrictions, but the full text is available at the following link: Access Free Full Text.

Teramachi J., Silbermann R., Yang P., Zhao W., Mohammad K.S., Guo J., Anderson J.L., Zhou D., Feng R., Myint K.Z., Maertz N., Beumer J.H., Eiseman J.L., Windle J.J., Xie X.Q., Roodman G.D, & Kurihara N. (2015). Blocking the ZZ Domain of Sequestosome1/p62 Suppresses Myeloma Growth and Osteoclast Formation In Vitro and Induces Dramatic Bone Formation in Myeloma-Bearing Bones In Vivo. Leukemia, 30(2), 390-398.

Publication 2015

precast gels anti-phospho PKCζ anti-phospho IκBα anti-phospho p38 MAPK anti-p38 MAPK anti-c-Fos anti-NFATc1 anti-V-CAM1 anti-TNF-α β-actin

Actin Antibodies Caspases 3 Gels Iκbα P38 mapk Polyacrylamide gels Precursor cell Sds page Sodium dodecyl sulfate Tnf α

Corresponding Organization : Richard L. Roudebush VA Medical Center

Other organizations : University of Pittsburgh, Virginia Commonwealth University

Top 5 similar protocols

Variable analysis

independent variables

BMSC or OCL precursor cell lysates

dependent variables

V-CAM1 expression
Cell signaling analyses

control variables

Reducing conditions using precast gels (Bio-Rad Laboratories, Hercules, CA)
β-actin as a control (Abcam, Cambridge, MA, catalog ab49900)

controls

Positive control: Not specified
Negative control: Not specified

Annotations

Based on most similar protocols

Etiam vel ipsum. Morbi facilisis vestibulum nisl. Praesent cursus laoreet felis. Integer adipiscing pretium orci. Nulla facilisi. Quisque posuere bibendum purus. Nulla quam mauris, cursus eget, convallis ac, molestie non, enim. Aliquam congue. Quisque sagittis nonummy sapien. Proin molestie sem vitae urna. Maecenas lorem.

As authors may omit details in methods from publication, our AI will look for missing critical information across the 5 most similar protocols.

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!