Protein Expression Analysis by SDS-PAGE and Western Blotting

Sodium dodecyl sulphate polyacrylamide gel electrophoresis and western blotting were performed as described previously.^{57 (link), 58 (link)} The following antibodies were used: p16 (Santa Cruz Biotechnology, Dallas, TX, USA, #sc-56330), MDR1/P-gp (Santa Cruz Biotechnology, #sc-1517), calnexin (Santa Cruz Biotechnology, #sc-80645), α-tubulin (Santa Cruz Biotechnology, #sc-32293), annexin V (Abcam, #ab14196), CD63 and HSP70 (System Biosciences, Palo Alto, CA, USA, #EXOAB-CD63A-1), p21 (BD Biosciences, #556430), GAPDH (Cell Signalling, #5174P) and Na/K ATPase (Novus Biologicals, Littleton, CO, USA #NB300-146). The differential CD63 bands potentially represents either differential glycosylation^{59 (link)} or the isolation method used to harvest EVs.^{60 (link)} Densitometry was completed to determine the expression of proteins. Densitometry was completed using Image J software with all results normalised to the loading control.

Free full text: Click here

Kavanagh E.L., Lindsay S., Halasz M., Gubbins L.C., Weiner-Gorzel K., Guang M.H., McGoldrick A., Collins E., Henry M., Blanco-Fernández A., Gorman P.O., Fitzpatrick P., Higgins M.J., Dowling P, & McCann A. (2017). Protein and chemotherapy profiling of extracellular vesicles harvested from therapeutic induced senescent triple negative breast cancer cells. Oncogenesis, 6(10), e388-.

Publication 2017

MDR1/P-gp calnexin α-tubulin annexin V HSP70 GAPDH Na/K ATPase

Annexin v Antibodies Biologicals Calnexin Densitometry Gapdh Hsp70 Isolation Na k atpase Novus Proteins Sodium dodecyl sulphate polyacrylamide gel electrophoresis α tubulin

Corresponding Organization :

Other organizations : University College Dublin, Dublin City University, Mater Misericordiae University Hospital, National University of Ireland, Maynooth

Top 5 similar protocols

Variable analysis

independent variables

Sodium dodecyl sulphate polyacrylamide gel electrophoresis and western blotting methods

dependent variables

Expression of proteins: p16, MDR1/P-gp, calnexin, α-tubulin, annexin V, CD63, HSP70, p21, GAPDH, and Na/K ATPase

control variables

Loading control used for densitometry analysis

controls

Positive controls: Not explicitly mentioned
Negative controls: Not explicitly mentioned

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!