Detection of diverse and high molecular weight nesprin-1 and nesprin-2 isoforms using western blotting

James Carthew, Iakowos Karakesisoglou

Research output: Chapter in Book/Report/Conference proceedingChapter (Book)Otherpeer-review

2 Citations (Scopus)

Abstract

Heavily utilized in cell and molecular biology, western blotting is considered a crucial technique for the detection and quantification of proteins within complex mixtures. In particular, the detection of members of the nesprin (nuclear envelope spectrin repeat protein) family has proven difficult to analyze due to their substantial isoform diversity, molecular weight variation, and the sheer size of both nesprin-1 and nesprin-2 giant protein variants (>800 kDa). Nesprin isoforms contain distinct domain signatures, perform differential cytoskeletal associations, occupy different subcellular compartments, and vary in their tissue expression profiles. This structural and functional variance highlights the need to distinguish between the full range of proteins within the nesprin protein family, allowing for greater understanding of their specific roles in cell biology and disease. Herein, we describe a western blotting protocol modified for the detection of low to high molecular weight (50–1000 kDa) nesprin proteins.

Original languageEnglish
Title of host publicationThe Nuclear Envelope
Subtitle of host publicationMethods and Protocols
EditorsSue Shackleton, Philippe Collas, Eric C. Schirmer
Place of PublicationNew York NY USA
PublisherHumana Press
Chapter14
Pages221-232
Number of pages12
Edition1st
ISBN (Electronic)9781493935307
ISBN (Print)9781493935284
DOIs
Publication statusPublished - 2016
Externally publishedYes

Publication series

NameMethods in Molecular Biology
PublisherHumana Press
Volume1411
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

Keywords

  • LINC complex
  • Nesprins
  • Nuclear envelope
  • Polyvinylidene difloride
  • SDS-PAGE
  • SYNE
  • Western blotting

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