Format

Send to

Choose Destination
See comment in PubMed Commons below
PLoS One. 2009;4(2):e4636. doi: 10.1371/journal.pone.0004636. Epub 2009 Feb 27.

A temporal threshold for formaldehyde crosslinking and fixation.

Author information

  • 1Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, United Kingdom.

Abstract

BACKGROUND:

Formaldehyde crosslinking is in widespread use as a biological fixative for microscopy and molecular biology. An assumption behind its use is that most biologically meaningful interactions are preserved by crosslinking, but the minimum length of time required for an interaction to become fixed has not been determined.

METHODOLOGY:

Using a unique series of mutations in the DNA binding protein MeCP2, we show that in vivo interactions lasting less than 5 seconds are invisible in the microscope after formaldehyde fixation, though they are obvious in live cells. The stark contrast between live cell and fixed cell images illustrates hitherto unsuspected limitations to the fixation process. We show that chromatin immunoprecipitation, a technique in widespread use that depends on formaldehyde crosslinking, also fails to capture these transient interactions.

CONCLUSIONS/SIGNIFICANCE:

Our findings for the first time establish a minimum temporal limitation to crosslink chemistry that has implications for many fields of research.

PMID:
19247482
PMCID:
PMC2645674
DOI:
10.1371/journal.pone.0004636
[PubMed - indexed for MEDLINE]
Free PMC Article
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Full text links

    Icon for Public Library of Science Icon for PubMed Central
    Loading ...
    Support Center