Nanoscale Protein Diffusion by STED-Based Pair Correlation Analysis

We describe for the first time the combination between cross-pair correlation function analysis (pair correlation analysis or pCF) and stimulated emission depletion (STED) to obtain diffusion maps at spatial resolution below the optical diffraction limit (super-resolution). Our approach was tested in systems characterized by high and low signal to noise ratio, i.e. Capsid Like Particles (CLPs) bearing several (>100) active fluorescent proteins and monomeric fluorescent proteins transiently expressed in living Chinese Hamster Ovary cells, respectively. The latter system represents the usual condition encountered in living cell studies on fluorescent protein chimeras. Spatial resolution of STED-pCF was found to be about 110 nm, with a more than twofold improvement over conventional confocal acquisition. We successfully applied our method to highlight how the proximity to nuclear envelope affects the mobility features of proteins actively imported into the nucleus in living cells. Remarkably, STED-pCF unveiled the existence of local barriers to diffusion as well as the presence of a slow component at distances up to 500–700 nm from either sides of nuclear envelope. The mobility of this component is similar to that previously described for transport complexes. Remarkably, all these features were invisible in conventional confocal mode.


Topics & Tags

Read full article:

Bianchini P, Cardarelli F, Luca MD, Diaspro A, and Bizzarri R:
Nanoscale Protein Diffusion by STED-Based Pair Correlation Analysis

PLoS ONE 9 (6): e99619 (2014); doi: 10.1371/journal.pone.0099619

Related Pages

Interested to know more?

Talk to our experts. We are happy to answer all your questions and concerns.

Contact Us

Do you prefer personal consulting?

  • Leica Microsystems Inc.
    1700 Leider Lane
    Buffalo Grove, IL 60089 United States
    Office Phone : +1 800 248 0123
    Service Phone : 1 800 248 0223
    Fax : +1 847-236-3009

You will find a more detailed list of local contacts here.