In:
American Journal of Physiology-Cell Physiology, American Physiological Society, Vol. 294, No. 4 ( 2008-04), p. C1096-C1102
Abstract:
Cell migration is an important physiological process among others controlled by ion channel activity. Calcium-activated potassium channels (K Ca 3.1) are required for optimal cell migration. Previously, we identified single human (h)K Ca 3.1 channel proteins in the plasma membrane by means of quantum dot (QD) labeling. In the present study, we tracked single-channel proteins during migration to classify their dynamics in the plasma membrane of MDCK-F cells. Single hK Ca 3.1 channels were visualized with QD- or Alexa488-conjugated antibodies and tracked at the basal cell membrane using time-lapse total internal reflection fluorescence (TIRF) microscopy. Analysis of the trajectories allowed the classification of channel dynamics. Channel tracks were compared with those of free QD-conjugated antibodies. The size of the label has a pronounced effect on hK Ca 3.1 channel diffusion. QD-labeled channels have a (sub)diffusion coefficient D QDbound = 0.067 μm 2 /s α , whereas that of Alexa488-labeled channels is D Alexa = 0.139 μm 2 /s. Free QD-conjugated antibodies move much faster: D QDfree = 2.163 μm 2 /s α . Plotting the mean squared distances (msd) covered by hK Ca 3.1 channels as a function of time points to the mode of diffusion. Alexa488-labeled channels diffuse normally, whereas the QD-label renders hK Ca 3.1 channel diffusion anomalous. Free QD-labeled antibodies also diffuse anomalously. Hence, QDs slow down diffusion of hK Ca 3.1 channels and change the mode of diffusion. These results, referring to the role of label size and properties of the extracellular environment, suggest that the pericellular glycocalyx has an important impact on labels used for single molecule tracking. Thus tracking fluorescent particles within the glycocalyx opens up a possibility to characterize the pericellular nanoenvironment.
Type of Medium:
Online Resource
ISSN:
0363-6143
,
1522-1563
DOI:
10.1152/ajpcell.00252.2007
Language:
English
Publisher:
American Physiological Society
Publication Date:
2008
detail.hit.zdb_id:
1477334-X
SSG:
12
Permalink
