Document Detail


Measurement of the lateral diffusion of human MHC class I molecules on HeLa cells by fluorescence recovery after photobleaching using a phycoerythrin probe.
MedLine Citation:
PMID:  11916842     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The mobility of cell surface MHC class I molecules on HeLa cells was measured by fluorescence recovery after photobleaching (FRAP). The probe used for these studies was the phycobiliprotein R-phycoerythrin coupled to Fab fragments of a monoclonal antibody specific for human monomorphic MHC class I molecules. It was found that the recovery curves could be equally well fitted by either a random diffusion model with an immobile component or by an anomalous diffusion model. In the latter case, the anomalous diffusion exponent was consistent with that previously determined by single-particle tracking (SPT) experiments using the same probe (P. R. Smith, I. E. G. Morrison, K. M. Wilson, N. Fernandez, and R. J. Cherry. 1999. Biophys. J. 76:3331-3344). The FRAP experiments, however, yielded a considerably higher value of D(0), the diffusion coefficient for a time interval of 1 s. To determine whether the results were probe dependent, FRAP measurements were also performed with the same monoclonal antibody labeled with Oregon Green. These experiments gave similar results to those obtained with the phycoerythrin probe. FRAP experiments with the lipid probe 5-N-(octadecanoyl) aminofluoroscein (ODAF) bound to HeLa cells gave typical results for lipid diffusion. Overall, our observations and analysis are consistent with anomalous diffusion of MHC class I diffusion on HeLa cells, but quantitative differences between FRAP and SPT data remain to be explained.
Authors:
George Georgiou; Sukhvinder S Bahra; Alan R Mackie; Caroline A Wolfe; Paul O'Shea; Shab Ladha; Nelson Fernandez; Richard J Cherry
Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Biophysical journal     Volume:  82     ISSN:  0006-3495     ISO Abbreviation:  Biophys. J.     Publication Date:  2002 Apr 
Date Detail:
Created Date:  2002-03-27     Completed Date:  2002-08-06     Revised Date:  2009-11-18    
Medline Journal Info:
Nlm Unique ID:  0370626     Medline TA:  Biophys J     Country:  United States    
Other Details:
Languages:  eng     Pagination:  1828-34     Citation Subset:  IM    
Affiliation:
Department of Biological Sciences, University of Essex, Colchester CO4 3SQ, United Kingdom.
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MeSH Terms
Descriptor/Qualifier:
Antibodies, Monoclonal
Chromatography, High Pressure Liquid
Diffusion
Fluorescein
Fluoresceins / chemistry
Fluorescent Dyes / pharmacology
Genes, MHC Class I*
Hela Cells
Humans
Immunoglobulin Fab Fragments
Immunoglobulin G / metabolism
Light
Lipids / chemistry
Phycoerythrin / chemistry
Spectrometry, Fluorescence / methods*
Time Factors
Chemical
Reg. No./Substance:
0/5-(N-octadecanoyl)aminofluorescein; 0/Antibodies, Monoclonal; 0/Fluoresceins; 0/Fluorescent Dyes; 0/Immunoglobulin Fab Fragments; 0/Immunoglobulin G; 0/Lipids; 0/oregon green 514; 11016-17-4/Phycoerythrin; 2321-07-5/Fluorescein
Comments/Corrections

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