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2015 ; 10
(7
): e0132142
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English Wikipedia
An Optimized Fluorescence-Based Bidimensional Immunoproteomic Approach for
Accurate Screening of Autoantibodies
#MMPMID26132557
Dutoit-Lefèvre V
; Dubucquoi S
; Launay D
; Sobanski V
; Dussart P
; Chafey P
; Broussard C
; Duban-Deweer S
; Vermersch P
; Prin L
; Lefranc D
PLoS One
2015[]; 10
(7
): e0132142
PMID26132557
show ga
Serological proteome analysis (SERPA) combines classical proteomic technology
with effective separation of cellular protein extracts on two-dimensional gel
electrophoresis, western blotting, and identification of the antigenic spot of
interest by mass spectrometry. A critical point is related to the antigenic
target characterization by mass spectrometry, which depends on the accuracy of
the matching of antigenic reactivities on the protein spots during the 2D
immunoproteomic procedures. The superimposition, based essentially on visual
criteria of antigenic and protein spots, remains the major limitation of SERPA.
The introduction of fluorescent dyes in proteomic strategies, commonly known as
2D-DIGE (differential in-gel electrophoresis), has boosted the qualitative
capabilities of 2D electrophoresis. Based on this 2D-DIGE strategy, we have
improved the conventional SERPA by developing a new and entirely
fluorescence-based bi-dimensional immunoproteomic (FBIP) analysis, performed with
three fluorescent dyes. To optimize the alignment of the different antigenic
maps, we introduced a landmark map composed of a combination of specific
antibodies. This methodological development allows simultaneous revelation of the
antigenic, landmark and proteomic maps on each immunoblot. A computer-assisted
process using commercially available software automatically leads to the
superimposition of the different maps, ensuring accurate localization of
antigenic spots of interest.
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