Biology Reference
In-Depth Information
Fig. 1. Workfl ow for a minimal CyDye labeling experiment. After CyDye labeling, the three samples are separated on the
same 2D gel and then imaged for the resulting fl uorescence associated with each CyDye. Permission to reproduce from
Westermeier and Scheibe ( 8 ) .
Table 1
Experimental design for a minimal labeling experiment,
incorporating a dye swap and including a pooled internal
standard (standard). This scenario allows for looking at two
different conditions: 1 and 2
Cy2
Cy3
Cy5
Gel 1
Gel 2
Gel 3
Gel 4
Standard
Standard
Standard
Standard
Sample 1a
Sample 2c
Sample 1c
Sample 2a
Sample 2d
Sample 1b
Sample 2b
Sample 1d
Standard = s1a + s1b + s1c + s1d + s2a + s2b + s2c + s2d
The letters a , b , c, and d denote biological replicates
compared to classical detection techniques ( 9 ) (see Fig. 2 ). The
2D DIGE system benefi ts from the pooled internal standard in
several ways; it is used to help normalize the signal between and
within each gel by comparing the ratio of each labeled protein spot
to the internal standard and then to the same protein spot in the
other gels. In addition, the pooled internal standard is used as a
standard map to match protein spots across multiple gels since all
of the spots in the internal standard should be present across all of
the gels.
Search WWH ::




Custom Search