Bio-Rad Criterion™ TBE-Urea Precast Gels - Manual (Page 37)

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Stain Sensitivity
(Lower
Limit)
Optimum
Protein Load
(µg/Band)
Advantages
Disadvantages
Imaging
Manual
SYPRO
Ruby protein
gel stain 1–10 ng ~0.1 Broad dynamic
range Requires laser-
or LED-based
imaging
instrument for
maximum
sensitivity Fluorescence
visualization
with UV,
LED, or laser
scanning 4006173
Flamingo ™
fluorescent gel
stain
0.25–0.5 ng ~0.02 Broad dynamic
range, mass
spectrometry
compatible 10003321
Criterion TGX Stain-Free and Criterion Stain Free Gels
Stain Free
imaging
2–28 ng ~0.5 Rapid (<5 min),
compatible with
blotting and mass
spectrometry,
simple protocol
with no additional
reagents Requires
tryptophan
residues in
proteins for
detection Fluorescence
visualization
using Criterion
Stain Free
imaging system 10014472
12.2 Peptide Gel Staining
Peptides and small proteins are prone to diffusion and loss during staining. The following protocol
includes a fixing step prior to staining to prevent sample loss and is suitable for detection of bands as
low as 10–20 ng.
Fixative solution 40% methanol, 10% acetic acid
Stain solution 0.025% (w/v) Coomassie Blue G-250, 10% acetic acid
Destain solution 10% acetic acid
Place gels in fixative solution and equilibrate for 30 min. Stain gels with stain solution for 1 hr. Stain
should be used only once; reuse may result in loss of sensitivity. Destain gels three times for 15 min or
until the desired background is achieved. Some peptides may not be completely fixed and may diffuse
out of the gels if fixing and staining times are greatly exceeded.
12.3 IEF Gel Staining
Samples on IEF gels can be detected using multiple methods. Use Table 12.2 as a guide to selecting an
appropriate staining method.
Table 12 .2 . IEF gel detection methods .
Method
Sensitivity
(Lower Limit)
Optimum
Protein Load
(µg/Band) Advantages Disadvantages
Silver Stain Plus kit 0.6–1.2 ng ~0.01 Sensitive, robust, mass
spectrometry compatible Requires TCA fixation
Silver stain 0.6–1.2 ng ~0.01 Stains complex proteins
(glyco- or lipoproteins) Not mass spectrometry
compatible
SYPRO Ruby protein
gel stain 1.0–10 ng ~0.1 Broad dynamic range, mass
spectrometry compatible Requires laser- or LED-
based imaging instrument
for maximum sensitivity
Flamingo fluorescent
gel stain 0.25–0.5 ng ~0.2 Broad dynamic range, mass
spectrometry compatible Requires laser- or LED-
based imaging instrument
for maximum sensitivity
Instruction Manual and Application Guide