Bio-Rad Mini-PROTEAN® TGX™ Precast Gels for 2-D Electrophoresis - Manual (Page 30)

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10.2 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.
10.3 TBE Gel Staining
Use Table 10.2 as a guide to selecting an appropriate staining method.
Table 10 .2 . TBE gel detection methods .
Method Sensitivity
(Lower Limit) Advantages Disadvantages
Ethidium bromide 50 ng Classic fluorescent DNA stain Carcinogenic
Silver stain 1–2 ng More sensitive than ethidium bromide Requires multiple steps
SYBR ® Green 0.02–2 ng High sensitivity Multiple steps, –20°C storage
SYBR ® Safe 0.5 ng Non-hazardous Multiple steps
10.4 TBE-Urea Gel Staining
Use Table 10.3 as a guide to selecting an appropriate staining method.
Table 10 .3 . TBE-urea gel detection methods .
Method Sensitivity
(Lower Limit) Advantages Disadvantages
Ethidium bromide 10 ng Classic fluorescent DNA stain Carcinogenic
SYBR ® Green 0.02–2 ng High sensitivity Requires multiple steps, −20°C
storage
Silver stain 1–2 ng More sensitive than ethidium bromide Requires multiple steps
Mini-PROTEAN Precast Gels