Sulforhodamine G

Guide (Following is our recommended protocol. This protocol only provides a guideline, and should be modified according to your specific needs).
Labeling of fluorescent internal protein:
A. Prepared of protein samples:
1. Protein samples to be analyzed are spiked with 0.1% of the total protein load of ALIS647 (ALIS internal standard) prior.
2. Protein sample is separated with 2-DE in the dark.
B. Purify Sulforhodamine G:
1. Sulforhodamine G (60% purity, 10 mg) dissolved in 100 mL of 1% v/v acetic acid to purify by RP chromatography.
2. Collect the pool with an absorbance maximum at 528 nm, lyophilized to dryness and stored as a dry powder at 47°C.
C. Stainning:
1. staining was performed in polypropylene staining dishes wrapped in aluminum foil to prevent photobleaching of the stains.
2. Sulforhodamine G staining is performed overnight in 35% methanol with a four-fold molar excess of dye: protein based on an average protein molecular weight of 50 kDa.
3. Following 4×15 min washes in 35% methanol and 2×15 min equilibrations in water.
4. Total protein and ALIS are visualized with a laser scanner using different channels.
5. Protein spots visualized using total protein stain (λex=532 nm) and ALIS (λex=633 nm) are quantified separately using 2-DE software, and statistical analysis.

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.