How to Prepare Coomassie Blue Staining Solution

1. 1Step 1: In a fume hood, add ~200 mL distilled water to a 1L glass beaker with magnetic stir bar
2. 2Step 2: Add 1.0g Coomassie Brilliant Blue R-250 dye powder while stirring
3. 3Step 3: Stir for 30-45 minutes until dye is completely dissolved (solution should be deep blue with no visible particles)
4. 4Step 4: Slowly add 500 mL methanol while continuing to stir
5. 5Step 5: Very slowly add 100 mL glacial acetic acid dropwise while stirring vigorously to prevent localized heating
6. 6Step 6: Add distilled water to bring final volume to exactly 1L
7. 7Step 7: Continue stirring for 10 minutes to ensure complete mixing
8. 8Step 8: Filter through Whatman filter paper or 0.22 μm filter to remove any undissolved particles

Filter through 0.22 μm filter if sterile solution is required. Cannot autoclave due to methanol evaporation which will alter composition.

MODERATE RISK: Requires fume hood for methanol and acetic acid vapors. Wear nitrile gloves, safety glasses, and lab coat. Glacial acetic acid is corrosive - add slowly to prevent violent reaction and vapor generation. Methanol is toxic and flammable - no open flames.

Store at room temperature in dark glass bottle or wrap in aluminum foil to prevent light degradation of dye. Shelf life 6 months - replace if methanol evaporation is suspected (check volume) or if precipitation occurs.