Preparation of Calcoflour stock solution

Ø Weigh 35 mg of Calcoflour White (C₄₀H₄₄N₁₂O₁₀S_{2 –} = Fluorescence Brightener 28, Sigma F3543) into a 15 ml tube.

Ø Add 7 mL of distilled water and a 2-3 drops of 10N NaOH (to increase pH to 10-11). Calcofluor does not dissolve well in neutral solutions.

Ø Dissolve the calcoflour

Ø Adjust the volume to 10 mL by adding distilled water.

Ø Store this solution in a lightproof tube (preferably covered with Al-foil) at -20°C.

Ø This is a 100X solution.

P.S. For staining, one should dilute it to 1X in Tris/HCl buffer solution (0.1 M and pH 9.0). 10% glycerol and phenylene diamine as anti-fade can be added. Live cells or formaldehyde fixed cells can be stainined. Stain for 15 minutes, followed by one wash in H2O or PBS.