To each lot, 10–15 ml of stain was added to the filter paper without suction, i.e. either bromocresol green or 1:1 mixture of bromocresol green+orange G or acid fuchsin. After 2–3 min, mild suction was applied to allow the stain to pass through the
filter paper, after which the filter paper was washed with 15–20 ml of 10% sulphuric acid followed by several small washes with water.