Cellulase screening by iodine staining: An artefact

Hydrolysis zones visualized by iodine (KI/I₂) staining of cellulose-agar media after growth of Trichoderma reesei QM6a, RUT C30, QM9136 (cellulase negative) or Thielavia terrestris cultures, or incubation of crude endoglucanases and amylases, were due primarily to degradation of a small amount of starch contaminant in commercial agar and not to cellulolysis as recently suggested. No zones were evident when amylase-digested agar or Gelrite was used as the gelling agent or when purified cellobiohydrolase and endoglucanase were used. Cellulase screening free from artefacts is best obtained by growing cultures on acid-swollen or crystalline cellulose with Gelrite as optimal gelling agent, followed by incubation at elevated temperature to enhance visualization of hydrolysis zones while restricting fungal growth, but without additional staining.