Chitin Identification via Calcofluor White

Calcofluor white (CFW, Fluorescent Brightener M2R, Sigma-Aldrich, St. Louis, MO, USA) staining was applied preliminarily to confirm the presence of chitin in materials isolated by the electrolysis-assisted method. This technique was used several times in different variants [9 (link),14 (link),70 (link),74 (link)]. However, in this study, the 30 µL of a solution containing 10 g glycerin and 10 g NaOH in 90 mL of water was applied as a buffer. Subsequently, after 5 min, the CFW was added to the samples, and the investigated materials were incubated in staining mixture for 6 h without sunlight at 25 °C. After this treatment, samples were extracted from the staining solution and extensively rinsed with distilled water to eliminate the free unbounded molecules of CFW stain, then dried at 25 °C and investigated using fluorescent microscopy.