Nile Red staining for detecting microplastics in biota: Preliminary evidence

Nile Red is a lipophilic, metachromatic and solvatochromic dye used as an alternative or complementary method to aid identification of microplastics in routine analysis of biological samples. It was rarely used in biota since organic residues after the digestion step can be co-stained with possible overestimation of microplastics. The limits of using Nile Red in biota were investigated in marine mussels experimentally contaminated with low-density polyethylene (LDPE) microplastics. Stained particles were detected through magnified images obtained by stitching together thirty photographs of the filter surface of each sample. LDPE particles appeared yellowish and fluorescent and could be confused with certain organic residues. The smaller the fragments, the greater the difficulty in recognizing them. In particular, it was difficult to recognize LDPE particles based on their fluorescence if <180 μm in size. Regardless of the size, fluorescence of the items aids the operator in LDPE particles identification also in biota.