Ethidium Bromide Nuclear Staining and Fluorescence Microscopy: An Alternative Method for Triploidy Detection in Fish

Abstract.– Erythrocyte nuclei from normal diploid (2N) and cold-shocked triploid (3N) Asian catfish Clarias macrocephalus were comparatively stained with fluorescent stain, ethidium bromide and with a bright-field stain, Giemsa. Stains were applied to dried blood smears on glass slides, with blood taken from fingerling fish without sacrificing the fish. Both stains produced comparable quality visual comparisons, with significant differences ( P < 0.001) between nuclear dimensions of 2N and 3N cells. Triploid nuclei were larger and more elongate than diploid nuclei. Karyotypic comparisons revealed that diploid fish had 54 chromosomes, while triploid fish had 81. Ethidium bromide nuclear staining required only 3 min after slides were air-dried and fixed, while Giemsa staining required more than 50 min.