Improved direct-count method by fluorescence microscope for low-biomass ice core

Abstract

Ice cores recovered from ice sheets contain microorganisms that were transported to the surface of the ice sheet along with airborne dust. The abundance of microorganisms in ice cores, as measured by direct cell count under a fluorescence microscope, is generally lower than that in other aquatic environments. It is possible that the abundance of microorganisms in ice cores is underestimated due to the degradation of fluorescent dye or because non-specific fluorescence from mineral particles inhibits the fluorescence signal from microorganisms. To investigate this possibility, we tested 4 types of antifade solution and 19 types of fluorescence dye, and selected the best combination in terms of prolonging the fluorescent signal from microorganisms and distinguishing biological signals from noise produced by non-specific particles. The results show that the optimal combination of solution and dye for cell counts of scarce microorganisms in ice cores is the nuclear acid dye ＂YOYO-1＂ (Molecular Probes) and the antifade solution ＂EverBrite Mounting Medium＂ (Biotium).