Exposure of the nuclear counterstain DAPI to just a few seconds of UV excitation light during fluorescence imaging can photoconvert the dye to stable green- and red-emitting forms, resulting in imaging artifacts in multi-color staining experiments. This article reviews the published literature on photoconversion and compares the effect of different mounting media on photoconversion of DAPI in stained tissue sections. Once photoconversion is recognized as a potential source of non-specific fluorescence, careful imaging practices or reagent selection can be used to avoid it.

LED illumination systems for fluorescence microscopy offer a wealth of benefits in comparison to traditional mercury and metal halide lamps, including ease of use, improved stability, and enhanced control. To fully realize these benefits, it is important to ensure that optical filters are configured correctly, which often can be confusing. However, without the correct filter configuration, experimental conditions can be suboptimal, and results may therefore be inaccurate. This article looks at optical filter setup in more depth, explaining the purpose and benefits of optimal LED filtering.

The objective lens is regarded as the critical component for an optimal image in light microscopy. Not every damage to the lens surface may be easily visible during maintenance when the objective is removed and observed. The internal lens organization produces a magnified image of the outer lens surface. Shining the light of a laser pointer into the lens, and reproducing the image of the light exiting onto a flat surface, produces different images for intact and damaged objective lenses.