Tests were performed on Olympus BX51 microscope with 10X UPlanFL 0.3NA, 20X UPlanFL 0.5NA, and 40X UPlanFL 0.75NA objectives and Chroma 39002 - AT - EGFP/FITC/Cy2/AlexaFluor 488 filter cube.

The light source was set to full intensity. Focusing of the light source was done by focusing the LED chip on the specimen without objective in the optical path.  The specimen (fluorescent plastic) was pre-focused with a 10X objective.

Camera - Basler color camera scA 1400-17gc, chip size 1390*1038 pixels, pixel size 6.45 μm * 6.45 μm, in 8-bit mode,  Gain 600.  Images in the table are full camera frames scaled down by a factor of 2.  Exposure time was chosen to slightly saturate the image.  No digital enhancement was done.

Specimens: Molecular Probes FluoCells Prepared Slide #1- bovine pulmonary artery endothelial (BPAE) cells with MitoTracker Red CMXRos, Alexa Fluor 488 Phalloidin, and DAPI and Spherotech Fluorescent Particle Slide, Ultra Rainbow, 5 Intensities, 5.05 μm, Cat. No. FPS-5057-UR5.