Position-addressable digital laser scanning point fluorescence microscopy with a Blu-ray disk pickup head.

A compact and position-addressable blue ray scanning microscope (BSM) based on a commercially available Blu-ray disk pickup head (PUH) is developed for cell imaging with high resolution and low cost. The BSM comprises two objective lenses with numerical apertures (NAs) of 0.85 and 0.6 for focusing blue and red laser beams, respectively, on the sample slide. The blue and red laser beams are co-located adjacent to each other and move synchronously. A specially designed sample slide is used with a sample area and an address-patterned area for sample holding and address recognition, respectively. The blue laser beam is focused on the sample area and is used for fluorescent excitation and image capturing, whereas the red laser beam is focused on the address-patterned area and is used for address recognition and dynamic focusing. The address-patterned area is divided into 310 sectors. The cell image of each sector of the sampling area has a corresponding address pattern. Fluorescence images of monkey-derived kidney epithelial cells and fibroblast cells in which the F-actin is stained with fluorophore phalloidin CF 405 are measured by the BSM, with results comparable to those measured by a Leica TCS CP2 confocal microscope. The cell image of an area of interest can be easily tracked based on the coded address, and a large-area sample image can be accurately reconstructed from the sector images.