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

Abstract

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.

Hacking CD/DVD/Blu-ray for Biosensing

“…B12, cover glass; B13, collimator astigmatic plate; B14, photodetector. Reprinted with permission from ref ( 74 ). Copyright 2014 The Optical Society.…”

“…The BSM uses the Blu-ray optical path to excite Simultaneously, the DVD optical path is focused on an address-patterned area to monitor the xy scanning area and zaxis focusing. 73 This BSM setup has been successfully used to measure monkey-derived kidney epithelial cells and fibroblast cells stained with fluorophore phalloidin CF405 (Biotum, Inc., Fremont, CA, USA), 74 as shown in Figure 16. Moreover, the compact size of the BSM allows embedding it into a cell culture chamber.…”

Cell depth imaging by point laser scanning fluorescence microscopy with an optical disk pickup head

“…3) In our previous study, we developed a compact, cost-effective, and positionaddressable digital laser scanning microscopy (DLSM) instrument using a commercially available Blu-ray disk pickup head (PUH). 4) The PUH comprises two objective lenses with numerical apertures (NAs) of 0.85 and 0.6, which are used to focus blue (405 nm) and red (650 nm) laser beams onto the sample slide, respectively. A specially designed sample slide with sample-holding areas and address-patterned areas for image capture and address recognition, respectively, is used.…”

“…The resolution of the measured fluorescence image is 0.38 µm, which is determined by the feature size of the address pattern. 4) In addition to its compact size and low cost, several advantages are expected for the position-addressable DLSM instrument with the commercially available Blu-ray disk PUH. A large-format cell image can be obtained by stitching the small-format images adjacent to each other.…”

“…A large-format cell image can be obtained by stitching the small-format images adjacent to each other. 4) The signalto-noise ratio of the image can be increased by scanning the same address image multiple times to multiply the signal intensity, while the variation of the standard deviation of background noise remains nominal. 5,6) The three-dimensional (3D) image can be obtained by stacking the images measured at different z-sections at the same x-y position.…”