Simultaneous imaging of multiple fluorescent probes in bio-cells.
In order to obtain the full spectrum from 400 to 800 nm of each pixel of a microscopic image, a unique spectro-imaging system was developed using an image slicer. The image slicer is composed of 100 photo fibers which are arranged in a matrix of 10 x 10 at the entrance and 100 x 1 at the exit. A line of this 100 signals is passed through a glism and projected onto a CCD. This system was applied to the fluorescent imaging of bio-cells. One of the demonstrative examples was simultaneous measurements of the Ca2+ concentration and the pH using of respective fluorescent probes. An electric signal was applied to BY-2 protoplasts and the fluorescent spectrum from 500 nm to 800 nm was measured every 5 s. The spectrum of the BY-2 protoplasts changed in response to the electric signal and the Ca2+ concentration, and the pH changes could be monitored. The wavelength resolution was satisfactory, but the space resolution was still rough in comparison with the usual microscopic systems. Notwithstanding these conditions, we could obtain discrete data from more than several tens of sites in a single-cell or a chain of several cells.
Matsuoka
Hideaki
H
Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology, Koganei, Tokyo 184-8588, Japan.
Kosai
Yuri
Y
Saito
Mikako
M
Takeyama
Norihide
N
Suto
Hiroshi
H
D002118:Calcium / Q000378:metabolism*; D004558:Electric Stimulation; D004867:Equipment Design; D005456:Fluorescent Dyes; D006863:Hydrogen-Ion Concentration; D007091:Image Processing, Computer-Assisted; D008856:Microscopy, Fluorescence / Q000295:instrumentation; D011523:Protoplasts / Q000166:cytology / Q000378:metabolism*; D015398:Signal Transduction; D014026:Nicotiana / Q000166:cytology
D005456:Fluorescent Dyes; D002118:Calcium