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FRET
Hardware and software support to optimize the environment for FRET.
Ratio changes when cameleon is manifested on the HeLa cell and stimulated by histamine then inhibited by cyproheptadine.
Cameleon genes provided by Dr. Miyawaki Atsushi in Brain Research Institute.
Equipment: FV300 and HeCd laser
Time period : 4 seconds.
The 440nm diode laser can be added for CFP/YFP FRET imaging.
A 440nm diode laser is optionally available for CFP/YFP imaging. The 440nm laser line ideally excites CFP, with minimal disturbance to YFP, and is therefore suitable for CFP/YFP FRET experiments. The high performance LSM objectives, PLAPO40XWLSM and PLAPO60XWLSM, are precisely corrected in this wavelength range, and ensure the highest measuring reliability.
*For simultaneous observation of CFP and YFP, 440nm
and 515nm laser lines are required.
Ratio imaging to analyze 2-wavelength images
Using time course software, the ratio image can be continuously displayed in pseudo- color. At the same time, the intensity of each channel can be monitored graphically. The analysis process is presented as an intuitive flow chart. (optional time course software: TIEMPO)
CFP Fluorescence wavelength 485nm | YFP Fluorescence wavelength 530nm |
Measurement
CFP/YFP FRET
Calcium ion concentration in a live HeLa cell using a cameleon (split type) indicator. Energy transfer between CFP and YFP is proportional to bound calcium. The time series shows the increase of calcium ion density caused by stimulation of histamine and the effect of blocking by proheputajin.
Input/output of external trigger signal
The optional time course software gives control over the input/output trigger signal by GUI. It is suitable for combined experiments such as those involving patch clamping.
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