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The excitation laser wavelength was 405 nm, and the emission channel was set to 420 535 nm.
However, we do not observe a difference in the red and green channel in terms of behavior, except for the added poleward arc of the green emission channel.
The area under the Ca2+ response curves was integrated for each emission channel.
Using the collapsed confocal stack images, software was developed to define the cells (blue emission channel image), count the number of beads per cell (green emission channel image) and determine if the beads are outside the cell (red emission channel image) using custom software developed in MATLAB® (The Mathworks, Inc., Natick, MA, USA).
Almost the entire collagen signal is in the blue emission channel, since the major signal that we detect from collagen is due to SHG.
In order to avoid ECFP bleeding into the EYFP emission channel, a special filter set was used that clearly separates the two fluorochromes.
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The instrument has two emission channels, 180° apart, each comprising a Glan-Taylor polarizer.
Suitable energy range of projectile for this production was selected by spotting of maximum cross section and minimum impurity due to other emission channels.
In addition, two different emission channels emerged in a diaminomaleonitrile derivative when the sensor was exposed to different concentration of copper cation in acetonitrile solution.
Flow cytometry was carried out using a BD FACSCalibur instrument (Becton Dickinson, USA) fitted with a 15 mW argon ion laser for excitation (488 nm), while monitoring with three different emission channels (530/30, 585/42, and 670 nm Lp).
The lymphatic tissue signal is similar in all three emission channels, suggesting that its origin is a single fluorophore with broad excitation and emission.
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