Abstract:
:Dual-excitation ratiometric dyes permit quantitative Ca2+ measurements by minimizing the effects of several artifacts that are unrelated to changes in the concentration of free Ca2+ ([Ca2+]). These dyes are excited alternately at two different wavelengths, and the pair of intensity measurements must be collected sequentially. Therefore, it is difficult to follow very fast Ca2+ dynamics or Ca2+ changes in highly motile cell samples. Here, we present a novel but simple dual-excitation ratiometric method which overcomes this problem. By the use of our home-made illuminator, each sample is illuminated by two orthogonal linear polarized lights of different wavelengths. Fluorescence images are captured by two CCD cameras through two analyzers, whose polarization directions are at right angles. This methodology allows us to perform simultaneous measurements of any dual-excitation ratiometric dye, and we demonstrate its validity by monitoring [Ca2+] changes in rat cardiac muscle cells loaded with Fura Red.
journal_name
Biochem Biophys Res Communjournal_title
Biochemical and biophysical research communicationsauthors
Fukano T,Shimozono S,Miyawaki Adoi
10.1016/j.bbrc.2004.03.009subject
Has Abstractpub_date
2004-04-23 00:00:00pages
77-83issue
1eissn
0006-291Xissn
1090-2104pii
S0006291X04004607journal_volume
317pub_type
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