scholarly journals Use of a new fluorogenic phosphatase substrate in immunohistochemical applications.

1995 ◽  
Vol 43 (1) ◽  
pp. 77-83 ◽  
Author(s):  
K D Larison ◽  
R BreMiller ◽  
K S Wells ◽  
I Clements ◽  
R P Haugland
Keyword(s):  
Alkaline Phosphatase ◽  
Stokes Shift ◽  
High Magnification ◽  
Adult Zebrafish ◽  
Large Stokes Shift ◽  
Retinal Tissue ◽  
Bright Yellow ◽  
Immunohistochemical Techniques ◽  
Phosphatase Substrate

We used the phosphatase substrate 2-(5'-chloro-2'-phosphoryloxyphenyl)-6- chloro-4-[3H]-quinazolinone, with standard alkaline phosphatase-mediated immunohistochemical techniques, to visualize a number of antibodies that bind to adult zebrafish retinal tissue. This compound, known as the ELF (enzyme-labeled-fluorescence) phosphatase substrate, produces a precipitate that fluoresces at approximately 500-580 nm (bright yellow-green). We show that the precipitated product from the ELF phosphatase substrate has a number of characteristics that make it superior to fluorescein-labeled secondary reagents. The staining produced with the ELF substrate is much more photostable than that produced by fluorescein-labeled secondary reagents, thus allowing time to examine, focus, and photograph the ELF-labeled tissue under high magnification. Moreover, the ELF precipitate exhibits a Stokes shift of greater than 100 nm, a characteristic that has enabled us to overcome the problem of distinguishing signal from background in this autofluorescent tissue. In addition, we show that the ELF product's large Stokes shift makes the ELF substrate ideal for multicolor applications.

scholarly journals The ELF-97 Alkaline Phosphatase Substrate Provides a Bright, Photostable, Fluorescent Signal Amplification Method for FISH

1997 ◽  
Vol 45 (3) ◽  
pp. 345-357 ◽  
Author(s):  
Violette B. Paragas ◽  
Yu-Zhong Zhang ◽  
Richard P. Haugland ◽  
Victoria L. Singer
Keyword(s):  
Alkaline Phosphatase ◽  
Cultured Cells ◽  
Mdck Cells ◽  
Blot Hybridization ◽  
Stokes Shift ◽  
Southern Blot Hybridization ◽  
Zebrafish Embryos ◽  
Tissue Sections ◽  
Whole Mount ◽  
Phosphatase Substrate

We used the ELF-97 (Enzyme-Labeled Fluorescence) phosphatase substrate, 2-(5'-chloro-2-phosphoryloxyphenyl)-6-chloro-4(3H)-quinazolinone, with alkaline phosphatase conjugates of streptavidin and appropriate antibodies to amplify signals from biotinylated and haptenylated hybridization probes. The dephosphorylated product, ELF-97 alcohol, is a bright yellow-green fluorescent precipitate optimally excited at ∼360 nm, with emission centered at ∼530 nm. This large Stokes shift allows ELF-97 signals to be easily distinguished from sample autofluorescence and signals arising from counterstains or other fluorophores. The ELF-97 precipitate was extremely photostable compared to fluorescein, allowing multiple photographic exposures of samples without significant signal intensity loss. For RNA in situ hybridization, labeling was specific and localized well to targets in cultured cells, tissue sections, and whole-mount zebrafish embryos. ELF-97 signals developed in seconds to minutes and were easily distinguished from pigmented tissues or cells, unlike those obtained using colorimetric substrates. We used the substrate with singly biotinylated short oligonucleotides to detect actin mRNA in MDCK cells and actin and β-galactosidase mRNA in LacZ+ mouse fibroblasts. We also used a biotinylated cDNA, complementary to the mRNA encoded by the constant region of the T-cell receptor β-chain, to specifically identify T-cells in mouse lymph node tissue sections. With digoxigenin-labeled probes, we detected several developmentally expressed mRNAs in whole-mount zebrafish embryos. Hybridization to centromere repeat regions in human metaphase and interphase chromosomes was also detected; ELF-97 signals were manyfold brighter than signals obtained with fluorescein conjugates. Finally, Southern blot hybridization using singly labeled oligonucleotide probes yielded a sensitivity similar to that obtained with radioactivity. (J Histochem Cytochem 45:345–357, 1997)

A novel near-infrared fluorescent probe based on isophorone for the bioassay of endogenous cysteine

2021 ◽  
Author(s):  
Hong-Bo Liu ◽  
Hai Xu ◽  
Xin Guo ◽  
Jian Xiao ◽  
Zheng-Hong Cai ◽  
...  
Keyword(s):  
Aqueous Solution ◽  
Fluorescent Probe ◽  
Near Infrared ◽  
Stokes Shift ◽  
Rapid Identification ◽  
Large Stokes Shift

A near-infrared (NIR) fluorescent probe with a large Stokes shift (143 nm) for the rapid identification of Cys over Hcy and GSH in aqueous solution was developed.

Spectra, stability and labeling of 1-(5-carboxypentyl)-4-(2-(N-ethyl-carbazole-3-yl) vinyl) pyridinium bromide with a large Stokes shift

Luminescence ◽  
2015 ◽  
Vol 31 (2) ◽  
pp. 380-387 ◽  
Author(s):  
Yingchun Gu ◽  
Dayong Lin ◽  
Ran Li ◽  
Yalin Tang ◽  
Xuening Fei ◽  
...  
Keyword(s):  
Stokes Shift ◽  
Pyridinium Bromide ◽  
Large Stokes Shift
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