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Dyes for Your Near-Infrared Imaging Needs
IRDye® 680LT Infrared Dye
IMPORTANT: IRDye 680LT dye products should not be used for small animal in vivo imaging or In-Cell Western™ Assays.
IRDye 680LT dye is highly soluble in water and is significantly brighter and more photostable than many other 700 nm near-infrared dyes. The spectral characteristics of IRDye 680LT are well suited for use on LI-COR imaging instruments with absorbance and emission maxima in aqueous solution and methanol of 676 and 693 nm, respectively.
Spectrally matched to LI-COR® imaging systems
Optimized for protein detection applications, including Quantitative Westerns
Low background at near-infrared wavelengths enables higher signal-to-noise ratios
IRDye® 680LT Applications
IMPORTANT: IRDye 680LT dye products should not be used for small animal in vivo imaging or In-Cell Western™ Assays.
Multiplex Western Blot Detection
[ABOVE] Lysates of EGF-treated A431 cells were separated and transferred to nitrocellulose. The blot was probed with anti-ERK and anti-phospho-ERK primary antibodies, and then detected with IRDye 680LT and IRDye 800CW secondary antibodies. Blot was imaged with Odyssey® Fc System for 2 min. This phospho-ERK antibody crossreacts with phospho-EGFR (upper green band).
Microscopy
[ABOVE] Immunofluorescence staining of HER2 protein localization to SK-BR-3 cell membrane. Cells were cultured on coverslips. After fixation and permeabilization, cells were stained with rabbit anti-HER2 mAb (Cell Signaling Technology), IRDye 680LT labeled goat anti-rabbit. Nuclei are stained with Sytox Green dye. Images were acquired with an Olympus microscope and deconvolved with the accompanying software.
IRDye® 680LT Products
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IRDye 680LT Dye Products |
IRDye 680LT Conjugates |
IRDye® 680LT Outperforms Alexa Fluor® 680
"Brightness" - Fluorescence intensity
IRDye 680LT matches or surpasses the brightness of Alexa Fluor® 680. Fluorescence determinations were made at a fixed antibody concentration of 10µg/mL in physiological buffer using dye-labeled goat anti-rabbit (GAR) conjugates prepared in-house. Fluorescence was measured using a PTI Fluorometer at the optimum excitation and emission wavelengths of each dye. The fluorescence intensity of each conjugate increased with increased degree of labeling. A plot of degree of labeling versus fluorescence (Shown Below) shows that the dynamic range for the 680LT GAR is much broader than the Alexa Fluor 680 which quickly levels off above D/P 3.4. The leveling off of the Alexa Fluor 680 conjugates is due to self-quenching. IRDye 680LT conjugates continue to increase in fluorescence intensity to at least D/P 6.4 which was the highest degree of labeling tested. Overall, IRDye 680LT is significantly brighter than Alexa Fluor 680.
[LEFT] Goat anti-rabbit IgG conjugates were analyzed at 10µg/mL in physiological buffer. Fluorescence was measured with a PTI fluorometer at optimum ex/em wavelengths for each dye.
Photostability
The photostability of IRDye 680LT was compared to Alexa Fluor 680. Test samples were prepared by spotting equimolar amounts of each dye onto nitrocellulose membrane. The membrane was then scanned repeatedly on the Odyssey Infrared Imaging System and the signal intensity was normalized to the control signal at time zero (Shown Below). The relative fluorescence of the IRDye 680LT exhibited greater photostability over Alexa Fluor 680.
[LEFT] Equimolar amounts of each dye (25 fmol) were spotted onto nitrocellulose membrane. The membrane was scanned repeatedly with the Odyssey® Classic Infrared Imaging System, and signal intensity was normalized to maximum intensity at time = 0.
