The fluorescent dye DAPI is useful for its association with and consequent amplification of an ∼460 nm emission maximum upon binding to dsDNA. Labelling with higher DAPI concentrations is a technique used to reveal Pi polymers [polyphosphate (polyP)], with a red-shift to ∼520–550 nm fluorescence emission. DAPI–polyP emissions of ∼580 nm are also generated upon 415 nm excitation. Red-shifted DAPI emission has been associated with polyP and RNA and has more recently been reported with polyadenylic acid (polyA), specific inositol phosphates (IPs) and heparin. We find that amorphous calcium phosphate (ACP) also demonstrates red-shifted DAPI emission at high DAPI concentrations. This DAPI spectral shift has been attributed to DAPI–DAPI electrostatic interactions enabled by molecules with high negative charge density that increase the local DAPI concentration and favour DAPI molecular proximity, as observed by increasing the dye/phosphate ratio. Excitation of dry DAPI (∼360 nm) confirmed a red-shifted DAPI emission. Whereas enzymatic approaches to modify substrates can help define the nature of DAPI fluorescence signals, multiple approaches beyond red-shifted DAPI excitation/emission are advised before conclusions are drawn about DAPI substrate identification.
Skip Nav Destination
Article navigation
February 2016
-
Cover Image
Cover Image
Scanning electron micrograph of a cell from the endosperm of a barley grain. The cell is tightly packed with large, disk-shaped (A-type) and much smaller, almost spherical (B-type) starch granules. The smooth areas in this image are the surface of the cell walls of neighbouring endosperm cells. For further details see pp. 157-163. Image kindly provided by Elaine Barclay and Vasilios Andriotis (John Innes Centre, Norwich). - PDF Icon PDF LinkTable of Contents
Review Article|
February 09 2016
A cautionary (spectral) tail: red-shifted fluorescence by DAPI–DAPI interactions
Sidney Omelon;
Sidney Omelon
1
*Department of Chemical and Biological Engineering, University of Ottawa, Ottawa, Canada K1N 6N5
1To whom correspondence should be addressed (email somelon@uottawa.ca).
Search for other works by this author on:
John Georgiou;
John Georgiou
†Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, Canada M5G 1X5
Search for other works by this author on:
Wouter Habraken
Wouter Habraken
‡Department of Biomaterials, Max Planck Institute of Colloids and Interfaces, 14476 Potsdam-Golm, Germany
Search for other works by this author on:
Publisher: Portland Press Ltd
Received:
November 20 2015
Online ISSN: 1470-8752
Print ISSN: 0300-5127
© 2016 Authors; published by Portland Press Limited
2016
Biochem Soc Trans (2016) 44 (1): 46–49.
Article history
Received:
November 20 2015
Citation
Sidney Omelon, John Georgiou, Wouter Habraken; A cautionary (spectral) tail: red-shifted fluorescence by DAPI–DAPI interactions. Biochem Soc Trans 15 February 2016; 44 (1): 46–49. doi: https://doi.org/10.1042/BST20150231
Download citation file:
Sign in
Don't already have an account? Register
Sign in to your personal account
You could not be signed in. Please check your email address / username and password and try again.
Captcha Validation Error. Please try again.