Biophysical Journal 72: 1986-1996 (1997)
© 1997 the Biophysical Society

This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Gadella, T W Articles by Bisseling, T Search for Related Content PubMed PubMed Citation Articles by Gadella, T W, Jr Articles by Bisseling, T

Microspectroscopic imaging of nodulation factor-binding sites on living Vicia sativa roots using a novel bioactive fluorescent nodulation factor.

Department of Molecular Biology, Wageningen Agricultural University, The Netherlands. dorus.gadella@laser.bc.wau.nl

ABSTRACT

A novel bioactive fluorescent nodulation (Nod) factor, NodRlv-IV(BODIPY FL-C16), has been synthesized by attaching a BODIPY FL-C16 acyl chain to the primary amino group of chitotetraose deacetylated at the nonreducing terminus by recombinant NodB. The binding of the fluorescent Nod factor to root systems of Vicia sativa was investigated with fluorescence spectral imaging microscopy (FSPIM) and fluorescence ratio imaging microscopy (FRIM). Spatially resolved fluorescence spectra of living and labeled Vicia sativa root systems were measured by FSPIM. Strong autofluorescence, inherent to many plant systems when excited at 488 nm, was corrected for by utilizing the difference in fluorescence emission spectra of the autofluorescence and NodRlv-IV(BODIPY FL-C16). A methodology is presented to break down the in situ fluorescence emission spectra into spatially resolved autofluorescence and BODIPY FL fluorescence spectra. Furthermore, an FRIM method was developed for correcting autofluorescence in fluorescence micrographs for this system. After autofluorescence correction it was shown that NodRlv-IV(BODIPY FL-C16) was concentrated in the root hairs, but was also bound to other parts of the root surface.

This article has been cited by other articles:

				G.-J. Kremers, E. B. van Munster, J. Goedhart, and T. W. J. Gadella Jr. Quantitative Lifetime Unmixing of Multiexponentially Decaying Fluorophores Using Single-Frequency Fluorescence Lifetime Imaging Microscopy Biophys. J., July 1, 2008; 95(1): 378 - 389. [Abstract] [Full Text] [PDF]

				M. T. Fujiwara, A. Nakamura, R. Itoh, Y. Shimada, S. Yoshida, and S. G. Moller Chloroplast division site placement requires dimerization of the ARC11/AtMinD1 protein in Arabidopsis J. Cell Sci., May 1, 2004; 117(11): 2399 - 2410. [Abstract] [Full Text] [PDF]

				E. Limpens, J. Ramos, C. Franken, V. Raz, B. Compaan, H. Franssen, T. Bisseling, and R. Geurts RNA interference in Agrobacterium rhizogenes-transformed roots of Arabidopsis and Medicago truncatula J. Exp. Bot., May 1, 2004; 55(399): 983 - 992. [Abstract] [Full Text] [PDF]

				F. J. M. van Kuppeveld, W. J. G. Melchers, P. H. G. M. Willems, and T. W. J. Gadella Jr. Homomultimerization of the Coxsackievirus 2B Protein in Living Cells Visualized by Fluorescence Resonance Energy Transfer Microscopy J. Virol., August 12, 2002; 76(18): 9446 - 9456. [Abstract] [Full Text] [PDF]

				W. D'Haeze and M. Holsters Nod factor structures, responses, and perception during initiation of nodule development Glycobiology, June 1, 2002; 12(6): 79R - 105R. [Abstract] [Full Text] [PDF]