|
|
|
|
 |
|||
|
|||
Neuronal tracing with DiI: decalcification, cryosectioning, and photoconversion for light and electron microscopic analysis
CS von Bartheld, DE Cunningham and EW Rubel
Hearing Development Laboratories, University of Washington Medical School, Seattle 98195.
The molecule 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI) is a fluorescent dye which diffuses within cell membranes. The properties of DiI diffusion and fluorescence are maintained in aldehyde-fixed tissue, thereby allowing selective neuronal tracing post mortem. We describe three modifications of this tracing method. First, while DiL diffuses along neuronal membranes the tissue can be decalcified in EDTA at 37 degrees C. Tracing in decalcified tissue extends the possible application of the DiI technique to the investigation of neuronal tissue enclosed in bony structures. Second, we describe a protocol that allows sectioning of DiI-injected tissue on a cryostat with minimal subsequent spread of DiI in dried sections. Third, we demonstrate that DiI label of fluorescent neurons in cryosections as well as Vibratome sections can be photo- oxidated and converted to a stable diaminobenzidine reaction product. The photo-converted DiI label is electron dense and allows analysis of labeled cell bodies and processes at the electron microscopic level. DiI does not stay confined to the surface cell membrane in fixed tissue but reaches internal organelles, presumably via membranes of the endoplasmic reticulum, and concentrates in microsomal structures adjacent to mitochondria. Photoconversion of DiI label is compatible with gold immunocytochemistry. Long-term incubation and subsequent photoconversion of post-mortem DiI-labeled neurons provides remarkable tissue preservation at the ultrastructural level.
Volume 38,
Issue 5,
pp. 725-733,
05/01/1990
Copyright © 1990 by The Histochemical Society
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati What's this?
This article has been cited by other articles:
|
|
 |
|
  Y. Momose-Sato, Y. Honda, H. Sasaki, and K. Sato Optical Mapping of the Functional Organization of the Rat Trigeminal Nucleus: Initial Expression and Spatiotemporal Dynamics of Sensory Information Transfer during Embryogenesis J. Neurosci., February 11, 2004; 24(6): 1366 - 1376. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. C. Tsai, R. L. van Bendegem, S. W. Hwang, and C. H. Tator A Novel Method for Simultaneous Anterograde and Retrograde Labeling of Spinal Cord Motor Tracts in the Same Animal J. Histochem. Cytochem., September 1, 2001; 49(9): 1111 - 1122. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. S. von Bartheld and R. Butowt Expression of Neurotrophin-3 (NT-3) and Anterograde Axonal Transport of Endogenous NT-3 by Retinal Ganglion Cells in Chick Embryos J. Neurosci., January 15, 2000; 20(2): 736 - 748. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. P. Dantuma, M. A. P. Pijnenburg, J. H. B. Diederen, and D. J. Van der Horst Electron Microscopic Visualization of Receptor–mediated Endocytosis of DiI–labeled Lipoproteins by Diaminobenzidine Photoconversion J. Histochem. Cytochem., September 1, 1998; 46(9): 1085 - 1090. [Abstract] [Full Text]
|
|
|
|
|
|
J.&n.;R. Lukas, M. Aigner, M. Denk, H. Heinzl, M. Burian, and R. Mayr Carbocyanine Postmortem Neuronal Tracing: Influence of Different Parameters on Tracing Distance and Combination with Immunocytochemistry J. Histochem. Cytochem., August 1, 1998; 46(8): 901 - 910. [Abstract] [Full Text]
|
|
|
|
|
|
T. Seki and U. Rutishauser Removal of Polysialic Acid-Neural Cell Adhesion Molecule Induces Aberrant Mossy Fiber Innervation and Ectopic Synaptogenesis in the Hippocampus J. Neurosci., May 15, 1998; 18(10): 3757 - 3766. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T Bossing and G. Technau The fate of the CNS midline progenitors in Drosophila as revealed by a new method for single cell labelling Development, January 7, 1994; 120(7): 1895 - 1906. [Abstract] [PDF]
|
|
| Guidelines | Subscriptions | About | exPRESS - Current - Archive | Business Information | Contact |