Anatomical Methods

Electron Microscopy Methods

(unless otherwise noted, methods are by David H. Hall)

1. Fixation and Embedding for TEM

Immersion Fixation Protocol for Structure
Microwave-Assisted Fixation for Ultrastructure
Microwave-Assisted Fixation for Immunocytochemistry
High Pressure Freeze (HPF) and Freeze Substitution
Pre-Embedding Antibody Staining for TEM

Older Methods
Conventional two-step fixation
Freeze substitution with tannic acid/osmium_long incubation by Robby Weimer - pdf file
Freeze substitution with tannic acid/osmium_short incubation by Robby Weimer - pdf file
High pressure freezing/freeze-substitution of C. elegans embryos and L1 worms by Richard Fetter
Freeze substitution in Potassium Permanganate by Robby Weimer - pdf file
Metal mirror (slam-freeze) fixation (MMF)
Flat embedding of worms (slam freezing) by Steve Fields (Rand Lab)
Laser hole fixation of embryos by Carolyn Norris (Hedgecock Lab)

2. Fixation and Embedding for SEM

Fixation for Block-Face Imaging, ATUM & FIB/SEM (OTO)
SEM preparation of dissected worms by David Greenstein

3. Sectioning

Serial Thin Sectioning - The Hartwieg Method
Cryo-Thin Sections
Alternate Methods for Post-Staining Thin Sections
Post-Embedding Antibody Staining for TEM
Grid Making
ATUM (automated) Thin Sections

4. Imaging (TEM and SEM)

Serial Block-Face SEM

5. Image Analysis and Serial Reconstruction

Electron Tomography
Computer-Based Analytical Tools

6. See Electron Microscopy Methods in Cell Biology in WormBook

Light and Fluorescence Microscopy Methods (Also see Wormbook-WormMethods section)

Antibody staining of formaldehyde-fixed animals by Gary Ruvkun and Michael Finney
DAPI (4'-6-Diamidino-2-phenylindole) staining of nuclei by Ross Francis (modified by Sudhir Nayak)
DiI and DiO staining methods by multiple sources
FITC (fluorescein isothiocyanate) staining of amphids and phasmids by Ed Hedgecock
FMRFamide staining by Chris Li
Formaldehyde fixation and cytoskeletal staining by Raffi Aroian
Freeze-crack and staining by Janet Duerr
Gonad-intestine staining by Barth Grant
Mounting animals for observation with Nomarski DIC Optics by Monica Driscoll
Staining of early embryos by Bruce Bowerman
SYTO staining of nucleic acids in live worms by Hill and L'Hernault