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Flourescent Immunostaining Protocol for a-Bungorotoxin (AChRs) in Zebrafish

LJ Lili Jing

Published: Feb 20, 2012 DOI: 10.21769/BioProtoc.183 Views: 13070

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Abstract

Zebrafish embryo is convenient for studying neuromuscular system due to its transparency. This protocol is a rapid method to visualize the AChR clusters on the postsynaptic muscle membrane.

Materials and Reagents

  1. Paraformaldehyde (USB Corporation, catalog number: 19943 )
  2. Phosphate buffered saline (PBS)
  3. DMSO
  4. Triton-100
  5. Na2HPO4
  6. NaH2PO4
  7. Collagenase (Sigma-Aldrich, catalog number: C-9891 )
  8. α-Bungorotoxin conjugated to Alexa 594 (Life Technologies, Invitrogen™, catalog number: B-13423)
  9. Vectashied (Vector Lab, catalog number: H-1400 )
  10. Primary antibody
  11. Secondary antibody
  12. Incubation Buffer (IB) (see Recipes)
  13. 0.1 M phosphate buffer(7.4) (see Recipes)

Equipment

  1. Rotator (Storvall Life Science, the Belly Dancer)
  2. Fluorescence microscope
  3. Transfer pipette

Procedure

Copyright: © 2012 The Authors; exclusive licensee Bio-protocol LLC.

Category

Neuroscience > Development > Immunofluorescence

Biochemistry > Protein > Immunodetection

Cell Biology > Cell imaging > Fluorescence

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