Developmental Biology Imaging Techniques Cell Biology

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Authors: Gargioli Cesare 

Introduction

With this technique it is possible to label the blood vessel in vivo in order to analyze in whole mount the quantity and quality of blood vessel.

Reagents

  1. 1.0 ml syringe
  2. China Black Ink
  3. Stadard anesthetic (Ketamine and Xylazine)

Procedure

  1. Anesthetize the mouse
  2. Using a 1 ml syringe withdraw 0,5 ml of mouse blood from the heart ventricle
  3. Inject 1 ml of China Ink and wait for 30 sec (until the eyes, mouth and legs became black)
  4. Quickly dissect and fix in 4% PFA over night at 4°C
  5. Treat the sample with a solution of 0,5% KOH for one day
  6. Treat the sample with a solution of 0,5% KOH/Glycarol 2:1 one day
  7. Treat the sample with a solution of 0,5% KOH/Glycerol 1:1 for one day
  8. Treat the sample with 100% Glycerol for one day (in order to visualize clearly all the inner blood vessels)

Timing

4 days

Critical Steps

Injection in the heart after blood withdrawal

Anticipated Results

eyes, mouth and legs black stained

Figures

Fig 1

Associated Publications

PlGF–MMP-9–expressing cells restore microcirculation and efficacy of cell therapy in aged dystrophic muscle, Cesare Gargioli, Marcello Coletta, Fabrizio De Grandis, Stefano M Cannata, and Giulio Cossu, Nature Medicine 14 (9) 973 - 978 27/07/2008 doi:10.1038/nm.1852

Author information

Gargioli Cesare, S. Raffaele Biomedical Park Foundation

Correspondence to: Gargioli Cesare ([email protected])

Source: Protocol Exchange (2008) doi:10.1038/nprot.2008.161. Originally published online 30 July 2008.

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