Journal
/
/
En forbedret protokol til at rense og direkte Mono-Biotinylate Recombinant BDNF i et rør for Cellular Trafficking Studies i Neuroner
JoVE Journal
Neuroscience
Author Produced
A subscription to JoVE is required to view this content.  Sign in or start your free trial.
JoVE Journal Neuroscience
An Improved Protocol to Purify and Directly Mono-Biotinylate Recombinant BDNF in a Tube for Cellular Trafficking Studies in Neurons
DOI:

13:46 min

July 11, 2020

, , , ,

Chapters

  • 00:00Introdcution
  • 00:48Transfection of HEK293 cells
  • 01:49Media Collection and Storage
  • 03:02Media Concentration and Purification
  • 04:58In vitro Mono-biotinylation Reaction
  • 06:01Biotinylation Analysis
  • 06:56Visualization of Retrograde Axonal Transport of BDNF-QD in Live Neurons
  • 07:58Represetative Results
  • 13:17Conclusion

Summary

Automatic Translation

Rekombinant BDNF indeholdende en Avi-sekvens (BDNFAvi) fremstilles i HEK293-celler på en omkostningseffektiv måde og renses ved affinitetskromatografi. BDNFavi monobiotinyliseres derefter direkte med enzymet BirA i et rør. BDNFavi og mono-biotinyleret BDNFavi bevarer deres biologiske aktivitet sammenlignet med kommercielt tilgængelige BDNF.

Related Videos

Read Article