Elektrophoretische Trennung von Proteinen

Biology

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Summary

In diesem Video zeigen wir eine Methode zur elektrophoretischen Trennung von Proteinen unter Verwendung von Poly-acrylimid Gelelektrophorese (PAGE).

Cite this Article

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Chakavarti, B., Chakavarti, D. Electrophoretic Separation of Proteins. J. Vis. Exp. (16), e758, doi:10.3791/758 (2008).

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Abstract

Elektrophorese wird verwendet, um komplexe Gemische von Proteinen (z. B. von Zellen, subzellulären Fraktionen, Säulenfraktionen oder Immunpräzipitaten) zu trennen, die Untereinheit Kompositionen zu untersuchen und um die Homogenität der Protein-Proben zu überprüfen. Es kann auch dazu dienen, um Proteine ​​für den Einsatz in weiteren Anwendungen zu reinigen. In Polyacrylamidgelelektrophorese, Proteine ​​in Reaktion auf ein elektrisches Feld durch die Poren in einem Polyacrylamid-Gel-Matrix zu migrieren, Porengröße nimmt mit steigender Acrylamid-Konzentration. Die Kombination von Porengröße und Protein Ladung, Größe und Form bestimmt die Wanderungsgeschwindigkeit des Proteins. In dieser Einheit ist die Standard-Laemmli-Methode für die diskontinuierliche Gelelektrophorese unter denaturierenden Bedingungen beschrieben, dh in Gegenwart von Natriumdodecylsulfat (SDS).

Protocol

Der vollständige Text Protokoll für diesen experimentellen Ansatz ist in Current Protocols in Molecular Biology .

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Comments

10 Comments

  1. Hi nice demo but I did not understand why the presenters kept on showing an agarose gel electrophoresed DNA gel in between as a representation instead of a protein gel.

    Reply
    Posted by: Anonymous
    June 15, 2008 - 1:11 PM
  2. SYPROorange and SYPROruby staining - two fluorescent-based protein detection methods - can appear like ethidium bromide stained DNA gels.  So those gels you are speaking of may actually be protein gels. 

    Reply
    Posted by: Anonymous
    June 16, 2008 - 9:34 AM
  3. Yeah possible,  but then why do we see only a single band.I guess SYPRO detection methods would detect the entire protein panel and not specific bands. unless otherwise they are detecting some specific enzyme activity using flourescent detection methods.

    Reply
    Posted by: Anonymous
    June 16, 2008 - 9:52 AM
  4. We loaded BSA.

    Thanks.

    Reply
    Posted by: Anonymous
    June 17, 2008 - 2:12 AM
  5. Please not that we showed polyacrylamide gel electrophoresis and not agarose gel electrophoresis.

    Thanks.

    Bulbul Chakravarti

    Reply
    Posted by: Anonymous
    June 17, 2008 - 2:08 AM
  6. why cannot i see the video...? pls help me..

    Reply
    Posted by: Anonymous
    October 10, 2008 - 3:35 AM
  7. i can see it now..thanks...

    Reply
    Posted by: Anonymous
    October 10, 2008 - 4:50 AM
  8. my network is poor, I can not enjoy the video

    Reply
    Posted by: Huang C.
    March 15, 2009 - 6:30 AM
  9. Hello, Please shoot me an email at nikitab@jove.com and we'll figure something out for you. Cheers, Nikita

    Reply
    Posted by: Anonymous
    March 22, 2009 - 10:26 PM
  10. Hi, thanks what can I download your video ? please help me

    Reply
    Posted by: violet s.
    January 14, 2012 - 10:35 AM

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