Separazione elettroforetica delle proteine

Biology

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Summary

In questo video, si dimostra un metodo per la separazione elettroforetica di proteine ​​mediante poli-acrylimide elettroforesi su gel (PAG).

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

Elettroforesi viene utilizzata per separare miscele complesse di proteine ​​(per esempio, dalle cellule, frazioni subcellulari, frazioni colonna o immunoprecipitati), per indagare composizioni subunità, e per verificare l'omogeneità dei campioni di proteine. Può anche servire a purificare le proteine ​​per l'uso in altre applicazioni. Elettroforesi in gel di poliacrilamide, proteine ​​migrano in risposta ad un campo elettrico attraverso i pori in una matrice di gel di poliacrilammide; dimensione dei pori diminuisce con l'aumentare la concentrazione di acrilamide. La combinazione di dimensione dei pori e la carica di proteine, le dimensioni e la forma determina il tasso di migrazione della proteina. In questa unità, il metodo standard Laemmli è descritto per elettroforesi su gel discontinuo in condizioni di denaturazione, cioè, in presenza di sodio dodecil solfato (SDS).

Protocol

Il protocollo testo completo di questo approccio sperimentale è disponibile in protocolli di corrente in Biologia Molecolare .

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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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