Kleuring Eiwitten in Gels

Biology
 

Summary

Na de scheiding door elektroforetische methoden, kunnen eiwitten in een gel worden gedetecteerd door verschillende kleuren methoden. Kleuring van eiwitten met Coomassie Blauw, Zilver kleuring, SYPRO Orange, SYPRO Ruby worden gedemonstreerd in deze video.

Cite this Article

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Gallagher, S., Chakavarti, D. Staining Proteins in Gels. J. Vis. Exp. (17), e760, doi:10.3791/760 (2008).

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Abstract

Na de scheiding door elektroforetische methoden, kunnen eiwitten in een gel worden gedetecteerd door verschillende kleuren methoden. Deze eenheid wordt beschreven protocollen voor het detecteren van eiwitten door vier populaire methoden. Coomassie blauwe kleuring is een makkelijke en snelle methode. Zilverkleuring, terwijl meer tijd in beslag, is aanzienlijk gevoeliger en kan dus worden gebruikt om kleinere hoeveelheden eiwit op te sporen. Fluorescerende kleuring is een populair alternatief voor de traditionele kleuren van de procedures, vooral omdat het gevoeliger dan Coomassie kleuring, en wordt vaak zo gevoelig als zilverkleuring. Kleuring van eiwitten met SYPRO Orange en SYPRO Ruby zijn ook hier aangetoond.

Protocol

De volledige tekst protocol voor deze experimentele aanpak is beschikbaar in Current Protocols in Molecular Biology .

Comments

8 Comments

  1. WHICH REACTION EQUATION OF CONVERT Cu+² TO Cu+1 in the protein test

    Reply
    Posted by: Anonymous
    August 3, 2008 - 7:45 AM
  2. thank you!very good!

    Reply
    Posted by: Anonymous
    September 6, 2008 - 8:51 PM
  3. It is very good, thanks

    Reply
    Posted by: Anonymous
    November 12, 2008 - 6:43 AM
  4. A good and cheap alternative to SYPRO Rubi is RuBPS staining. Here are some protocols. More information can be found on www.ruthenium.ag.vu   RuBPS staining protocol I (quality) 1. Fix the gel in 30% EtOH, 10% acetic acid overnight ². Rinse the gel in ²0% EtOH for 30 min and repeat 3 times 3. Incubate the gel in 1 mM RuBPS solution for 6 h 4. Equilibrate the gel in water for 10 min and repeat once 5. Destain the gel with 40% EtOH/10% acetic acid for 15 h 6. Equilibrate the gel in water for 10 min repeat once and scan   all% are in V/V   Procedure as published in Proteomics ²004, 4, 599–608.     RuBPS staining protocol II (fast)   1. Incubate the gel in 50 ml of 40% Ethanol/10% acetic acid containing    1 mM RuBPS for 1 h. ². Destain the gel for ²0 min in 40% Ethanol/10% acetic acid 3. Wash the gel for 10 min in water and scan   all% are in V/V   Procedure as published in PLoSONE. ²007 Feb ²8;²(²)e²63.       RuBPS staining protocol III (co-electrophoretical)   1. Add 1 ml of ²0 mM stock solution to one pocket of your SDS gel. Run     the gel according to your standard procedure. ². Incubate the gel in 50 ml of 40% Ethanol/10% acetic acid for ²0 min 3. Incubate the gel in 50 ml water for 10 min and scan       RuBPS staining protocol IV (loading buffer)   1. Add 1 ml of ²0 mM stock solution to your loading buffer (omit all other     dyes like bromophenol blue). Run the gel according to your standard     procedure. ². Incubate the gel in 50 ml of 40% Ethanol/10% acetic acid for ²0 min 3. Incubate the gel in 50 ml water for 10 min and scan  

    Reply
    Posted by: Anonymous
    February 16, 2009 - 1:20 PM
  5. Itz realy ² gud!

    Reply
    Posted by: sathya r.
    May 18, 2009 - 11:01 AM
  6. Thank yoy

    Reply
    Posted by: JASSIM A.
    May 20, 2009 - 9:39 AM
  7. thank you

    Reply
    Posted by: ibrahim b.
    August 27, 2010 - 12:31 AM
  8. thanks! This is very useful for my job!

    Reply
    Posted by: Anonymous
    August 31, 2010 - 4:33 AM

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