18 Questions
What is the approximate pH of the electrophoresis buffer?
pH 8.3
What is the role of glycine in the electrophoresis buffer?
To set up a steep voltage gradient in the stacking gel
What is the primary effect of the chloride ions in the tank buffer and stacking gel?
To form a moving ion front
What is the purpose of the large pores in the stacking gel?
To concentrate protein-SDS complexes
What happens when the glycine ions enter the running gel?
Dramatic changes occur
What is the result of the difference in chloride and glycine mobility?
A steep voltage gradient in the stacking gel
What is the effect of the alkaline pH of the running gel on glycine molecules?
They assume a negative charge
What is the primary cause of proteins migrating at different rates in the running gel?
The sieving properties of the gel
What is the purpose of pre-stained protein standards in electrophoresis?
To monitor the approximate positions of proteins during electrophoresis
What is the effect of adding chromophores to a protein standard on the gel?
It produces more diffuse bands on the gel
What is the purpose of rinsing the gel with water after electrophoresis?
To remove the buffer salts used for electrophoresis
What is the characteristic of the stain Coomassie Brilliant Blue G-250?
It binds non-covalently and with very little specificity to proteins
What is the main function of the electrophoresis buffer in SDS-PAGE?
To provide a consistent ionic strength and pH environment
What are the two main components of the SDS-PAGE system?
Stacking and running gels
What is the role of ammonium persulfate in the SDS-PAGE system?
To generate free oxygen radicals that catalyze the polymerization of acrylamide and bis-acrylamide
What is the effect of increasing the concentration of acrylamide or bis-acrylamide in the gel matrix?
It decreases the pore size of the gel
What is the function of glycerol in the SDS-PAGE system?
To allow protein samples to settle into the bottom of the gel wells
What is the difference between the stacking and running gels?
They have different pore sizes, ionic strengths, and pHs
Learn about the composition and role of the electrophoresis buffer in SDS-PAGE, including its pH and glycine ionization. Understand how it affects protein separation. Test your knowledge of this critical step in protein analysis.
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