Separation of Proteins by

Gel Electrophoresis

Carolina Biological Kit #68-9800

Presented by

Bryan Smith

Why this Kit?

• Just Curious!– Done chemical tests for proteins– Done enzyme activity labs– Done Gel Electrophoresis to analyze DNA by

separating molecules w/different #Bp– Read about different methods of protein

separation– Never use any methods while in school or in

teaching

What do we know about protein structure?

• Building blocks?• Primary structure?

• Secondary structure?

• Tertiary Structure?

• Quaternary Structure?

What makes one Protein Different from another?

There are two properties of proteins used to separate them

• Size or MW

• Electric Charge

This lab will use Agarose gel electrophoresis to separate proteins

Lab particulars

• Our protein sample contains two pre-stained proteins both w/overall negative charge– Albumin– Hemoglobin

• Work in pairs • Gel is 1% agarose • Class will run two gels

Procedures

• Pour two gels using largest well comb• After solidifying, remove comb and orient so

wells are on “negative” end• Fill chamber with TBE buffer• Each pair will load one 40uL sample of protein

using micropipette• Voltage?

Go over discussion questions

How many different scenarios can you think of where separating proteins is essential to a biologist?XXXX – isolating insulin from pancreatic cells