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BIOL 10016 Experiment 5 Using Ion Exchange Chromatography to Separate Proteins (2023)(NEW)
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BIOL 10016 Experiment 5 Using Ion Exchange Chromatography to Separate Proteins
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MOHAWK COLLEGE OF APPLIED ARTS AND TECHNOLOGYCHEMICAL ,ENVIRONMENTAL & BIOTECHNOLOGY DEPARTMENT
Lab Report
ROOM NO: FE E309
Class : BIOL 10016 Lab.
EXPERIMENT NO : 5
TITLE : Using Ion Exchange Chromatography to Separate Proteins
Submitted by :
Partners :
Instructor : Farag Soliman
Date lab performed : March 24, 2023
Date of submission : March 31, 2023
FENNELL CAMPUS
HAMILTON, ONTARIO
, Purpose
The purpose of using ion exchange chromatography (IEC) to separate proteins is
to exploit the differences in their net charge at a given pH, which allows for the
selective retention and separation of proteins with distinct charge properties. IEC
is particularly useful for the separation and purification of proteins in complex
mixtures and can be applied to a wide range of proteins with varying charges,
sizes, and shapes.
In the specific case of separating lysozyme and albumin, these two proteins exhibit
opposing charges at pH 7.2. Lysozyme carries a positive net charge, while albumin
has a negative net charge. This contrast in charge characteristics provides an
opportunity to separate them using ion exchange chromatography. By selecting
the appropriate ion exchange resin and optimizing the chromatographic conditions,
it is possible to retain lysozyme and albumin on the stationary phase based on
their respective charges, ultimately facilitating their separation from each other.
Procedure
Biotechnology 1 Laboratory Manual BIOL 10016 Lab. Experiment 5 Using Ion
Exchange Chromatography to Separate Proteins. Pages 2-4.
Materials
Tools
Test Tubes
Chromatographic Column
Micropipette
Reagents
Equilibrium buffer (50 mM Sodium monophosphate, pH 7.2)
Elution Buffer (50 mM Sodium monophosphate/0.5 M NaCl, pH 7.2)
Lysozyme protein solution 0.2 mg/ml
Albumin protein solution 0.1 mg/ml
Lysozyme + Albumin protein solution
BRADFORD Reagent Refrigerate
DEAE SEPHAROSE anion exchange resin
Instrument
UV-Spectrophotometer
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