PRACTICAL FOR ELECTROPHORESIS OF SERUM PROTEIN

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LABORATORY REPORT
PRACTICAL 2
(ELECTROPHORESIS OF SERUM PROTEIN)

DATE : 10 OCTOBER 2013

PART A
SEPARATION OF SERUM PROTEINS USING THE ELECTROPHORESIS METHOD.

OBJECTIVES:
1. To understand the analytical methods involved in analyzing serum proteins.
2. To study the serum protein electrophoresis pattern to aid the understanding of the relationship between the structure and the function of proteins.

PRINCIPLE OF PRACTICAL:
This technique is based on the movement of charged particles such as proteins when placed in an electric field. The movement of these particles is based on their charges where positively charged particles move towards cathode while negatively charged particles move towards anode. The rate of movement is dependent on factors such as size and charges.
This technique is useful in diagnosis. For example, in diagnosing myocardial infection, electrophoresis is used to separate isoenzymes of lactase dehydrogenase and creatine kinase. Electrophoresis can also be used to separate blood proteins for accurate diagnosis of protein impaired metabolism such as patient with abnormal haemoglobin. Apart from that, electrophoresis can also be used as a method to separate and purify proteins.

MATERIALS AND REAGENTS:
1. Electrophoresis tank with voltage power supply.
2. 2.5 x 12 cm sized cellulose acetate paper.
3. Paper to line the table.
4. Tray for the dye and acetic acid.
5. Tray for the buffer.
6. Forceps.
7. Capillary tubes.
8. Clock.
9. Filter paper.
10. Scissors.
11. Spectrophotometer (570 nm wavelength)
12. 5% (v/v) of acetic acid (glacial acetic acid 50 ml/L)
13. 0.2% of Ponceau S that contains:
Ponceau S (2g) 1 Liter
Trichloroacetic acid (30g)
14. 0.06 M barbitone buffer at pH 8.2 and pH 8.6 contains:
Diethylbarbiturate acid (1.84g) 1 Liter
Sodium diethylbarbiturate (10.3g)
15. 0.25 M of sodium hydroxide, NaOH solution.
16. 0.1 ml serum.



References: Glucose-6-phosphate Dehydrogenase Deficiency, Dr. Rull G., (2011). http://www.patient.co.uk/doctor/Glucose-6-phosphate-dehydrogenase-deficiency.htm, Retrieved on October 10, 2013. Blood:NADPH, Not Glutathione, Status Modulates Oxidant Sensitivity in Normal and Glucose-6-Phosphate Dehydrogenase-Deficient Erythrocytes, Scott M.D., Zuo L., Lubin B.H., & Chiu D.T., (2013). American Society of Hematology. Washington DC.

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