Protein Assay Reflection.
Objectives
Procedure
1. 20 g of fish flesh for freshwater fish and marine fish is weighed using analytical balance.
2. After weighing, the fish flesh is sliced into smaller size
3. Smaller size of fish is blend in the blender and Phosphate Buffer saline at 1:10 ratio is added
4. The fish is blended separately between freshwater fish and marine fish.
5. Then it is incubate for 24 hours at 4C in 100 rpm shaker
6. After 24 hours, the sample is filter to obtain the pure stock by using filter funnel and filter paper.
7. Pure stock that is obtained is put into centrifuge tube using micropipette
8. Then the centrifuge tube is centrifuge at 4C, 1400 rpm, 15 minutes.
9. After centrifuge, the protein content is measured by using Lowry or Biuret method
10. As for the Biuret method that we used in this experiment, we mix 0.50 mL of protein with 2.50 mL of Biuret reagent.
11. To measure the abosrbance, we use spectrophotometer
12. Then, the stocks is put into a cuvette and we measure the absorbance at 540 nm after 10 minutes.
13. Step 12 is repeated five times for each marine fish and freshwwater fish so that we are able to obtain the average value of the absorbance we needed.
14. Protein absorbance for each sample we used is then calculated by using Biuret equation y=0.1276 In (x) + 0.0346
Result:
- To measure the protein concentration in marine fish and freshwater fish by using Lowry and Biuret method.
- To compare the protein concentration between freshwater fish and marine fish.
Procedure
1. 20 g of fish flesh for freshwater fish and marine fish is weighed using analytical balance.
2. After weighing, the fish flesh is sliced into smaller size
3. Smaller size of fish is blend in the blender and Phosphate Buffer saline at 1:10 ratio is added
4. The fish is blended separately between freshwater fish and marine fish.
5. Then it is incubate for 24 hours at 4C in 100 rpm shaker
6. After 24 hours, the sample is filter to obtain the pure stock by using filter funnel and filter paper.
7. Pure stock that is obtained is put into centrifuge tube using micropipette
8. Then the centrifuge tube is centrifuge at 4C, 1400 rpm, 15 minutes.
9. After centrifuge, the protein content is measured by using Lowry or Biuret method
10. As for the Biuret method that we used in this experiment, we mix 0.50 mL of protein with 2.50 mL of Biuret reagent.
11. To measure the abosrbance, we use spectrophotometer
12. Then, the stocks is put into a cuvette and we measure the absorbance at 540 nm after 10 minutes.
13. Step 12 is repeated five times for each marine fish and freshwwater fish so that we are able to obtain the average value of the absorbance we needed.
14. Protein absorbance for each sample we used is then calculated by using Biuret equation y=0.1276 In (x) + 0.0346
Result:
What is the new thing i learn in this experiment?
well, for the first time in my life i can measure the content of protein in a fish on my own ! im so excited to learn the two new method. I also learned about the centrifuge mean and function. This lab seriously attract me soo much.
well, for the first time in my life i can measure the content of protein in a fish on my own ! im so excited to learn the two new method. I also learned about the centrifuge mean and function. This lab seriously attract me soo much.