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Bovine Serum Albumin (BSA) is a globular protein derived from cattle. The objective of this laboratory experiment was to determine the concentration of BSA in an unknown solution (test tube 8) by comparing the absorbance levels at 540 nm with those of known BSA concentrations. The Biuret reagent was added to the BSA solutions, and the resulting color saturation was observed. The hypothesis posited a direct relationship between color change and BSA concentration, enabling the accurate determination of the unknown BSA concentration.
The experiment involved eight test tubes, each filled with known BSA concentrations and an unknown BSA concentration (provided by the instructor).
To these tubes, 3 ml of Biuret reagent was added, and gentle mixing was performed. A 30-minute incubation period allowed the reagent to react with BSA. The standard BSA concentrations for each test tube were provided in the lab manual.
| Tube | ml Water | Biuret ml | BSA standard ml | BSA unknown | Final concentration of BSA (mg/ml) |
|---|---|---|---|---|---|
| 1 | 3.0 | 0.0 | 0.0 | 0.0 | 0.00 |
| 2 | 2.8 | 0.2 | 0.0 | 0.10 | |
| 3 | 2.6 | 0.4 | 0.0 | 0.20 | |
| 4 | 2.3 | 0.7 | 0.0 | 0.35 | |
| 5 | 2.0 | 1.0 | 0.0 | 0.50 | |
| 6 | 1.5 | 1.5 | 0.0 | 0.75 | |
| 7 | 1.0 | 2.0 | 0.0 | 1.00 | |
| 8 | 2.0 | 0.0 | 1.0 | ??? |
The table displays the BSA standard concentrations in each test tube and the final BSA concentration (mg/ml) after adding the Biuret reagent.
After incubation, the absorbance of each solution was measured at 540 nm using a spectrophotometer.
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It was essential to zero the spectrophotometer with the reagent blank tube (tube 1) before measurement.
The following table displays the absorbance values at 540 nm for each test tube:
| Tube | Absorbance at 540 nm |
|---|---|
| 1 | 0.000 |
| 2 | 0.025 |
| 3 | 0.052 |
| 4 | 0.084 |
| 5 | 0.141 |
| 6 | 0.205 |
| 7 | 0.296 |
| 8 | 0.081 |
The absorbance values measured at 540 nm for each test tube.
The BSA concentration of test tube 8 was calculated to be 0.299 mg/ml.
The experiment revealed a positive correlation between BSA concentration and absorbance rate.
Test tube 8 had an absorbance of 0.081 at 540 nm, similar to test tube 4 with an absorbance of 0.084 at 540 nm. Consequently, it was inferred that the BSA concentration in test tube 8 was akin to that in test tube 4. Indeed, the BSA concentration in test tube 8 was calculated to be 0.299 mg/ml, while test tube 4 had a standard BSA concentration of 0.35 mg/ml. Test tubes 8 and 4 exhibited the closest values of BSA concentration and absorbance, aligning with the hypothesis that BSA concentration could be determined accurately by comparing absorbance changes and their correlation with known BSA concentrations.
Recognizing proteins in samples is crucial for medical purposes. For instance, protein concentration measurement in urine samples can help detect conditions like pregnancy, leukemia, diabetes, and HIV/AIDS (Bank, 2017).
To enhance the lab, more accurate measurements could have been taken, and color interpretation could have been improved. Accurate measurements would contribute to more precise BSA concentration determinations, while refined color interpretation would reduce potential errors.
The experiment successfully determined the BSA concentration in an unknown solution by comparing its absorbance at 540 nm with that of known BSA concentrations. The positive correlation between BSA concentration and absorbance supported the accurate determination of the unknown BSA concentration. This practical application of spectrophotometry has implications in medical diagnostics, where protein concentration measurement plays a vital role in detecting various medical conditions.
Lab Report: Determination of BSA Concentration. (2024, Jan 03). Retrieved from https://studymoose.com/document/lab-report-determination-of-bsa-concentration
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