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Questions and Answers
What is the formula for calculating the specific activity of each fraction in the purification table?
What is the formula for calculating the specific activity of each fraction in the purification table?
What is the primary purpose of using cuvettes in the purification process?
What is the primary purpose of using cuvettes in the purification process?
What is the role of the phenol-hypochlorite reaction in the enzymatic assay?
What is the role of the phenol-hypochlorite reaction in the enzymatic assay?
What is the significance of the fold purification calculation in the purification process?
What is the significance of the fold purification calculation in the purification process?
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What is the purpose of the literature review in a research article?
What is the purpose of the literature review in a research article?
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What is the unit of measurement for the total activity in the purification table?
What is the unit of measurement for the total activity in the purification table?
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What is the purpose of the yield calculation in the purification process?
What is the purpose of the yield calculation in the purification process?
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What is the significance of the ∆ Abs value in the purification process?
What is the significance of the ∆ Abs value in the purification process?
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What is the role of the urease enzyme in the purification process?
What is the role of the urease enzyme in the purification process?
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What is the purpose of the problem statement in a research article?
What is the purpose of the problem statement in a research article?
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Study Notes
Phenol Red Urease Detection Assay (PRUDA)
- The procedure involves pipetting 10 μL of positive control (urease), negative control (gel mobile buffer), and negative control (testing buffer) into a 96-well plate.
- 10 μL of protein fractions are added to columns 3-10, and 150 μL of testing buffer is added to all wells.
- The plate is incubated at 37°C for 10 minutes and read at 595 nm.
SDS-PAGE Sample Preparation
- A sample buffer is added to 50 μg of protein, and the mixture is heated at 100°C for 5 minutes.
- The sample is then stored at -20°C.
Phenol-Hypochlorite Urease Assay of IEX Pool
- 6 polystyrene cuvettes are set up with 1.25 mL of reagent A and 10 μL of E (or purification product).
- The reaction is started by adding 200 μL of urease assay buffer, and the mixture is incubated at 56°C for 6 minutes.
- The absorbance is read at 637 nm.
Purification Data Analysis
- Specific activity is calculated as total activity (μmol/min) divided by total protein (mg).
- Fold purification is calculated as specific activity at a particular step divided by specific activity of the initial step.
- Yield is calculated as total activity at a particular step divided by total activity of the initial step.
Research Findings Reporting
- Research findings can be reported through different modes of dissemination, including articles, posters, and presentations.
- A typical research article structure includes a literature review, problem statement, proposed solution, and impact of the solution.
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Description
Quiz on laboratory techniques in biochemistry, including Phenol Red Urease Detection Assay (PRUDA) and SDS-PAGE sample preparation.
