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purplecricket936Lv1

28 Sep 2019

Suppose your lab instructor asked you to make quantitativedeterminations of the following:

a) Total protein concentration in whole blood.
b) Concentration of a solution of purifiedhemoglobin.
c) Concentration of hemoglobin in whole blood.

Would you expect the BCA assay to give accurate values for eachcase (a, b, and c) above? Provide an explanation for your answerfor each case.

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Nelly StrackeLv2

28 Sep 2019

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I have some questions about SDS-page and BCA assay.

1. Why do ascorbic acid, iron, and tryptophan all increase the background in the BCA assay.

A. They oxidize the copper

B. They absorb in the same wavelength

C. They react with the bicinchoninic acid

D. They are reducing agents

2. Why does BCA require a standard curve?

A. the protein sequence influences color development

B. the development of color is time dependent

C. this controls for pipetting mistakes

D. this controls for dilution in the sample

3. If you have an interfering chelator in your sample, what should you do?

A. Give up, chelation is the worst

B. increase the amount of copper with more reagent A

C. increase the amount of copper with more reagent B

D. add an oxidizing reagent

4. A common theme between agarose and acrylamide gels is using different concentrations to separate different molecular weights. Which concentrations separate large species better?

A. high concentration agarose and high concentration acrylamide

B. high concentration agarose and low concentration acrylamide

C. low concentration agarose and high concentration acrylamide

D. low concentration agarose and low concentration acrylamide

5. What would happen if you skipped step 3 of the NextGel protocol?

A. It would be fine

B. The gel would leak out of the casting rig

C. The gel would never polymerize

D. There would not be a well to hold protein

peachpig172

In Proteomics Practicals

a) Why is it necessary to determine the amount and concentration of protein in a sample extract? (short answer)

b) If you use a number of different assays to determine the amount of protein in a sample, would you expect the value to be exactly the same? Explain your answer.

c) If you use a number of different assays to determine the amount of protein in a sample, would you expect the relative values between samples to be the same? In other words, if two samples had a relative total protein amount of 1:2 (twice as much in sample 2) in one assay, would the relative total amount determined by another assay be 1:2?

please answer all the question answering only one question is not helpful .
Practice Calculations on Serial Dilutionsâbe sure to include the amount of water added and include all units

17. Calculate and describe (or diagram) how you would use a 10% SDS solution to prepare a final volume of 20 ml of each of the following solutions: 8%, 4%, 2%, 1% 0.5%.

18. Calculate and describe (or diagram) how you would use a 5 M Tris solution to prepare 200 ml of each of the following solutions: 1M, 100 mM ,10 mM , 1 mM, 0.1 mM.

How much of the first dilution should you make to perform the following dilutions?

19. 300 ml of a 1:5 dilution

20. 100 ml of a 1:10 dilution

21. What is the minimum volume you should pipette with the following? (include units with your answers):

P2 P10 P20 P100 P200 P1000

22. How accurate was your pipetting?

23. Was it better with the larger volume or smaller volume pipettor? Provide rationale.

(If there was a consistent problem with one pipettor please tell the instructor.)

Standard Curves

24. Complete a standard curve for the data above and tape a printed version into your notebook.

25. What is the R-squared value of your trend line? Is this good or badâ¦ i.e. would you repeat the assay?

26. What is the protein concentration of (Explain/show your calculations.)

Unknown #1 = Unknown #2 =

Data for DNA Concentration Procedure

Attach a labeled printout of the plate reader raw data as well as your generated standard curve to this lab.

26. What is the concentration of your unknown DNA sample?

27. What is the R ² for your standard curve data? Why is it important that the R² is >0.95?

28. Comment on the purity of your DNA... discuss- How can you determine the purity of DNA? What are common contaminants of DNA purified from cells?

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