Unit 5: Analysis of proteins - Practice Quiz

1 What does the abbreviation SDS in SDS-PAGE stand for?

2 What does PAGE stand for in SDS-PAGE?

3 In SDS-PAGE, proteins are separated primarily on the basis of their:

4 What charge does SDS impart to proteins?

5 Which reducing agent is commonly added to the sample buffer to break disulfide bonds in proteins?

6 Which dye is most commonly used to stain and visualize protein bands after SDS-PAGE?

7 Which tracking dye is added to the sample buffer to monitor the migration front during electrophoresis?

8 What is the purpose of running a molecular weight marker (ladder) alongside samples in SDS-PAGE?

9 The two monomers that polymerize to form the polyacrylamide gel are acrylamide and:

10 Which two reagents are used to initiate the polymerization of the polyacrylamide gel?

11 In a typical SDS-PAGE gel, which portion is used to concentrate the proteins into a sharp band before separation?

12 The gel region where actual protein separation by size occurs is called the:

13 Toward which electrode do SDS-coated proteins migrate during electrophoresis?

14 SDS-PAGE is described as qualitative analysis because it primarily tells us about the:

15 In quantitative analysis, the intensity of a Coomassie-stained band is related to the:

16 Which technique is used to measure the intensity of protein bands for quantitative analysis?

17 What is the general relationship between a protein's molecular weight and how far it migrates in SDS-PAGE?

18 Which staining method is more sensitive and can detect very small amounts of protein compared to Coomassie staining?

19 Before loading, protein samples are usually boiled in sample buffer. Why?

20 Which buffer system is most commonly used in the running (electrode) buffer for standard SDS-PAGE?

21 In SDS-PAGE, sodium dodecyl sulfate (SDS) primarily allows separation of proteins based on which property?

22 A protein migrates as a single band under non-reducing conditions but as two bands of different sizes under reducing conditions. What does this indicate?

23 In a discontinuous SDS-PAGE system, what is the main function of the stacking gel?

24 A researcher wants to resolve small proteins in the range of 10–40 kDa. Which acrylamide gel percentage is most appropriate?

25 In SDS-PAGE, why does the relationship between mobility and molecular weight follow a linear plot of versus relative migration distance ()?

26 A protein has a calculated of 0.5. Using a standard curve where , what is its approximate molecular weight?

27 Which staining method offers the highest sensitivity for detecting low-abundance protein bands on an SDS-PAGE gel?

28 During SDS-PAGE sample preparation, why is a tracking dye like bromophenol blue added?

29 A glycoprotein often appears to have a higher apparent molecular weight on SDS-PAGE than its true polypeptide mass. Why?

30 In the Laemmli buffer system, which reagent in the sample buffer is responsible for reducing disulfide bonds?

31 What is the purpose of adding glycerol to the SDS-PAGE loading buffer?

32 A gel run shows 'smiling' bands (edges migrating slower than the center). What is the most likely cause?

33 To quantify a protein band using densitometry, the integrated band intensity is compared against what?

34 Why must the amount of protein loaded stay within a limited range for accurate densitometric quantification?

35 In SDS-PAGE, the resolving gel typically has a pH of about 8.8 while the stacking gel has a pH of about 6.8. This pH difference is essential for what phenomenon?

36 Which catalyst pair initiates and accelerates acrylamide polymerization when casting a gel?

37 Two proteins of identical molecular weight but very different native charges are run on SDS-PAGE. What is the expected result?

38 After Coomassie staining, why is a destaining step performed?

39 If a molecular weight marker lane is run alongside samples, what is its primary purpose?

40 A sample not fully denatured shows an unexpectedly low apparent molecular weight band. What is the most likely reason?

41 A protein with a native molecular mass of 240 kDa migrates as a single band at approximately 60 kDa on a reducing SDS-PAGE gel, but shows two bands at 60 kDa and 30 kDa when the reducing agent is omitted. What is the most consistent interpretation of the quaternary structure?

42 In SDS-PAGE, the relative mobility () of standard proteins plotted against gives the line . An unknown protein migrates with . What is its approximate molecular weight?

43 A highly glycosylated protein of true polypeptide mass 45 kDa consistently migrates at ~70 kDa on standard SDS-PAGE calibrated with non-glycosylated standards. The primary reason for this anomalous migration is:

44 In a discontinuous (Laemmli) SDS-PAGE system, glycine ions in the stacking gel are largely uncharged (isoelectric) at pH 6.8 while chloride is fully charged. What is the direct consequence of this behavior?

45 A researcher observes that a purified 100 kDa protein splits into two bands (~52 kDa and ~48 kDa) only when the sample is heated to 100 °C in loading buffer, but runs as one 100 kDa band when loaded at room temperature. The best explanation is:

46 During densitometric quantification of Coomassie-stained bands, a band appears saturated (non-linear response). Which strategy most reliably restores accurate quantification?

47 A protein migrates faster (appears smaller) than its true mass on SDS-PAGE. Which molecular characteristic is the most likely cause of this anomalous behavior?

48 For resolving proteins in the 10–40 kDa range with maximum resolution, which acrylamide percentage and rationale is most appropriate?

49 A sample shows a smeared, poorly resolved high-molecular-weight streak at the top of the resolving gel, with reduced intensity of the expected monomer band. What is the most probable cause and remedy?

50 Two proteins of identical molecular mass (50 kDa) but very different isoelectric points (pI 4 and pI 10) are run on standard SDS-PAGE. What is the expected outcome?

51 A calibration curve of vs is strongly linear from 20–100 kDa but curves at both extremes on a single-percentage gel. To accurately size a broad mixture (10–250 kDa) in one run, the best approach is:

52 In quantitative SDS-PAGE, silver staining is chosen over Coomassie for a low-abundance sample. A major limitation to keep in mind when quantifying with silver is:

53 A recombinant protein predicted at 27 kDa runs at exactly 27 kDa under reducing SDS-PAGE but at ~24 kDa under non-reducing conditions. This shift most likely indicates:

54 During gel casting, a stacking gel of pH 6.8 and a resolving gel of pH 8.8 are used. If both gels were mistakenly cast at pH 8.8, what would be the most noticeable effect on results?

55 A protein sample gives a single band of 66 kDa, but mass spectrometry shows the true monomer mass is 33 kDa. Reducing agent was present and the sample was boiled. Which explanation is most consistent?

56 For accurate relative quantification of a target band across multiple lanes, why is normalization to a total-protein stain or loading control essential?

57 A membrane protein sample boiled at 100 °C in Laemmli buffer shows a ladder of high-MW bands, whereas incubation at 37 °C for 30 min gives a clean monomer band. The best explanation is:

58 In a Ferguson plot analysis, mobility is measured across gels of varying acrylamide percentage. Two proteins whose lines are parallel but have different y-intercepts differ in:

59 A quantitative assay requires detecting a 5-fold change in a low-abundance protein. Coomassie gives a linear range of roughly 0.5–2 (intensity units). If the low sample reads 0.6 and the high reads 2.5, what is the correct action for reliable quantification?

60 A protein of true mass 55 kDa is phosphorylated at multiple sites and appears as a doublet at ~55 and ~58 kDa on SDS-PAGE. The upward shift of the phosphorylated form is best explained by: