Question 1

Which principle does size exclusion chromatography primarily use to separate proteins?

Accepted Answer

Proteins are separated based on their size.

Question 2

In size exclusion chromatography, which type of protein elutes first from the column?

Accepted Answer

Proteins with the largest molecular weight.

Question 3

A mixture contains CytC ($M_r = 12,000$), Immunoglobulin G ($M_r = 145,000$), Ribonuclease A ($M_r = 13,700$), and Serum albumin ($M_r = 68,500$). Which protein will elute last in size exclusion chromatography?

Accepted Answer

CytC

Question 4

Which statement about size exclusion chromatography is FALSE?

Accepted Answer

It separates proteins based on their net charge.

Question 5

A protein elutes earlier from a size exclusion column after treatment with 6 M guanidine hydrochloride (GuHCl). What does this suggest about the protein?

Accepted Answer

The protein has unfolded, increasing its hydrodynamic radius.

Question 6

Which equation best describes the relationship between elution volume ($V_e$) and molecular weight ($M$) in size exclusion chromatography?

Accepted Answer

$V_e = a 	imes \log(M) + b$

Question 7

A heteromeric protein of 60 kDa is treated with 6 M GuHCl and elutes as two peaks of 34 kDa and 26 kDa. What does this indicate?

Accepted Answer

The protein is a heterodimer composed of two different subunits.

Question 8

Which of the following is NOT a reason why size exclusion chromatography is used in protein purification?

Accepted Answer

To separate proteins based on their isoelectric point.

Question 9

In anion-exchange chromatography, which protein will elute first?

Accepted Answer

The protein with the lowest negative charge.

Question 10

A protein mixture is separated by anion-exchange chromatography. The elution order is A, B, C, D, E. Which protein likely has the lowest isoelectric point ($pI$)?

Accepted Answer

Protein E

Question 11

Which buffer condition would best separate proteins using anion-exchange chromatography?

Accepted Answer

A buffer with pH above the $pI$ of all proteins.

Question 12

Which of the following statements about gel filtration chromatography is TRUE?

Accepted Answer

Large proteins elute first because they are excluded from the pores.

Question 13

A protein sample is run through a size exclusion column and elutes at a volume corresponding to 145 kDa. After treatment with a reducing agent, it elutes at 72 kDa. What is the most likely explanation?

Accepted Answer

The protein is a dimer held together by disulfide bonds.

Question 14

Which chromatographic technique would be most appropriate to separate proteins with similar molecular weights but different charges?

Accepted Answer

Ion-exchange chromatography

Question 15

If a protein elutes at a lower volume than expected in size exclusion chromatography, what could be a possible reason?

Accepted Answer

The protein is aggregated into a larger complex.

Question 16

Which of the following best describes the stationary phase in size exclusion chromatography?

Accepted Answer

Porous beads that allow small molecules to enter

Question 17

A protein mixture is separated by size exclusion chromatography. Which protein will have the largest elution volume?

Accepted Answer

The smallest protein

Question 18

Which of the following is a limitation of size exclusion chromatography?

Accepted Answer

It cannot separate proteins with similar sizes.

Question 19

Which chromatographic method would you use to remove salt from a protein solution after ion-exchange chromatography?

Accepted Answer

Size exclusion chromatography

Protein Purification: Size Exclusion and Ion-Exchange Chromatography

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