← Back to 1. The Foundations of Biochemistry

📝 isopycnic centrifugation (11 MCQs)

📖 From Principles of Biochemistry • 1. The Foundations of Biochemistry • 11 questions available

←Previous Topic:Differential Centrifugation and Cell Fractionation📝 isopycnic centrifugationOrganelle isolation by cell fractionation:Next Topic→

What is isopycnic centrifugation?

Definition:
Isopycnic centrifugation is a high-resolution density-based separation technique where macromolecules or subcellular particles are centrifuged in a density gradient until they reach a position where their own buoyant density exactly matches the density of the surrounding medium, meaning they have reached isopycnic (equal density) equilibrium, with no further movement, allowing for the purification of molecules like DNA, RNA, and lipoproteins.

Working:
This method works by using a pre-formed gradient of a dense medium (e.g., cesium chloride or sucrose), and under ultracentrifugation at high speeds for extended periods, each particle sediments to the point where its density equals that of the gradient, following the equation $\rho_p = \rho_m$ at equilibrium, where $\rho_p$ is the particle density and $\rho_m$ is the medium density, and particles with different densities form distinct bands at their respective isopycnic positions.

Example:
A classic example is the separation of DNA molecules with different guanine-cytosine (GC) content, where DNA is mixed with cesium chloride and centrifuged at 140,000 × g for 24 hours, and the DNA molecules form bands at different positions based on their density, allowing the isolation of pure DNA fractions for downstream analysis.

Reason:
Isopycnic centrifugation is essential in molecular biology and biochemistry for purifying nucleic acids, viruses, and organelles, as it provides high purity and resolution, and is widely used in research for studying macromolecular structure, function, and interactions.

Easy

Medium

Hard

📝 All isopycnic centrifugation MCQs

Q1. A researcher wants to separate DNA molecules that differ in buoyant density rather than simply by molecular size. Which principle makes isopycnic centrifugation especially suitable for this goal?

A.Molecules stop when they reach a position where their density matches the surrounding medium ✅

B.Molecules stop when their molecular masses become equal to the rotor speed

C.Molecules separate because larger molecules always sediment more slowly

D.Molecules remain permanently at the top because the medium is denser

💡 Difficulty: medium | ✅ Correct: A

📖 Explanation: In isopycnic centrifugation, particles migrate through a density gradient until they reach a position where their buoyant density matches that of the surrounding medium. Separation therefore depends primarily on density rather than simply molecular size or mass.

Q2. A sample contains two DNA populations with buoyant densities of $1.70$ and $1.75$ g/mL. The gradient spans densities from $1.60$ to $1.80$ g/mL. After sufficient centrifugation, what should happen if equilibrium is successfully reached?

A.Both DNA populations form bands at the bottom of the tube

B.Both populations remain at their original loading positions

C.Each DNA population forms a band near the position where the gradient density equals its own density ✅

D.The less dense DNA disappears from the gradient

💡 Difficulty: medium | ✅ Correct: C

📖 Explanation: At equilibrium, each DNA population migrates until the surrounding solution has approximately the same density as the DNA. Since both densities fall within the gradient range, distinct bands should form at different positions.

Q3. A student argues that increasing centrifugation time will eventually force every DNA molecule to the bottom of an isopycnic gradient. Which response best evaluates the argument?

A.Correct, because centrifugation always overcomes buoyancy

B.Incorrect, because particles at density equilibrium have no net tendency to continue moving downward ✅

C.Correct, because DNA becomes denser during centrifugation

D.Incorrect, because DNA cannot move through density gradients

💡 Difficulty: hard | ✅ Correct: B

📖 Explanation: The student's reasoning confuses sedimentation with equilibrium positioning. Once a particle reaches a region whose density matches its own buoyant density, the opposing effects of sedimentation and buoyancy balance, so it does not simply continue toward the bottom.

Q4. A laboratory accidentally prepares a gradient whose maximum density is $1.72$ g/mL, while one DNA species has a buoyant density of $1.76$ g/mL. What is the most likely consequence?

A.That DNA species will form an equilibrium band at

1.76

g/mL inside the tube

B.That DNA species cannot reach its matching-density position within the prepared gradient ✅

C.Both DNA species will automatically have identical band positions

D.The DNA species will become chemically converted into a lower-density form

💡 Difficulty: medium | ✅ Correct: B

📖 Explanation: An equilibrium band requires the gradient to contain a region with a density matching the particle's buoyant density. Because $1.76$ g/mL exceeds the gradient's maximum of $1.72$ g/mL, that equilibrium position is unavailable.

Q5. Two DNA samples are centrifuged in the same density gradient. Sample X forms a band higher in the tube than Sample Y. Assuming the gradient becomes progressively denser toward the bottom, which interpretation is most reasonable?

A.X has a higher buoyant density than Y

B.X has a lower buoyant density than Y ✅

C.X must have a larger molecular mass than Y

D.Y must have a smaller molecular mass than X

💡 Difficulty: hard | ✅ Correct: B

📖 Explanation: In a gradient that becomes denser downward, a less-dense particle reaches its matching-density position higher in the tube, while a denser particle travels farther downward. Therefore, X's higher band indicates a lower buoyant density than Y.

Q6. A graph of band position versus buoyant density shows that as DNA buoyant density increases, the equilibrium band moves progressively toward the bottom of the tube. A new DNA sample produces a band between two known standards. What is the best experimental strategy?

A.Estimate the unknown density from its position relative to the standards ✅

B.Assume the unknown has the same density as the upper standard

C.Use molecular mass alone to determine its density

D.Ignore the band position because equilibrium gradients cannot provide density information

💡 Difficulty: medium | ✅ Correct: A

📖 Explanation: Known standards establish a relationship between band position and buoyant density. If the unknown band lies between two standards, its density can be estimated by interpolation, provided the gradient and experimental conditions are comparable.

Q7. A researcher compares isopycnic centrifugation with ordinary sedimentation-based separation. The researcher needs to distinguish particles that have similar sizes but different densities. Which method and reasoning are most appropriate?

A.Isopycnic centrifugation, because equilibrium positions depend strongly on buoyant density ✅

B.Ordinary sedimentation, because size is the only relevant property

C.Either method must give identical results because all particles have mass

D.Ordinary sedimentation, because density differences prevent equilibrium bands

💡 Difficulty: hard | ✅ Correct: A

📖 Explanation: Isopycnic centrifugation is particularly useful when density differences are important. Particles with similar size can nevertheless occupy different equilibrium positions if their buoyant densities differ, allowing density-based discrimination.

Q8. A student observes that two DNA bands are very close together and concludes that the molecules must have identical buoyant densities. What is the strongest criticism of this conclusion?

A.Close bands can still represent different densities if the gradient or detection resolution is limited ✅

B.Identical densities always produce bands at opposite ends of the tube

C.Band spacing is determined only by DNA concentration

D.A density gradient cannot produce more than one DNA band

💡 Difficulty: easy | ✅ Correct: A

📖 Explanation: Band position reflects buoyant density, but experimental resolution is finite. Two populations with slightly different densities may produce closely spaced bands. Therefore, visual proximity alone does not prove that their densities are identical.

Q9. A density gradient is represented schematically by increasing solution density from top to bottom. Three DNA populations have densities $1.65$ , $1.70$ , and $1.78$ g/mL. If the gradient contains all three matching densities, which ordering of equilibrium bands from top to bottom is expected?

1.78,\ 1.70,\ 1.65

1.65,\ 1.70,\ 1.78

✅

1.70,\ 1.65,\ 1.78

D.All three must occupy the same position

💡 Difficulty: easy | ✅ Correct: B

📖 Explanation: Because the gradient becomes progressively denser toward the bottom, the least dense DNA reaches equilibrium in the upper region, the intermediate-density DNA occupies a middle position, and the densest DNA forms the lowest band.

Q10. A sample initially contains DNA at several densities. During centrifugation, the bands become sharper and then stop changing position even though the rotor continues operating. Which explanation best integrates the observations?

A.The DNA has reached density-dependent equilibrium positions within the gradient ✅

B.The DNA has been completely removed from the tube

C.The rotor has stopped generating a centrifugal field

D.All DNA molecules have acquired the same buoyant density

💡 Difficulty: easy | ✅ Correct: A

📖 Explanation: Initially, particles migrate through the gradient, causing bands to develop and sharpen. Once each population reaches the region where its buoyant density matches the surrounding medium, net migration approaches zero, so continued centrifugation does not continually move the bands.

Q11. A scientist wants to separate two DNA populations whose buoyant densities differ by only $0.01$ g/mL. The scientist can either improve the density-gradient resolution or simply double the DNA concentration. Which choice is more directly useful for distinguishing the populations?

A.Improve gradient resolution because the separation depends on resolving different equilibrium densities ✅

B.Double DNA concentration because concentration determines equilibrium density

C.Increase molecular mass because it automatically creates a larger density difference

D.Reduce the gradient range until both DNA populations have identical densities

💡 Difficulty: easy | ✅ Correct: A

📖 Explanation: When the density difference is small, the central challenge is resolving closely spaced equilibrium positions. Improving gradient resolution can make small density differences easier to distinguish, whereas increasing sample concentration does not inherently increase the density difference between populations.

🔗 Related Topics (MCQs)

📝 Cellular Foundations MCQs 📝 Cell structure and functional units MCQs 📝 Cell dimensions and oxygen diffusion MCQs 📝 Archaea (archaebacteria) MCQs 📝 Bacteria (eubacteria) MCQs

←Previous Topic:Differential Centrifugation and Cell Fractionation📝 isopycnic centrifugationOrganelle isolation by cell fractionation:Next Topic→