Chromatography Techniques Flashcards
7 cards from real BACE practice questions. Tap to flip, then mark Knew It or Still Learning — missed cards come back until you master them.
Read the first 7 Chromatography Techniques flashcards as text
What is a chromatogram?
Answer: A plot of detector signal versus elution volume or time
A chromatogram is a graphical output showing detector signal (e.g., UV absorbance) on the Y-axis versus elution volume or time on the X-axis, with peaks representing separated components.
Which of the following best describes 'gradient elution' in ion exchange chromatography?
Answer: Progressively increasing salt concentration in the elution buffer
Gradient elution involves gradually increasing the salt concentration (or changing pH) to sequentially release proteins with different affinities for the resin.
What is the function of a guard column in HPLC?
Answer: To protect the analytical column from particulates and contaminants
A guard column is a short, inexpensive column placed before the analytical column to trap particulates and strongly retained contaminants, extending analytical column life.
In thin layer chromatography (TLC), the Rf value is calculated as:
Answer: Distance traveled by spot / distance traveled by solvent
Rf (retention factor) = distance traveled by the analyte spot ÷ distance traveled by the solvent front, and ranges from 0 to 1.
When performing protein purification by chromatography, SDS-PAGE is typically run on collected fractions in order to:
Answer: Confirm the presence and purity of the target protein
SDS-PAGE separates proteins by size and visualizes bands, allowing the technician to confirm whether the target protein is present and to assess purity.
Which buffer property is most critical to control when performing ion exchange chromatography?
Answer: pH and ionic strength
pH determines the charge state of both the protein and the resin, while ionic strength affects binding strength; both must be tightly controlled for reproducible separations.
After an affinity chromatography purification step, the column is often regenerated by:
Answer: Flushing with a regeneration solution to remove residual bound material, then re-equilibrating
Regeneration involves washing away any remaining bound material with an appropriate stripping solution, followed by re-equilibration in storage or binding buffer to restore column capacity.