Research Methodology & Design Flashcards

In a clinical PCR validation study, which design best estimates diagnostic sensitivity and specificity?

Answer: Testing positive and negative samples against an accepted reference method

Sensitivity and specificity require comparison of both positive and negative samples to a reference standard.

An operator who knows sample group identity interprets ambiguous melt curves. Which bias is most likely introduced?

Answer: Observer bias

Knowing group assignment can unconsciously influence subjective interpretations; blinding prevents this.

Which experiment establishes the limit of detection (LoD) of a PCR assay?

Answer: Replicate testing of low-concentration dilutions to find the level detected at least 95% of the time

LoD is commonly defined as the lowest concentration detected in ≥95% of replicates.

A standard curve slope of -3.32 corresponds to approximately what amplification efficiency?

Answer: 100%

Efficiency = 10^(-1/slope) - 1, and a slope of -3.32 gives about 100%.

Why should sample processing order be randomized across treatment groups?

Answer: To prevent time-dependent drift from being confounded with treatment

Randomization distributes unknown systematic effects evenly across groups.

Which statement correctly describes the 2^-ΔΔCq method's key assumption?

Answer: The target and reference genes have similar, near-100% amplification efficiencies

The ΔΔCq method assumes approximately equal and optimal efficiencies; otherwise the Pfaffl method is preferred.

Running the same sample on different days, by different operators, primarily assesses which performance characteristic?

Answer: Reproducibility (intermediate precision)

Variation across days and operators assesses intermediate precision/reproducibility rather than within-run repeatability.