SCYM Data Acquisition & Analysis 3

Question 1

In a bivariate dot plot of FSC-A vs. FSC-H, a population appearing above the diagonal line (FSC-H lower than expected for its FSC-A) most likely represents:

Accepted Answer

Doublets or aggregates

Answer

Doublets have greater area (FSC-A) relative to height (FSC-H) because two cells passing together produce a widened pulse without proportionally increased peak height.

Question 2

Which gating strategy step should immediately follow excluding debris with an FSC/SSC gate in a standard immunophenotyping panel?

Accepted Answer

Doublet exclusion using pulse geometry

Answer

After the initial scatter gate removes debris, doublet exclusion ensures only single cells proceed to fluorescence analysis.

Question 3

The 'coefficient of variation' (CV) of a flow cytometry peak is calculated as:

Accepted Answer

Standard deviation divided by mean × 100

Answer

CV = (SD / Mean) × 100; a lower CV indicates a tighter, better-resolved population.

Question 4

When analyzing rare events (e.g., circulating tumor cells at 1 per million), what is the most critical acquisition parameter to adjust?

Accepted Answer

Total events collected—acquire far more events to detect rare populations

Answer

Statistical reliability for rare populations requires collecting a very large number of total events; at 1 per million, millions of cells must be acquired for meaningful counts.

Question 5

Which file format is the standard for storing flow cytometry list-mode data?

Accepted Answer

FCS (Flow Cytometry Standard)

Answer

FCS (e.g., FCS 3.1) is the ISAC-approved standard format encoding all event parameters and metadata for flow cytometry data.

(SCYM) Specialist in Cytometry Practice Test

(SCYM) Specialist in Cytometry Practice Test

SCYM Data Acquisition & Analysis 3