SCYM Data Analysis 2

Question 1

In flow cytometry, what does the term 'compensation' correct for?

Accepted Answer

Spectral spillover of fluorochromes into adjacent channels

Answer

Compensation mathematically removes spectral spillover, where emission from one fluorochrome bleeds into a detector channel assigned to another fluorochrome.

Question 2

Which gating strategy is most appropriate for identifying rare cell populations comprising less than 0.01% of total events?

Accepted Answer

Sequential hierarchical gating with back-gating validation

Answer

Sequential hierarchical gating progressively enriches the population of interest, and back-gating confirms the identified population is correctly placed in earlier scatter plots.

Question 3

What is the purpose of using a 'fluorescence minus one' (FMO) control?

Accepted Answer

To determine the boundary of positivity in the presence of spectral spillover

Answer

FMO controls contain all fluorochromes except the one being measured, revealing where the negative population boundary falls after compensation is applied.

Question 4

When analyzing a bivariate dot plot, a 'cloud' of events forming a diagonal line from low-low to high-high typically indicates:

Accepted Answer

A positive correlation between the two measured parameters

Answer

A diagonal cluster from low-low to high-high in a bivariate plot indicates that the two parameters co-vary positively, suggesting co-expression or biological co-regulation.

Question 5

In mass cytometry (CyTOF), which approach replaces traditional spectral compensation?

Accepted Answer

Minimal signal spillover due to narrow isotope mass windows

Answer

Mass cytometry uses heavy metal isotope tags with minimal mass overlap, so the need for spectral compensation is largely eliminated compared to fluorescence-based cytometry.

(SCYM) Specialist in Cytometry Practice Test

(SCYM) Specialist in Cytometry Practice Test

SCYM Data Analysis 2