Data Acquisition & Analysis Flashcards
7 cards from real SCYM practice questions. Tap to flip, then mark Knew It or Still Learning — missed cards come back until you master them.
Read the first 7 Data Acquisition & Analysis flashcards as text
What is the primary purpose of using a threshold (discriminator) during flow cytometry data acquisition?
Answer: To exclude events below a set signal level from data collection
A threshold eliminates low-level electronic noise and debris by only collecting events whose signal exceeds the defined minimum value.
When setting up a multicolor panel, the process of measuring how much fluorochrome signal appears in unintended detector channels is called:
Answer: Spectral overlap
Spectral overlap occurs because fluorochrome emission spectra are broad and extend into adjacent detector channels.
Which compensation control provides the most accurate matrix for correcting spectral spillover?
Answer: Single-stained beads or cells at the same brightness as the experimental sample
Compensation controls must match the brightness of the actual sample; single-stained controls at equivalent intensity yield the correct spillover coefficients.
In flow cytometry, what does the term 'coincidence' (or 'doublet') refer to during acquisition?
Answer: Two or more cells passing through the interrogation point at the same time
Coincidence events occur when multiple cells pass through the laser beam together, creating artificially large or complex signals.
A histogram displaying a broad, flat peak for a single fluorescent population most likely indicates:
Answer: High coefficient of variation (CV) suggesting instrument or staining problems
A high CV (wide peak) reflects poor resolution, often caused by suboptimal PMT voltage, aggregated cells, or inconsistent staining.
Fluorescence Minus One (FMO) controls are used in flow cytometry primarily to:
Answer: Define positive/negative gates in the presence of spillover from other fluorochromes
FMO controls contain all fluorochromes except the one being gated, revealing where the negative population sits after spillover fills the channel.
During acquisition, what is the recommended approach when a sample shows an unexpected shift in bead position on quality control plots?
Answer: Stop acquisition, investigate instrument alignment and laser stability, then recalibrate
A QC bead shift indicates an instrument performance change; data collected during that period may be unreliable and the cause must be identified before continuing.