SCYM Sample Preparation & Staining Techniques 5

Question 1

When staining cells for apoptosis using Annexin V/PI, what is the interpretation of Annexin V+/PI- cells?

Accepted Answer

Early apoptotic cells with intact membranes

Answer

Annexin V+/PI- represents early apoptosis: phosphatidylserine is externalized (binding Annexin V) but the plasma membrane remains intact (excluding PI).

Question 2

Which condition is essential when performing Annexin V staining to ensure proper binding?

Accepted Answer

Calcium-containing binding buffer

Answer

Annexin V binding to phosphatidylserine is calcium-dependent; the staining buffer must contain physiological calcium concentrations (typically 2.5 mM CaCl₂).

Question 3

A researcher wants to measure proliferation of T cells using CFSE. Which statement correctly describes the assay principle?

Accepted Answer

CFSE covalently labels proteins, and fluorescence halves with each cell division

Answer

CFSE covalently binds intracellular proteins; at each division, the dye is distributed equally between daughter cells, so fluorescence intensity halves with each generation.

Question 4

What is the recommended storage condition for cells after paraformaldehyde fixation when same-day acquisition is not possible?

Accepted Answer

4°C protected from light for up to 24-72 hours

Answer

Fixed samples are stable at 4°C in the dark for 24-72 hours; longer storage can cause autofluorescence increases and epitope degradation.

Question 5

Which pre-analytical variable most significantly affects the absolute count of CD34+ hematopoietic progenitor cells in peripheral blood?

Accepted Answer

Sample temperature and time from collection to processing

Answer

CD34+ cells are particularly sensitive to delayed processing and temperature extremes; cells begin to die and expression levels change within a few hours if samples are not handled properly.

(SCYM) Specialist in Cytometry Practice Test

(SCYM) Specialist in Cytometry Practice Test

SCYM Sample Preparation & Staining Techniques 5