SCYM Sample Preparation & Staining Techniques 2

Question 1

Which fixative is most commonly used to preserve cell surface antigens for flow cytometric immunophenotyping?

Accepted Answer

Paraformaldehyde 1-4%

Answer

Paraformaldehyde (1-4%) is preferred for surface antigen preservation because it cross-links proteins while maintaining epitope integrity better than stronger fixatives.

Question 2

When preparing peripheral blood for flow cytometry, which lysis method is preferred for maintaining lymphocyte scatter characteristics?

Accepted Answer

Ammonium chloride-based red cell lysis

Answer

Ammonium chloride-based lysis selectively lyses red blood cells while preserving lymphocyte morphology and scatter properties.

Question 3

What is the primary purpose of blocking with normal serum prior to antibody staining in flow cytometry?

Accepted Answer

To saturate Fc receptors and reduce non-specific antibody binding

Answer

Blocking with normal serum saturates Fc receptors on monocytes and other cells, preventing non-specific antibody binding that would produce false-positive results.

Question 4

Which parameter should be used to gate out dead cells when no viability dye is available?

Accepted Answer

Low forward scatter and high side scatter

Answer

Dead cells characteristically show decreased FSC (cell shrinkage) and increased SSC (increased granularity from cellular debris), allowing them to be excluded by scatter gating.

Question 5

When performing intracellular cytokine staining, why is stimulation with brefeldin A or monensin required?

Accepted Answer

To block protein transport and accumulate cytokines inside cells

Answer

Brefeldin A and monensin block the Golgi apparatus, preventing secretion so that newly synthesized cytokines accumulate intracellularly and can be detected.

(SCYM) Specialist in Cytometry Practice Test

(SCYM) Specialist in Cytometry Practice Test

SCYM Sample Preparation & Staining Techniques 2