LST Proliferation Assays 2

Question 1

In CFSE dilution assays analyzed by flow cytometry, each successive generation of dividing lymphocytes is characterized by:

Accepted Answer

Fluorescence intensity halved relative to the previous generation

Answer

Because CFSE is equally partitioned between two daughter cells at each division, each generation displays fluorescence that is exactly half that of the parent generation, creating discrete peaks on a histogram.

Question 2

The Proliferation Index (PI) in a CFSE assay differs from the Division Index in that PI measures:

Accepted Answer

The average number of divisions undergone by cells that actually divided

Answer

The Proliferation Index is the average number of divisions per dividing cell (excludes undivided cells from the calculation), whereas the Division Index includes undivided cells in the denominator.

Question 3

Which cell density is generally optimal for seeding peripheral blood mononuclear cells (PBMCs) in a standard 96-well proliferation assay?

Accepted Answer

1–2 × 10⁵ cells per well

Answer

A seeding density of 1–2 × 10⁵ PBMCs per well in a 96-well plate provides adequate cell-to-cell contact for co-stimulation while avoiding overcrowding that suppresses proliferation or depletes nutrients.

Question 4

In a tritiated thymidine proliferation assay, cells are typically harvested onto glass fiber filters using a:

Accepted Answer

Semi-automated cell harvester

Answer

A semi-automated cell harvester lyses cells and deposits their DNA onto glass fiber filter mats, efficiently washing away unincorporated ³H-thymidine before scintillation counting.

Question 5

EdU (5-ethynyl-2'-deoxyuridine) offers a technical advantage over BrdU for proliferation assays primarily because:

Accepted Answer

EdU detection does not require harsh DNA denaturation steps that damage cell surface antigens

Answer

EdU is detected via a click chemistry reaction that does not require the acid or heat denaturation needed to expose BrdU epitopes, thereby preserving cell surface markers and enabling robust multi-parameter flow cytometry.

LST - Lymphocyte Stimulation Test Practice Test

LST - Lymphocyte Stimulation Test Practice Test

LST Proliferation Assays 2