Applications of Flow cytometry Flashcards
Name some ways that Flow Cytometry can be used in research
- Immunophenotyping
- Cell sorting
- Cell Cycle Analysis
- Apoptosis
- Cell Proliferation
- Intracellular calcium flux
Name some ways that flow cytometry can be used in a clinical setting
- Transplantation
- Transfusion
- Immunophenotyping
- DNA analysis
- Clinical immunology
- Testing for cancers or blood disorders
- Testing immune function
Describe the sets of cytoplasmic flow cytometry
- Treat cells with a drug or stimulant
- Fix and permeabilize cells
- Stain cells with antibodies
- Perform flow cytometry
What are the 3 types of hematopoietic neoplasia (blood cancer)
Leukemia
Lymphoma
Myeloma
What does the following stand for …
ALL
AML
CLL
CML
CMML
Acute lymphocytic leukemia
Acute myeloid leukemia
chromic lymphocytic leukemia
chronic myeloid leukemia
chronic myelomonocytic leukemia
What are the 2 types of lymphoma
Hodgkin’s lymphoma
Non-Hodgkin’s lymphoma
What is the measurable residual disease test and what complementary approaches are used
- Detection of rare neoplastic cells during post treatment follow up using…
1. Multiparametric flow cytometry - immunophenotyping analysis to detect abnormal expression of certain antigens
2. Molecular diagnostics (PCR) - to detect specific DNA signatures
Check notes for information on cell cycle and drug testing
What are the 3 ways you can measure apoptosis?
- SS, apoptotic cells have a high SS due to the membrane blebbing that occurs
- Annexin - when a cell is undergoing apoptosis the phosphatidyl serine (PS) that should be on the inner membrane flips to the outer membrane, annexin binds to PS and can then be measured. This happens in early apoptosis.
- PI - In late apoptosis the membrane begins to break apart allowing PI in which will attach to the nucleus and can also be measured.
Describe the application of flow cytometry in DNA analysis
- Malignant cells often have an abnormal amount of chromosomes and this is significant in prognosis.
- The DNA content of a tumor may be expressed as a DNA index - the ratio of cancer cell content vs normal cell content
How can cell proliferation be measured in flow cytometry
As cells divide the fluorescence in the daughter cells will be exactly half of the fluorescence of the parent cells etc…
What are the ways flow cytometry can be used in transplantation?
- HLA antibody detection
- Flow cross matching of organs
- HLA typing
- CD34 analysis of hematopoietic stem cell transplantation
What is the significance of HLA antibody detection in transplantation?
- Utilises beads coated in HLA
- Can be done on both FACS or Luminex
- Complement fixing antibody - Hyperacute rejection
- Non-complement fixing antibody - early rejection, long term survival may be achieved
- IgG against HLA Class I and II show poor prognosis
Describe CD34 analysis of HSC
- CD34 is a marker of stem cells
- CD34is present on 2-4% of all normal mononuclear cells
- HSC can be mobilised in blood by granulocyte-colony stimulating factor (GM-CSF)
- Flow methods utilize a mix of anti CD34, anti CD45 and fluorospheres to generate an absolute count.
What is the equation for CD34 absolute count
Number of CD34 cells + fluorospheres / Number of fluorospheres counted
Multiplies by dilution factor
