Flow Cytometry Flashcards
Argon ___ electricity heats the argon gas producing light of a single wavelength.This emitted light excites the fluorescent compounds attached to the antibodies. Fluorescent compounds emit light of a higher wavelength which is detected by the instrument.
Function of laser
Function of Photomulitplier Tube (PMT)
Detects photons. The more photons, the more fluorescence being emitted.
Function of Computer display.
Electrical impulses are sent to the computer for storage, analysis, and viewing as histograms
Explain hydrodynamic focusing.
Technique that allows cells to pass through the laser beam in a single file. one cell at a time.
Identify the cellular component measured by forward angle light scatter (FALS or FS)
Detects particle (cell) size
Identify the cellular component measured by 90 degree light scatter (SS)
Detects the granularity or internal composition of cells. Measured by a separate PMT
Be able to identify the cell populations of normal scattergrams
Pictures
List the five fluorescent compounds commonly used with an argon laser in flow cytometric analysis.
- FITC-Fluorescein isothiocyanate-green
- PE-phycoerythrin-orange
- ECD-Red
- PeCy5-Deep red
- PeCy7
- PI-propidium iodide-red
List three different types of samples that are acceptable for flow cytometric analysis
Peripheral blood, BM, tissue, fine needle aspirate, lavage specimen
List two methods of preparing a single cell suspension from a peripheral blood sample.
- Sample is mixed with a hypotonic solution to lyse RBCs, and centrifuged, incubated for 30 mins, and run on flow cytometer
- automated instrument: T-Q prep
Define gating
Electronic isolation of a population of cells for analysis.
List three reasons why immunophenotypic by flow cytometry might be requested on a patient.
- Diagnosis of leukemias and lymphomas
- Detection of minimal residual disease before the overt relapse of acute leukemia
- T-cells subset enumeration in HIV patients
Define helper: suppressor (H:S) ratio
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State the adult normal range for Helper:suppressor ratio
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Define CD and explain the purpose of CD numbers
CD: Cluster of differentiations; cell surface membrane receptors or markers used to characterize cells by their functions. used to identify cell clones associated with lymphatic and myelogenous leukemias & lymphomas
List CD markers for granulocytic/monocytic
CD13, CD14, CD15, CD33, CD34
List CD markers for Erythroid
CD71
List CD markers for megakaryocytes
CD41, CD42, CD61
List CD markers for B cell
CD19, CD20, Kappa and lambda light chains
List CD markers for T cell
CD3, CD4, CD5, CD7, CD8
What is normal myeloid:erythroid ratio (M:E)?
1.5:1-3.3:1
Calculate the cellularity in an adult using the normacellular formula and determine if it is normal, hyperplasia,or hypoplasia.
(100-Age) +/- 10
Review Cytochemistry lecture for case studies
MPO ect
Calculate an LAP score and normal range
Score times # cells. Then add up all to get LAP score
Normal range: 20-100
