FACS lecture
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49 terms
Terms | Definitions |
|---|---|
 Definition of flow cytometry/FACS | Measuring properties of cells in flow. |
| Flow sorting definition | Sorting/separating cells baserd on properties measured in flow. Can be used for optical sorting of any particles. |
| Basics of Immunophenotyping | Detection of antigenic pattern (immune penotype- antigenic properties of cells/subcellular particles-mitochondria) of cells allowing to assign to one of cell lineages. Characterize maturity and functional properties of cells, determine heterogenicity of tested cells, aberrant features of cells. Labelling of cells with fluorescent antibodies against various antigenic markers followed by either luminescent microscopy of flow cytometry (microfluorometery of labeled cells/specific proteins. Normal/pathogenic? Flowing/active? |
| Optical properties of cells | Refraction of light (forward scatter), reflection of light (side scatter). Measuring how many times does light change directions because of change in optical density. |
| What is measured by side scatter? | Granularity and complexity or cells. More-neutrophils and immature myeloid cells. |
| What is measured in forward scatter? | Mainly size of cells. More=Monocytes, immature myeloid cells. |
| 2D Histogram | Measures forward scatter on X-axis and side scatter on Y-axis. Close to zero: lymphocytes: small and simple. |
| Monoclonal antibody production | Immunization of lab rate--> antibody forming cells-->fuse with tumor cells from tissue culture-->hybridoma-->seeding of cells in a medium allowing selection of hybrid cells-->hybridomas screened for Ab production, Ab producing hybridomas cloned-->monoclonal antibodies isolated for cultivation. Labeled with fluorescence for specific Ab. |
| 2D histograms in leukemia | Can see different singular proportions of myeloid and lymphocytes. |
| Cluster of differentiation | Enormous diversity of monoclonal antibodies to leukocytes. Most used to characterize surface of leukocytes. 25 CD numbers at first, now more than 350. |
| CD Antigenic properties of stem cells | CD34 |
| Lymphoid CD markers | CD10, CD34, CD38 |
| Myelomonocytic progenitor CD markers | CD13, CD33, CD34 |
| Erythroid CD antigen | CD36 |
| Eosinophil CD antigen | CD34 |
| Basophil CD Antigen | CD33 |
| Megakaryocytic CD antigens | CD41, CD61 |
| Immature T-cell CD Markers | CD34, CD7 |
| Immature B cell CD marker | CD34, CD79a, CD10 |
| NK cell CD markers | CD2, CD8, CD56, CD16 |
| Pro-T-cell CD markers | cyTdT, CD7 |
| Early cortical T-cells CD markers | cyTdT, CD7, cyCD3 |
| Late cortical T cells | CD2, CD7, CD4, CD8 |
| Medullary T-cells | CD2, CD7, sCD3 |
| T helper CD markers | CD2, CD3, CD4 |
| T suppressor CD markers | CD2, CD3, CD8 |
| T killer CD markers | CD2, CD3, CD8, CD57 |
| Pro B-cells CD markers | CD34, TdT, CD79a, CD19, CD10 |
| Pre-B cells | CD79a, CD19, CD10 |
| Immature B-cells CD markers | CD79a, CD19, CD10, CD20 |
| Mature B-cells CD markers | CD79a, CD19, CD20, CD45 |
| Lineage specific antigens with broad expression | Pan myeloid antigens: CD13, CD33, MPO. Pan-B-cell antigen: CD22, CD19, cyCD79a. Pan-T-cell antigens: cyCD3, CD2, CD7, CD5 |
| Antigens associated with immaturity of cells | Immaturity: cyTdT, CD7, CD34, HLA-DR, CD117 |
| Antigens associated with activation of cells | CD23, CD25, CD38, HLA-DR |
| Antigens with lineage specific and maturation dependent expression: | Myeloid cells: CD11b, CD15, CD16, CD64. Monocytoid: CD14, CD64. Platelets: CD41, CD42, CD61, CD62. Erythroid: CD36, CD71, GlyA. B-cells: Cd5, CD10, CD20, CD21, CD79b, FMC7, Ig. T-cells: CD1/1a, sCD3, CD4, CD8. NK cells: CD16, CD56, CD57. |
| Markers of cell clonality | Kappa and delta chains of IG: TCR alpha/beta and TCR gamma/delta. |
| B-ALL Common type CD markers | cyCD79a, CD10, CD19, CD20, CD22, CD34, cyTdt |
| B-All Mature type CD markers | cyCD79a, CD10, CD19, CD20, CD22, CD45, sIg |
| T-ALL Mature type CD markers | CD2, sCD3, CD4 or CD8, CD7, CD45? |
| AML M0-M1 CD markers | CD7, CD13, CD33, CD34, CD45, CD117, HLA-DR |
| AML M3 CD markers | SS high, CD13, CD33, CD45, CD117, cyMPO, no CD4, NO HLA-DR (unique) |
| AML M4 CD markers | CD34, CD33, CD45, CD117, CD14, CD11b, HLA-DR, cyMPO, maybe CD4, CD36, CD64 |
| AML M6 CD markers | CD71, glycophorin A (CD235a-lymphoblastic) |
| AML M7 (megakaryocyte) CD markers | CD41, CD61 |
| Antigenic markers of Myeloid precursor cells | CD33, CD34, CD38 |
| BCLL CD markers | CD5, CD19, CD23, sIg diminished (monoclonal) |
| CD markers of T-large granular cell leukemia | CD3, CD2 |
| CD markers of NK large granular cell leukemia | CD2, CD56 |
| Markers of Hematological Precursor malignancies | TDT in immature lymphoids, surface vs cytoplasmic: mature (lymphoproliferative over lymphoma), HLADR in B cells |
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