Download
slide1 n.
Skip this Video
Loading SlideShow in 5 Seconds..
Advances in Cytometry PowerPoint Presentation
Download Presentation
Advances in Cytometry

Advances in Cytometry

138 Views Download Presentation
Download Presentation

Advances in Cytometry

- - - - - - - - - - - - - - - - - - - - - - - - - - - E N D - - - - - - - - - - - - - - - - - - - - - - - - - - -
Presentation Transcript

  1. Advances in Cytometry

  2. TheEvolution of Cytometry: FromCytologytoCytomics Cytology Cytometry Cytomics

  3. FromCytometrytoCytomics: BetterTechnologyfor a BetterBiology BetterTechnology BetterBiology More information Better performance Novel strategies Novel Instruments New applications Better data mining

  4. Betteror Novel CytomicInstrumentation HydrodynamicFlowCytometry ConfocalMicroscopy AcousticFlowCytometry MassSpectrometry Cytometry Laser Scanning Cytometry Multispectral Image-in-flowCytometry HCA AutomatedBioimaging

  5. Betterinstrument performance • Acceleratingtherate of cell data acquisition • Acceleratingtherate of specimenanalysis High-TroughputCytometry

  6. Increasingcellacquisitionrate: Acousticfocusing in flowcytometry • Acoustic focusing cytometry uses ultrasound waves (over 2 MHz) to position cells into a single focused line along the central axis of a flow channel without high velocity or high volumetric sheath fluid. • The acoustic focusing actually concentrates the cells in the center of the fluid with sound energy. This means considerable flexibility in the sample concentration. The end result is more accurate and precise data. • Acoustic focusing separates the alignment of cells from the particle flow rate. This allows to increase or decrease flow without disrupting the focus of cells in the capillary.

  7. MassCytometry: Overcomingtheproblems of fluorescence

  8. Lanthanides as AdvantageousTagsforBiologicalMeasurements

  9. Lanthanides as AdvantageousTagsforBiologicalMeasurements • As defined by IUPAC, rare earth elements or rare earth metals are a set of seventeen chemical elements in the periodic table, specifically the fifteen lanthanides plus scandium and yttrium. • Scandium and yttrium are considered rare earth elements since they tend to occur in the same ore deposits as the lanthanides and exhibit similar chemical properties.

  10. Lanthanides as AdvantageousTagsforBiologicalMeasurements

  11. Lanthanides as AdvantageousTagsforBiologicalMeasurements • A time-of-flight mass spectrometer (TOF-Ms) uses the differences in transit time through a drift region to separate ions of different masses. • The ions from the ion source are accelerated by an electric field pulse. • The accelerated particles then pass into a field-free drift tube that is about a meter in length. • In TOF-Ms all ions are accelerated to the same kinetic energy. • Their velocities are inversely proportional to the square roots of their masses. • The lighter ions arrive at the detector earlier than the heavier ions, of lower velocity.

  12. TheMassFlowCytometer (CyTOF): A new multiparametricdimension Journal of Immunological Methods 361 (2010) 1–20

  13. TheMassFlowCytometer (CyTOF): A new multiparametricdimension http://www.DVSsciences.com

  14. TheMassFlowCytometer (CyTOF): A new multiparametricdimension

  15. TheMassFlowCytometer (CyTOF): A new multiparametricdimension

  16. TheMassFlowCytometer (CyTOF): A new multiparametricdimension

  17. Workflow in MassCytometry

  18. DesigningAffinityReagentsforMassCytometry

  19. DesigningAffinityReagentsforMassCytometry

  20. CustomLabelling of AntibodiesforMassCytometry MAXPAR® labeling reagent kits for 33 different metals are available from DVS Sciences Inc. for preparation of labeled antibodies or intracellular proteins. Kits are optimized and quality control tested for use with the CyTOF® instrument and are available in pack sizes sufficient to label 100ug or 1mg of antibody. Labeling methods are simple and convenient and require only 2.5 hours to complete. The labeling kits utilize a traditional approach of adding a metal tag using disulfide reduction (typically in the Fc region of the antibody) through a maleimide linker. Labeled antibodies are stable for approximately 6 months after preparation.

  21. Performance of MassCytometry

  22. MassCytometry and FlowCytometryyield similar results

  23. MassCytometry and FlowCytometryyield similar results Humancelllines Human AML cases Human normal PBMC

  24. Analysis of IntracellularAntigensbyMassCytometry

  25. Analysis of CellViabilitybyMassCytometry

  26. Bead-basedAssaysbyMassCytometry

  27. New Tags in theCatalog!

  28. New Tags in theCatalog!

  29. FutureTags in theCatalog: FromIsotypestoIsotopes??

  30. When the Map approaches the 1:1 scale…or beyond!

  31. Combining phenotypic Maps with Activation Landscapes by CyTOF

  32. A novel bioinformatic description of subpopulations in Hematopoesis

  33. Cell signaling across subpopulations in Hematopoesis

  34. Cell signaling modulation across subpopulations in Hematopoesis

  35. Gracias porvuestraatención… jose.e.oconnor@uv.es http://www.uv.es/~oconnor/MASTER_UPV_2012