Presentation is loading. Please wait.

Presentation is loading. Please wait.

New Research Directions in the Rowlen Group Detection of cancer cells in blood Isolation of cancer cells using dielectrophoresis Multispectral analysis.

Similar presentations


Presentation on theme: "New Research Directions in the Rowlen Group Detection of cancer cells in blood Isolation of cancer cells using dielectrophoresis Multispectral analysis."— Presentation transcript:

1 New Research Directions in the Rowlen Group Detection of cancer cells in blood Isolation of cancer cells using dielectrophoresis Multispectral analysis of single cells for identification

2 “One out of every three Americans will be diagnosed with cancer” Early diagnosis saves lives A 1 g tumor, generally undetectable, can exfoliate up to 10 6 cells per day into blood stream. However, 10 9 – cells/mL in blood Detect 4 in 10 9 ? If malignant cells could be distinguished/detected: Early diagnosis Monitor response to treatment Evaluate minimum residual disease

3 Fact: Most cancer cells have a lower membrane potential than most normal cells Hypothesis: membrane potential can be used as a biomarker for malignant cells in blood

4 Cellular Membrane Potential Na + Cl - K + Cl - Cl - Na + Cl - K + Cl - Na + Normal cell: -40 to -90 mV Malignant cell: -10 to – 30 mV

5 Resting Membrane Potential estimated from Goldman Eqn: Can be measured using “distributional” probes:

6 Bis (1,3-dibutyl-barbituric acid) trimethine oxonol [DiBAC 4 (3)] Ideal characteristics of a fluorescent dye sensitive to cytoplasmic membrane potential: 1) response proportional to membrane potential 2) increased fluorescence with decreased polarization 3) photostable, 4) minimal toxicity to cell 5) immune to drug efflux pump

7 Assay: Stain cells, flow cytometry detection of rare events Fluorescence Intensity Cell Count

8 Objective: “… to develop novel technologies for capturing, enriching, and preserving exfoliated abnormal cells in body fluids or effusions and to develop methods for concentrating the enriched cells for biomarker studies.” “… the number of exfoliated tumor cells [in body fluids] is often small compared to the number of non-neoplastic cells. Therefore, the detection of exfoliated abnormal cells by routine cytopathology is often limited because few atypical cells may be present in the specimen.…” “Thus, the development of enrichment methods is a prerequisite for the routine detection of small numbers of exfoliated cells and small amounts of subcellular materials in biological fluids for molecular analysis.” National Institutes of Health Program Announcement

9 Dielectrophoresis Force on particle due to interaction between induced dipole and local electric field If particle more polarisable than medium, positive dielectrophoresis (as shown) If particle less polarisable than medium, negative dielectrophoresis, dipole aligns counter to field, repelled by field

10 Dielectrophoretic Field Cages (all field cage images taken from literature)

11 Electric Field in Cage

12 Dielectrophoretic Field Cages 200  m Micro-Channel Incoming Cells Single Cell Trapped in Cage Picture courtesy of Evotec OAI

13

14

15

16 Medical diagnostics Biocomplexity On-line absorbance, fluorescence, and Raman Multispectral Analysis for Rapid Identification of Single Cells Technology (e.g., semiconductor industry)

17 Single Nanoparticle Enumerator “SNaPE” 3D Stage Ar + Laser PMT PC Computer Syringe Pump Premonochromator 5X Beam Expander With 50  m Pinhole Mirror Lens (f = 150 mm) Longpass Filter 500  m Pinhole Notch Filter Dichroic Mirror 100X Objective 10  m I.D. Capillary

18 Bacterial Morphology SFM image of E. coliSEM image of P. aeruginosa Images adapted from:

19 Bacterial Fluorescence

20 Raman Intensity Raman Shift (cm-1) K562 Jurkat Confocal Raman spectra of Cancer Cells: (glass substrate subtracted)

21 Bioinformatics Raman Extinction Fluorescence Scatter Raman Shift (cm -1 ) Wavelength (nm)

22 Thanks to the Rowlen Group Matt Michele Jessica Carrie Michael Not pictured: Peter and Jenna


Download ppt "New Research Directions in the Rowlen Group Detection of cancer cells in blood Isolation of cancer cells using dielectrophoresis Multispectral analysis."

Similar presentations


Ads by Google