1 / 23

CARS Microscopy Third Annual Workshop

Introduction. I attended the 3rd Annual CARS Workshop in Harvard in June 2003Here are some of the basic ideas of the technique.. Workshop. Sunney Xie: ChemistCame to Harvard in 1999Pioneered CARS developmentsThird annual CARS workshopPhysicists, Biologists, Industry peopleLecturesLab prac

addison
Download Presentation

CARS Microscopy Third Annual Workshop

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

    1. CARS Microscopy Third Annual Workshop

    2. Introduction I attended the 3rd Annual CARS Workshop in Harvard in June 2003 Here are some of the basic ideas of the technique………….

    3. Workshop Sunney Xie: Chemist Came to Harvard in 1999 Pioneered CARS developments Third annual CARS workshop Physicists, Biologists, Industry people Lectures Lab practicals

    4. Outline What is CARS? Why use it? Non Resonant Background Sources Detection schemes Applications Problems? Discussion

    5. What is CARS? Non linear Raman process Generated at the focus of the beam ?AS = 2?p - ?s

    7. Why use CARS? Intrinsic vibrational contrast Strong, directional signal => Sensitive Requires moderate average powers good for biological samples Only generated at focus => 3D sectioning capability Higher in frequency than one-photon fluorescence => easily detected in presence of a strong fluorescent background. Near IR => little scattering, deep penetration in tissues Near IR => little absorption, Low photodamage

    8. Non-resonant Background Non-resonant Background from the bulk!

    9. Two colours Difference between pump and Stokes span the vibrational spectrum Modelocked sources Picosecond pulses Time-bandwidth product Tunability for exciting different vibrations OPO pumped by ps Laser

    10. Detection Schemes F – CARS: Forward detected E – CARS: Epi detected P- CARS: Polarisation dependent FM CARS: Frequency modulated

    11. F CARS

    12. F CARS

    13. Detection Schemes F – CARS: Forward detected E – CARS: Epi detected P- CARS: Polarisation dependent FM CARS: Frequency modulated

    14. Avoid non resonant background! Objects smaller than ?p/3 (too short for destuctive interference) Interfaces with different ?3 - to propagation Back-scattering of F CARS, reflactive/turbid media i.e. tissue

    15. E CARS Images of a hairless mouse ear. The Raman shift is set at 2,845 cm1 (p 816.8 nm) to address the lipid CH2 symmetric stretch vibration. The frames are averaged for 2 s. (A) Stratum corneum with bright signals from the lamellar lipid intercellular space that surrounds the polygonal corneocytes. Bright punctuated dots are ducts of sebaceous glands. (B) Sebaceous glands at 30 m from skin surface. (C) Individual cells of the gland compartment can be recognized, with nuclei visible as dark holes (arrow). (D) Adipocytes of the dermis at 60 m from skin surface. (E) Adipocytes of the subcutaneous layer at a depth of 100 m. (F) 2D projection of 60 depth-resolved slices separated by 2 m. Panels to the right and under F show the yz and xz cross sections taken at the white lines, respectively. Images of a hairless mouse ear. The Raman shift is set at 2,845 cm1 (p 816.8 nm) to address the lipid CH2 symmetric stretch vibration. The frames are averaged for 2 s. (A) Stratum corneum with bright signals from the lamellar lipid intercellular space that surrounds the polygonal corneocytes. Bright punctuated dots are ducts of sebaceous glands. (B) Sebaceous glands at 30 m from skin surface. (C) Individual cells of the gland compartment can be recognized, with nuclei visible as dark holes (arrow). (D) Adipocytes of the dermis at 60 m from skin surface. (E) Adipocytes of the subcutaneous layer at a depth of 100 m. (F) 2D projection of 60 depth-resolved slices separated by 2 m. Panels to the right and under F show the yz and xz cross sections taken at the white lines, respectively.

    16. E CARS

    17. Detection Schemes F – CARS: Forward detected E – CARS: Epi detected P- CARS: Polarisation dependent FM CARS: Frequency modulated

    18. P CARS

    20. Detection Schemes F – CARS: Forward detected E – CARS: Epi detected P- CARS: Polarisation dependent FM CARS: Frequency modulated

    22. Sensitive probe for lipids Lipid bilayer, thin objects, small objects Fast dynamic scanning of processes in living cells High damage threshold In vivo capabilites

    23. Non resonant background term very strong Expensive laser sources Have to know beforehand the vibrational band of interest Currently limited tunability of sources – improving in line with the laser sources….

    24. Discussion?

More Related