Multimodal Microscopy

Imaging modalities such as confocal reflectance microscopy, fluorescence by single- or two-photon excitation, second harmonic generation and coherent anti-Stokes Raman spectroscopy (CARS) provide different contrast mechanisms that can be combined to give structural, functional and molecular information of living tissue. Our group has developed a multimodal microscope in which up to three of these imaging modalities can be realized simultaneously. Images are acquired at video rate, which allows real-time monitoring of fast events in the living tissue.

Adaptive Optics

Ophthalmic imaging resolution suffers from aberrations present in the eye. With the introduction of an adaptive optics (AO) system into a scanning laser ophthalmascope (SLO) one can achieve the best possible resolution for the system. We have implemented such an AO system into a SLO designed for imaging the mouse retina.

Gfp_microglia_1 (a)
Gfp_microglia_2 (b)
Images of GTP labeled microglia in the living mouse retina. (a) Aberrated image (AO system off). (b) Corrected image (AO system running).

In vivo flow cytometry

Our group has developed an in vivo flow cytometer, based on confocal design. Contrary to the conventional flow cytometer, the in vivo flow cytometer can provide real-time detection and quantitative information on fluorescently labeled cells while in circulation in a live animal model. As the labeled cells pass through a slit of light focused across a blood vessel, fluorescence is excited. Its confocal detection makes it possible to observe the cell population of interest without the need to extract a blood sample. Furthermore, the same cell population can be tracked continuously and over long periods of time to examine the dynamic changes in the circulation of different types of cells in the same animal. The in vivo flow cytometer has been used to measure the circulation lifetime of different tumor cells and leukocyte populations in the peripheral circulation in response to immunological stress or therapeutic manipulation.

Cells (circles) flow through the blood vessel as shown. A slit of excitation light (green box) illuminates passing cells. When a fluorescently labeled cell passes through the “slit” it emits light.
Sample flow cytometer trace. (Novak et al, Opt. Lett. 2004)

Charles Lin