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Imaging leukocyte trafficking in vivo with two-photon-excited endogenous tryptophan fluorescence.

Li C, Pastila RK, Pitsillides C, Runnels JM, Puoris'haag M, Côté D, Lin CP - Opt Express (2010)

Bottom Line: We describe a new method for imaging leukocytes in vivo by exciting the endogenous protein fluorescence in the ultraviolet (UV) spectral region where tryptophan is the major fluorophore.Two-photon excitation near 590 nm allows noninvasive optical sectioning through the epidermal cell layers into the dermis of mouse skin, where leukocytes can be observed by video-rate microscopy to interact dynamically with the dermal vascular endothelium.Because the new method alleviates the need to introduce exogenous labels, it is potentially applicable for tracking leukocytes and monitoring inflammatory cellular reactions in humans.

View Article: PubMed Central - PubMed

Affiliation: Wellman Center for Photomedicine and Center for Systems Biology, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, USA. Li.Chunqiang@mgh.harvard.edu

ABSTRACT
We describe a new method for imaging leukocytes in vivo by exciting the endogenous protein fluorescence in the ultraviolet (UV) spectral region where tryptophan is the major fluorophore. Two-photon excitation near 590 nm allows noninvasive optical sectioning through the epidermal cell layers into the dermis of mouse skin, where leukocytes can be observed by video-rate microscopy to interact dynamically with the dermal vascular endothelium. Inflammation significantly enhances leukocyte rolling, adhesion, and tissue infiltration. After exiting the vasculature, leukocytes continue to move actively in tissue as observed by time-lapse microscopy, and are distinguishable from resident autofluorescent cells that are not motile. Because the new method alleviates the need to introduce exogenous labels, it is potentially applicable for tracking leukocytes and monitoring inflammatory cellular reactions in humans.

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(a-c) Tryptophan fluorescence images of C57BL/6 mouse ear at (a) 10 mins, (b) 60 mins, and (c) 120 mins post UV exposure (scale bar 50 µm). (d) Single-frame excerpts from video recordings of leukocyte migration in skin tissue (Media 3). (e) Trajectory of the centroid of a migrating leukocyte at 80-second interval (scale bar 20 µm).
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g007: (a-c) Tryptophan fluorescence images of C57BL/6 mouse ear at (a) 10 mins, (b) 60 mins, and (c) 120 mins post UV exposure (scale bar 50 µm). (d) Single-frame excerpts from video recordings of leukocyte migration in skin tissue (Media 3). (e) Trajectory of the centroid of a migrating leukocyte at 80-second interval (scale bar 20 µm).

Mentions: We also induced a mild degree of inflammation on C57BL/6 mice (pigmented) ear skin by exposing them to the UV radiation and imaged the mouse ear dermis every 10 minutes and continued for 2 hours after UV exposure. Figure 7 (a)Fig. 7


Imaging leukocyte trafficking in vivo with two-photon-excited endogenous tryptophan fluorescence.

Li C, Pastila RK, Pitsillides C, Runnels JM, Puoris'haag M, Côté D, Lin CP - Opt Express (2010)

(a-c) Tryptophan fluorescence images of C57BL/6 mouse ear at (a) 10 mins, (b) 60 mins, and (c) 120 mins post UV exposure (scale bar 50 µm). (d) Single-frame excerpts from video recordings of leukocyte migration in skin tissue (Media 3). (e) Trajectory of the centroid of a migrating leukocyte at 80-second interval (scale bar 20 µm).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3369551&req=5

g007: (a-c) Tryptophan fluorescence images of C57BL/6 mouse ear at (a) 10 mins, (b) 60 mins, and (c) 120 mins post UV exposure (scale bar 50 µm). (d) Single-frame excerpts from video recordings of leukocyte migration in skin tissue (Media 3). (e) Trajectory of the centroid of a migrating leukocyte at 80-second interval (scale bar 20 µm).
Mentions: We also induced a mild degree of inflammation on C57BL/6 mice (pigmented) ear skin by exposing them to the UV radiation and imaged the mouse ear dermis every 10 minutes and continued for 2 hours after UV exposure. Figure 7 (a)Fig. 7

Bottom Line: We describe a new method for imaging leukocytes in vivo by exciting the endogenous protein fluorescence in the ultraviolet (UV) spectral region where tryptophan is the major fluorophore.Two-photon excitation near 590 nm allows noninvasive optical sectioning through the epidermal cell layers into the dermis of mouse skin, where leukocytes can be observed by video-rate microscopy to interact dynamically with the dermal vascular endothelium.Because the new method alleviates the need to introduce exogenous labels, it is potentially applicable for tracking leukocytes and monitoring inflammatory cellular reactions in humans.

View Article: PubMed Central - PubMed

Affiliation: Wellman Center for Photomedicine and Center for Systems Biology, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, USA. Li.Chunqiang@mgh.harvard.edu

ABSTRACT
We describe a new method for imaging leukocytes in vivo by exciting the endogenous protein fluorescence in the ultraviolet (UV) spectral region where tryptophan is the major fluorophore. Two-photon excitation near 590 nm allows noninvasive optical sectioning through the epidermal cell layers into the dermis of mouse skin, where leukocytes can be observed by video-rate microscopy to interact dynamically with the dermal vascular endothelium. Inflammation significantly enhances leukocyte rolling, adhesion, and tissue infiltration. After exiting the vasculature, leukocytes continue to move actively in tissue as observed by time-lapse microscopy, and are distinguishable from resident autofluorescent cells that are not motile. Because the new method alleviates the need to introduce exogenous labels, it is potentially applicable for tracking leukocytes and monitoring inflammatory cellular reactions in humans.

Show MeSH
Related in: MedlinePlus