Effects of Temperature on Fluorescence in Human Tissue

D.B. Masters,1,* Alex Walsh,1 Ashley J. Welch,2 E. Duco Jansen,1 and Anita Mahadevan-Jansen1

1Department of Biomedical Engineering, Vanderbilt University, 5824 Stevenson Center, Box 1631, Station B, Nashville, TN 37235 USA

2Biomedical Engineering Program, The University of Texas at Austin, 639 Engineering Science Building, Austin, TX, 78712-1084, USA

1

Disclosures

2

No disclosures.

Investigational research: not FDA approved.

No off label uses.

Motivation

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Applications• Fluorescence for therapy

guidance/ diagnosis─ Procedures with variable

temperature• RFA/microwave ablation• Electrocauterization• Laser ablation

Background

• Fluorescence intensity and temperature – Usually inversely related– Depends on substance

• Tissue– Small temperature range– Very complex

• Optical property changes– Temperature Dependent– Modulate fluorescence emission

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• Other possible mechanisms− Loss of cell viability− Collisional quenching

Fluorescence emission

µa

µs’

Goal

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Goal:Examine mechanism of fluorescence change due to temperature:

4.Optical Properties5.Fluorophore degradation

Materials & Methods: In vitro

6

Materials• Human Tissue Samples:

– From liposuction and breast reduction surgeries– Skin

• Flash frozen samples

Spectra and Temperature acquired every 2.5°C

Methods

7

Materials and Methods

Data Processing

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Spectra (Fl., Rd.)

Spectral Processing

Inverse Monte Carlo1

µs’

µa

Max. Intensity

Rd

For every temperature, approximately every 2.5°C.

λ : 400-800 nm

Spectral AnalysisFl.

1Palmer, G.M. Appl. Opt., 2006. 45(5): p. 1062-1071.

• Reflectance data used in inverse Monte Carlo algorithm as input

• Output: µa, µs’

•Fluorescence max. intensity as a function of temperature

• Normalized so that peak intensity at 23°C was equal to 1.

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Results

•Consistent fluorescence decrease•Optical property changes do not explain fluorescence decrease

Fl.

Pe

ak

He

igh

t (a

.u.)

0 20 40 60 800.5

0.6

0.7

0.8

0.9

1

1.1

1.2

1.3

µ s′ (

cm-1

)

temperature °C

Skin

0 20 40 60 80

40

45

50

55

60

µ a (c

m-1

)

0 20 40 60 801

1.2

1.4

1.6

1.8

2

2.2

2.4

2.6

2.8

3

Average (n=8)

Average(n=4)St. Dev.

Results: Reversibility

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•All skin samples showed some reversibility

•Hysteresis expected

Fl.

Pe

ak H

eig

ht (a

.u.)

temperature °C

Skin Reversibility

0 10 20 30 40 50 60 70 800.5

0.6

0.7

0.8

0.9

1

1.1

1.2

1.3

1.4

Average (n=8)

Average (n=4)St. Dev.

Cooling: Max. Temp. 70οC (n=4)

Cooling: Max. Temp. 50οC (n=4)

Conclusions: In Vitro

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Fluorescence intensity decreases with increasing temperature in human tissue

Fluorophore degradation above a certain temperature

Optical properties do not explain fluorescence decrease at 20°C-50°C

Materials & Methods: In vivo

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Materials• Human lateral forearm• 7 volunteersMethods

Results: In Vivo

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• Fluorescence decrease is reproduced in vivo– No damage– Completely

reversible

In Vivo Skin

Fl.

Pe

ak H

eig

ht (a

.u.)

temperature (°C)

10 15 20 25 30 350.6

0.7

0.8

0.9

1

1.1

1.2

Average (n=7)

St. Dev.

In Vivo: Conclusions

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Fluorescence decrease can be reproduced in vivo

No damage or coagulation

Reversible

Conclusions

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In vitro•Fluorescence intensity decreases with increasing temperature in human tissue•Optical properties do not cause fluorescence decrease from 20°C to 50°C

In vivo•Fluorescence decrease can be reproduced in vivo

•No damage•Reversible

OverallIn human tissue, optical properties and tissue damage are not the only factors that cause a change in fluorescence due to temperature.

Acknowledgements

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•All the members of the Biomedical Optics Lab•Raiyan Zaman at the University of Texas at Austin•NIH R21 CA 133477•USAF Grant for Graduate Students and Post-Doctoral Fellows Currently Involved Full-

Time in Biomedical Laser Research travel grant