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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

2

Disclosures

No disclosures.

Investigational research: not FDA approved.

No off label uses.

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Motivation

Applications• Fluorescence for therapy

guidance/ diagnosis─ Procedures with variable

temperature• RFA/microwave ablation• Electrocauterization• Laser ablation

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Background

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

• Tissue– Small temperature range– Very complex

• Optical property changes– Causes

• Coagulation• Dehydration• Denaturation

– Modulate fluorescence emission

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

Goal:Examine mechanism of fluorescence change due to temperature:1. Optical Properties2. Fluorophore

degradation

Materials & Methods: In vitro

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Materials• Human Tissue Samples:

– From liposuction and breast reduction surgeries– Skin

• Flash frozen samples

Passively warmed (to 23°C )

Actively heated (to 50°C or 70°C)

Allowed to cool (to 23°C)

Fluorescence and Temperature acquired every 2.5°C

Methods

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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

1.4

s (cm

-1)

temperature C

Skin

0 20 40 60 8035

40

45

50

55

60

65

a (cm

-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)

St. Dev.

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Results: Reversibility

•All skin samples showed some reversibility

•Hysteresis expected

Fl.

Pe

ak

He

igh

t (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)

St. Dev.

Cooling: Max. Temp. 70C (n=4)

Cooling: Max. Temp. 50C (n=4)

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Conclusions: In Vitro

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

Cooled Skin with Ice Pack

Skin Passively Warms to Body Temp.

Heated Skin with Heat Pack

Skin Passively Cools to Body Temp.

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Results: In Vivo

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

reversible

In Vivo Skin

Fl.

Pe

ak

He

igh

t (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.

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In Vivo: Conclusions

Fluorescence decrease can be reproduced in vivo

No damage or coagulation

Reversible

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Conclusions

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.

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Acknowledgements

•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