This document examines the effects of temperature on fluorescence in human tissue. It finds that fluorescence intensity decreases with increasing temperature in human tissue samples from 20°C to 50°C. Optical property changes alone do not explain this decrease. The fluorescence decrease can also be reproduced in vivo in human skin, with no tissue damage and the change being fully reversible. The conclusion is that in human tissue, factors other than optical properties and tissue damage cause fluorescence changes with temperature, likely fluorophore degradation above a certain temperature.

1. 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

2. Disclosures
No disclosures.
Investigational research: not FDA approved.
No off label uses.
2

3. Motivation
Applications
• Fluorescence for therapy
guidance/ diagnosis
─ Procedures with variable
temperature
• RFA/microwave ablation
• Electrocauterization
• Laser ablation
3

4. Background
• Fluorescence intensity and
temperature
– Usually inversely related
– Depends on substance
• Tissue
– Small temperature range
Goal:
– Very complex Examine mechanism of
• Optical property changes fluorescence change due
– Causes to temperature:
• Coagulation
• Dehydration 1. Optical Properties
• Denaturation
2. Fluorophore
– Modulate fluorescence emission
degradation
• Other possible mechanisms
− Loss of cell viability
− Collisional quenching
4

5. Materials & Methods: In vitro
Materials
• Human Tissue Samples:
– From liposuction and breast reduction surgeries
– Skin
• Flash frozen samples
Methods
Passively warmed (to 23°C )
Fluorescence and
Actively heated (to 50°C or
70°C) Temperature
acquired every
2.5°C
Allowed to cool (to 23°C)
5

6. Materials and Methods
6

7. Data Processing
For every
temperature, approximately
• Reflectance data used in
every 2.5°C. inverse Monte Carlo
λ : 400-800 nm µs ’
algorithm as input
Inverse • Output: µa, µs’
Rd Monte
Carlo1 µa
Spectra Spectral •Fluorescence max.
(Fl., Rd.) Processing intensity as a function of
temperature
Spectral Max.
Fl. Analysis Intensity • Normalized so that
peak intensity at 23°C
was equal to 1.
1Palmer, G.M. Appl. Opt., 2006. 45(5): p. 1062-
1071. 7

8. Results
Skin
1.4 65 3
Average (n=8)
1.3 2.8 St. Dev.
60
2.6
1.2
Fl. Peak Height (a.u.)
2.4
1.1 55
2.2
(cm-1)
(cm-1)
1
50 2
0.9
a
s
1.8
0.8 45
1.6
0.7
1.4
40
0.6 1.2
0.5 35 1
0 20 40 60 80 0 20 40 60 80 0 20 40 60 80
temperature C
• Consistent fluorescence decrease
•Optical property changes do not explain fluorescence
decrease 8

9. Results: Reversibility
Skin Reversibility
1.4
Average (n=8)
St. Dev.
1.3 Cooling: Max. Temp. 70 C (n=4)
Cooling: Max. Temp. 50 C (n=4)
•All skin samples showed
1.2 some reversibility
1.1
•Hysteresis expected
Fl. Peak Height (a.u.)
1
0.9
0.8
0.7
0.6
0.5
0 10 20 30 40 50 60 70 80
temperature C 9

10. Conclusions: In Vitro
Fluorescence intensity decreases with
increasing temperature in human tissue
Optical properties do not explain
fluorescence decrease at 20°C-50°C
Fluorophore
degradation above a
certain temperature
10

11. Materials & Methods: In vivo
Materials
• Human lateral forearm
• 7 volunteers
Methods
Cooled Skin with Ice Pack
Skin Passively Warms to Body
Temp.
Heated Skin with Heat Pack
Skin Passively Cools to Body Temp.
11

12. Results: In Vivo
In Vivo Skin
• Fluorescence
Average (n=7)
St. Dev.
decrease is 1.2
reproduced in vivo
– No damage
1.1
Fl. Peak Height (a.u.)
– Completely 1
reversible
0.9
0.8
0.7
0.6
10 15 20 25 30 35
temperature ( C) 12

13. In Vivo: Conclusions
Fluorescence decrease can be reproduced
in vivo
No damage or
coagulation
Reversible
13

14. 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
Overall
In human tissue, optical properties and tissue
damage are not the only factors that cause a
change in fluorescence due to temperature.
14

15. 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
15