# Ex Vivo Exposure to Soft Biological Tissues by the 2-um All-Fiber Ultrafast Holmium Laser System
## 4 Conclusions  
In this work, we studied the exposure of ultrashort pulsed laser radiation in the spectral range of 2-2.2 \(\upmu\)m with a central wavelength of 2.06 \(\upmu\)m to porcine longissimus tissue ex vivo. A simple, robust and compact all-fiber Ho-doped laser system generated ultrashort pulsed radiation with a duration of less than 1 ps and a repetition rate of 20 MHz. This type of ultrafast all-fiber Holmium laser system with a high pulse frequency and low energy for soft tissue irradiation has not yet been reported in the literature. After ex vivo single-spot ablation experiments on the muscle tissue in the range of applied energy 1.5-18 J, the depth and diameter of coagulation zones, ablation craters and thermal damage zones were evaluated.  
Comparison of USP laser exposure at an average power of 0.3 W with a CW laser at the same power and exposure time showed a difference in results. The continuous-wave laser radiation exposure led to reversible damage, namely the formation of a heat-affected zone on the tissue surface without destructive changes in the cell structure. The ultrashort pulsed radiation with the same average power caused more pronounced destructive tissue damage, which could be associated with the mechanism of laser-tissue interaction, and possibly nonthermal. The relative power safety for this USP laser exposure, resulting in non-destructive and reversible effects, is less than 1.5 J. Comparison of ablative efficiency has shown that USP laser ablation can be achieved at lower average power values compared to CW laser ablation. In addition, morphological studies have shown that the tissue at the edge of the formed crater during USP laser ablation was less susceptible to carbonization.  
It should be noted that tissue carbonization is often undesirable in surgical practice because charred tissue has a tendency to a longer healing period and, as a consequence, to a higher risk of infection [54]. We can assume that, as proposed in [36,55], cutting the high-repetition-rate pulse sequence and creating pulse bursts with a relatively low repetition rate could contribute to a decrease in thermal damage of tissues, and consequently a reduction of carbonization.  
The results obtained from ex vivo experiments with tissues of postmortem animals are part of a major study of the effect on biological tissues of two-micron laser radiation. The next stage will be in vivo experiments on animals using the laser systems studied in this work and the previous one [41].  
Conceptualization, V.B.T.; Methodology, M.S.K., S.A.F., V.A.K., A.I.T. and E.I.K.; Investigation, M.S.K., S.A.F., V.A.K. and E.I.K.; Data analysis, M.S.K., V.V.A., S.A.F. and V.B.T. Writing--original draft preparation and editing, M.S.K., S.A.F., V.B.T., V.A.K., V.V.A. and V.B.L.; Visualization, M.S.K. and S.A.F.; Supervision, S.A.F.; Project administration, V.B.T. All authors have read and agreed to the published version of the manuscript.  
Figure 7: (**a**) Ablation area and (**b**) ablation efficiency as a function of applied energy of USP and CW laser radiation for different times of exposure (5, 30 and 60 s) and laser power (0.3 \(W\), 0.5 W), shown as applied energy.  
**Funding:** This research was funded with the financial support of the Ministry of Science and Higher Education of Russian Federation, grant number 075-15-2020-912 and carried out on the premises of the World-Class Research Center <<Photonics<<.  
**Institutional Review Board Statement:** Not applicable.  
**Informed Consent Statement:** Informed consent was obtained from all subjects involved in the study.  
**Data Availability Statement:** The data supporting the findings of this study are available within the article.  
**Conflicts of Interest:** The authors declare no conflict of interest.