Development of a fiber laser with independently adjustable properties for optical resolution photoacoustic microscopy

ABSTRACT Photoacoustic imaging is based on the detection of generated acoustic waves through thermal expansion of tissue illuminated by short laser pulses. Fiber lasers as an excitation

source for photoacoustic imaging have recently been preferred for their high repetition frequencies. Here, we report a unique fiber laser developed specifically for multiwavelength

photoacoustic microscopy system. The laser is custom-made for maximum flexibility in adjustment of its parameters; pulse duration (5–10 ns), pulse energy (up to 10 μJ) and repetition

frequency (up to 1 MHz) independently from each other and covers a broad spectral region from 450 to 1100 nm and also can emit wavelengths of 532, 355, and 266 nm. The laser system consists

of a master oscillator power amplifier, seeding two stages; supercontinuum and harmonic generation units. The laser is outstanding since the oscillator, amplifier and supercontinuum

generation parts are all-fiber integrated with custom-developed electronics and software. To demonstrate the feasibility of the system, the images of several elements of standardized

resolution test chart are acquired at multiple wavelengths. The lateral resolution of optical resolution photoacoustic microscopy system is determined as 2.68 μm. The developed system may

pave the way for spectroscopic photoacoustic microscopy applications via widely tunable fiber laser technologies. SIMILAR CONTENT BEING VIEWED BY OTHERS MULTIMODAL NONLINEAR ENDOMICROSCOPIC

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2024 INTRODUCTION Photoacoustic microscopy (PAM) is a promising imaging modality that combines optical and ultrasound imaging. It takes advantage of high optical contrast and high ultrasonic

spatial resolution owing to its hybrid nature. When a short laser pulse illuminates tissue, absorbed light leads to acoustic emission via thermoelastic expansion1,2,3,4,5,6,7,8,9,10.

Generated ultrasonic waves are conventionally detected by transducers. Recorded signals are used to map the distribution of the locations of optical absorbers. Relatively low scattering of

ultrasonic waves in biological tissues provides deeper penetration beyond the optical transport mean free path4. The contrast of PAM is endogenously produced by optical absorption of

chromophores within the tissue11,12. The laser system needs to produce short enough pulses, _i.e._, several nanoseconds, in order to generate photoacoustic signals efficiently and emit

wavelengths in the visible range to cover absorption peaks of tissue chromophores in their spectra4,13,14. To obtain adequate penetration depth, it is also desirable to utilize a wavelength

in the near infrared range, from 600 to 1200 nm, where biological tissues are relatively transparent15,16. Several kinds of lasers have been used for photoacoustic imaging. Pulsed laser

diodes draw researchers’ attention by being compact and inexpensive. While the peak power is relatively modest15,17, it is sufficient to obtain adequate signal-to-noise ratio for _in-vivo_

optical resolution photoacoustic microscopy (OR-PAM), as demonstrated in several publications18,19,20,21,22. On the other hand, they found only limited place in photoacoustic applications

due to their lack of continuous tunability in wavelength. Q-switched Nd:YAG lasers operating at 1064 nm (and/or acquiring 532 nm by frequency doubling) are frequently utilized for PAM15,23.

They are generally preferred because of their easy accessibility. However, their fixed wavelength output is a serious drawback for multispectral photoacoustic applications which quantify

unique spectral features of each absorber by a set of wavelengths. On the other hand, Q-switched Nd:YAG pumped dye lasers, Ti:Sapphire lasers, and optical parametric oscillators (OPOs) are

usually preferred for providing necessary wavelength tuning with high pulse energies (>1 mJ)7,8,14,24,25,26,27,28,29,30,31,32; yet, they have some major limitations of their practical

applications such as having low pulse repetition rate (generally less than 50 Hz, recently up to several kHz for OPOs33,34), being bulky and expensive, and requiring external cooling

units35. For the sake of enabling spectroscopic measurements, multiwavelength spectrum is obtained from a single wavelength emitting Q-switched Nd:YAG microchip laser, either through

stimulated Raman scattering (SRS) or nonlinear broadening by coupling its output to a fiber36,37,38,39,40,41,42,43,44,45. For lasers utilizing SRS, major energy is distributed on a series of

fixed individual wavelength peaks that result from nonlinear interaction between incoming photons through the fiber and the molecules in the fiber itself, thus offers a limited wavelength

tunability46. Koeplinger _et al_.41 reported four bands in a polarization maintaining single mode fiber (PM-SMF), and Loya _et al_.40 improved the system with a broader wavelength tuning

range also with a higher repetition rate and pulse energy per band. It was also demonstrated that both discrete lines and a continuum can be produced by using four-wave mixing in a special

fiber (SMF-28e)42. As a different technique, Buma _et al_.43 used a birefringent optical fiber and produced discrete spectral bands in near infrared region. Much broader wavelength tuning

can potentially be achieved by a supercontinuum source such as photonic crystal fiber (PCF), which relies on spectral broadening through nonlinear processes36,47,48,49. PCF is a silica

optical fiber with an ordered array of microscopic air holes running along its length50,51. Billeh _et al_.36 utilized PCF for developing spectroscopic photoacoustic microscopy system. Lee

_et al_.37 also built a supercontinuum laser system for both PAM and optical coherence tomography (OCT). Afterwards, Lee _et al_.38 determined oxygen saturation of hemoglobin and hemoglobin

concentration via the same laser source. Whensoever the applications by coupling the output of Q-switched Nd:YAG microchip to PCF are considered, energy per band is reported to be lower in

supercontinuum case than SRS, which may be a drawback for many applications46. In order to achieve wider tunability in the wavelength with high energy per band, Shu _et al_.39 proposed a

master oscillator power amplifier (MOPA) laser system with a homebuilt yterrbium-doped (Yb) fiber amplifier for power boost. The amplifier was coupled to a specially designed PCF taper that

connects a large-core fiber that has a much more resistance to high-pulse energy at the input to a small-core PCF for spectrum broadening. Pulse energy per band increased dramatically and

became comparable to the ones produced through SRS39,45. Apart from wavelength tunability, a laser system with high pulse repetition frequency (PRF) is also desired for fast image

acquisition. The repetition frequencies of solid-state lasers are limited up to several kHz; but recently, fiber lasers with high repetition rates emerge as an alternative excitation source

for PAM. Through their high repetition rate, near real and real time imaging can be achieved46,52,53,54. It has already been reported that in comparison to conventional systems with solid

state lasers, the ones with fiber lasers are at least two orders of magnitude faster without compromising lateral resolution52,55. Fiber laser sources are also used for _in vivo_ and _in

vitro_ studies also including flow cytometry applications52,54,55,56,57,58. The main disadvantage of these systems is their fixed wavelength that does not allow for multispectral functional

imaging. To overcome the limitations, fiber laser technology seeking for tunability in wavelength is put forward. Hajireza _et al_.59 developed an SRS fiber laser source for photoacoustic

imaging. They coupled the output of an Yb fiber laser into a PM-SMF in varying lengths at different PRFs and extended the number of wavelengths at SRS peaks that were previously

limited46,60. In recent years, due to high power capabilities, MOPA laser systems have begun to be developed61,62,63. The first demonstration of a short pulse MOPA fiber laser at 1 μm was

the study by Ilday _et al_.64. Allen _et al_.61 produced a fiber laser system with a high repetition frequency in MOPA configuration with a single emission wavelength of 1064 nm. Mahmud _et

al_.62 demonstrated an OR-PAM system by using a commercial picosecond MOPA laser system consisting of a fiber-based tunable oscillator and three amplifier stages with a high power booster

amplifier. However, the wavelength tunability was limited with 50 nm bandwidth. Here, to address the limitations of each approach, we develop a tunable fiber based MOPA laser system

producing nanosecond pulses, covering spectrum from 450 nm to 1100 nm, specifically for PAM. The supercontinuum part is all fiber-integrated; guided-beam-propagation renders its misalignment

free and largely immune to mechanical perturbations. Free space harmonic generation creates higher pulse energy for a specific band, i.e. 532 nm, and also generates ultra violet (UV) light

with wavelengths of 355 and 266 nm. Total supercontinuum output power is over 1 W and visible output power is around 270 mW at 65 kHz repetition rate corresponding to 4 μJ pulse energy. One

of the novelties here is the improvement of wavelength tunability, output power and pulse energy when fiber-based lasers are benchmarked. This is the first demonstration of spectroscopic PAM

by developing a supercontinuum all-fiber based MOPA source. The tunability of the laser parameters allows using only one laser for many different PAM applications, and also high repetition

rate enables fast scanning. The coverage of near-UV spectrum gives an opportunity to image cell nuclei. As certain morphological changes such as size and shapes irregularities in the nuclei

are known indicators of various cancers30,65,66, we believe our system may also be useful for cell nuclei studies as well. RESULTS A standardized resolution test target (USAF-1951, Thorlabs)

was imaged for determination of the lateral resolution of our OR-PAM system. A transducer (V384, Panametrics) with a 3.5 MHz center frequency was used to acquire photoacoustic signals at

the optical wavelength of 1064 nm filtered from the supercontinuum output. For focusing the light, a 5× objective (LMH-5 × −1064, Thorlabs) was used. The target was immersed in water, then

2D raster scanning by a motorized linear translation stage (LNR50SEK1, Thorlabs) along the _x-y_ plane in steps of 1 μm for an area of 300 × 330 μm2 was performed. The acquired signals were

averaged over 128 consecutive signal cycles. The trigger signal from the field programmable gate array (FPGA) of the laser was used to trigger a data acquisition card (DAQ) for

synchronization. Following the triggering of each laser pulse, photoacoustic signals were initially amplified by 40 dB using a pre-amplifier (5678, 40 MHz bandwidth, Olympus) and then 39 dB

via a pulser/receiver (5073PR, Olympus). The signals were digitized through a DAQ (Razor Express CompuScope 1422, Gage Applied Technologies, Inc.), then data processing and reconstruction

were performed. Figure 1a shows the optical microscopy image and Fig. 1b presents the maximum amplitude projection (MAP) image of the scanned area (Group 6 and 7) of the test target. The

lateral full width at half-maximum (FWHM) value from the imaged highlighted well resolved bars (Group 7, Element 6) was determined as 2.68 μm, as shown in Fig. 1d. Furthermore, for the

demonstration of our multiwavelength PAM system, Group 5 Element 6 of the test target were also imaged with six different wavelengths of 532, 650, 697, 732, 785, and 880 that can be seen in

Fig. 2a,b,c,d,e and f, respectively. These wavelength values except from 532 nm which was obtained by second harmonic generation (SHG), were filtered from the supercontinuum output of the

laser for each experiment. A 10× objective (Plan Achromat, 0.25 NA, Olympus) was used to focus light to the relevant area. DISCUSSION In order to evaluate the performance of our laser

system, pulse energy, average power and repetition rate values are compared with the ones in existing systems within the literature including fiber components and independent of seeding

laser type. Billeh _et al_.36 sent the output of a Q-switched Nd:YAG microchip laser with a repetition frequency of 6.6 kHz to a 7 m-long PCF and reported seven wavelengths of 575, 625, 675,

725, 775, 825, and 875 nm with a bandwidth of 40 nm for each wavelength and pulse energies were measured as 7, 15, 24, 31, 31, 31, and 33 nJ, respectively. Lee _et al_.38 also sent the

output of the same type of laser to a 10 m-long PCF and stated pulse energy of the generated supercontinuum light as 500 nJ. The pulse energies of two bands, 500 to 560 and 560 to 660 nm

were measured as 0.6 and 1.8 nJ, respectively. As can be seen, these pulse energies are quite low despite the wide bandwidths. There are many attempts to develop multiwavelength laser

systems generating higher pulse energies from the output of an integrated fiber for various photoacoustic imaging applications36,39,40,41,45. However, this condition requires PCFs to

withstand such high energies. Since non-linearity increases as the effective mode area of fiber gets smaller; thus, it is advantageous to decrease core diameter for generation of more

efficient supercontinuum. Yet, energy per surface area of the fiber has a major effect on the maximum optical pulse peak power which a fiber can withstand67,68,69. Therefore, there is a

trade-off between supercontinuum efficiency and energy to be coupled into the fiber. In order to overcome this limitation, tapered fibers are designed. Bondu _et al_.45 used a nonlinear

fiber that combines a large-core fiber for high-pulse energy handling with a small-core fiber for efficient spectral broadening. They used five different PCFs with varying core diameters,

two of them were tapered for supercontinuum generation. They also demonstrated that total energy at the output of the straight PCF with core diameters of 5, 9, and 10 μm as 10, 29.5, and 30 

μJ, respectively with visible output energies of 1.7, 5.4, and 4.6 μJ. Total output energy of tapered PCF of length of 1 m with an input core diameter of 10 μm tapered down to 5 μm was

stated as 22 μJ with visible output energy of 6 μJ39,45. Taking advantage of SRS inside a fiber is another method to increase the number of wavelengths from a fixed wavelength output.

Polarization-maintaining single-mode fiber (PM-SMF) as well as PCF have been used for generation of SRS peaks40,41,46,59,60. Koeplinger _et al_.41 sent the output of a Q-switched Nd:YAG

microchip laser with a repetition frequency of 7.5 kHz to a frequency-doubling KTP crystal. Then, this output was sent to a 6 m-long PM-SMF and acquired four distinct bands 546, 560, 574,

and 600 nm with a pulse energy of 80 nJ for the each wavelength. Loya _et al_.40 coupled the output of a Q-switched Nd:YAG laser operating at 30 kHz repetition rate to a 30 m-long large mode

area photonic crystal fiber (LMA-PCF) and individual pulse energies were reported as 270, 360, 520, 530, and 400 nJ at wavelengths of 532, 546, 568, 589, and 600 nm, respectively. Hajireza

_et al_.46,59,60 coupled an Yb-doped fiber laser into a PM-SMF in varying lengths at different PRFs and extended the number of wavelengths at SRS peaks. The acquired pulse energies were in

between 100 to 500 nJ. Our tunable fiber-based laser system has three outputs; supercontinuum (from 450 to 1100 nm), 1064 nm from single-wavelength emitting port, and harmonic generation

(532, 355, and 266 nm). The average power of 1064 nm output is around 3 W which seeds harmonic generation unit but also can be used for its own applications. The maximum average power values

of SHG (532 nm), third harmonic generation (THG, 355 nm), and fourth harmonic generation (FHG, 266 nm) are 500, 3, 10 mW, respectively. Total output power of supercontinuum is measured over

1 W with visible output power around 270 mW with a powermeter (S314C, Thorlabs) at 65 kHz repetition rate that corresponds to 17 μJ total and 4 μJ visible energy. Various bandpass filters

are used to obtain wavelength of interest from supercontinuum output and power measurements are performed to compare with the values in the literature. In order not to damage bandpass

filters, a 1000 nm shortpass filter is firstly employed. Average power values at wavelengths of 680 and 830 nm with 10 nm bandwidths are measured as 5 and 11 mW by a powermeter (S142C,

Thorlabs) after the achromatic lens that corresponds to 76 and 169 nJ pulse energy. For wider bandwidths, average power values for wavelengths of 650, 697, 732, 785, and 880 nm with 80, 75,

68, 62, and 70 nm bandwidths are 92, 93, 82, 84, 142 mW, respectively. Corresponding pulse energies are 1.4, 1.4, 1.3, 1.3, 2.2 μJ. These energies are higher than the ones produced through

coupling the output of Q-switched Nd:YAG microchip laser to PCF which is at most 33 nJ36. As mentioned above, for the special case of tapered PCFs, visible output energy was reported as 6 μJ

at 25 kHz, for our system that is 4 μJ at 65 kHz and comparable to that output39,45. In addition to that, our laser source can provide higher pulse repetition rate, up to 1 MHz, at the

expense of lower pulse energies. For the systems utilizing SRS, the energies per band were reported several hundreds of nJ with an utmost energy of 500 nJ46,60. SRS peaks are produced with a

bandwidth around 10 nm, pulse energies are higher than our system for such narrow bandwidths for visible region. However, when filters with wider bandwidths are selected, pulse energies

become higher than ones that SRS peaks possess. To be also noted, pulse energies of SRS peaks decreases (estimated around 100 nJ) elongating near-infrared spectral region. The edge of peaks

was noted as 788 nm46, our spectrum covers up to 1100 nm. Allen _et al_.61 produced an all-fiber laser source with a PRF up to 2 MHz but the output wavelength was fixed. Mahmud _et al_.62

also reported a fiber based laser source. By means of electronic modulations in the oscillator, tuning the repetition rate (0.1–120 MHz), the pulse-width (0.1–5 ns) and the wavelength

(1030–1080 nm) were carried out. Green light was also generated through frequency doubling. The output power was reported up to 1.1 W and pulse energy up to 500 nJ. However, wavelength

cannot be tuned in a broad range which does not allow for various spectroscopic photoacoustic applications. There are many other advantages of our system. All the laser parameters, which are

reported as independently adjustable, could be achieved by changing FPGA configuration and currents to the pump diodes electronically without any mechanical intervention. The only exception

to this is the switching among the supercontinuum and harmonic generation ports, which is achieved by a mechanically switchable mirror, that can also readily be motorized, if desired. In

addition to this, it is very compact with dimensions of 40 × 40 × 9 _cm_3 except from free-space harmonic generation unit and does not require any big cooling unit. Thanks to its high PRF,

it may be a promising source for cytometry as well57. In our system, the light is transmitted through the splice between Yb-doped fiber and PCF for rendering all-fiber integrity with an

efficiency of 40%. One of the disadvantages of current configuration is the heating at the splice point. Despite the cooling fan, the splice should be renewed once in a while in order to

compensate for decreasing power in time. In order to handle the issue for robust and long-term operation, the splicing between the gain fiber and the PCF is optimized for low-loss and high

tensile strength (using GPX-3000 series splicer, Vytran, Inc.), as demonstrated in the context of _in-situ_ absorption spectroscopy of plasmas using a similar supercontinuum source and the

same type of fibre70. Free space coupling is also possible between Yb-doped fiber and PCF; in that case transmission can be performed with higher efficiency and higher pulse energies can be

produced if all-fiber integrity is disregarded. The present limitations to the continuously and independently adjustable laser parameters arise from the requirement of simultaneous

satisfaction of the following conditions during laser design: ensuring that each amplification stage is seeded with sufficient power to prevent generation of laser noise in the form of

amplified spontaneous emission (ASE), ensuring that the targeted, final pulse duration will depend on the seed pulse duration in a complex manner due to gain saturation and that there is

sufficient peak power to accomplish the supercontinuum generation in the PCF. We believe that even a large range of parameters are possible, albeit at the cost of increased system complexity

(by adding a second AOM and additional amplifier stages). The present parameter range was decided based on the balance between system complexity and sufficiency for most typical OR-PAM

applications. To sum up, when all-fiber based laser systems are taken into consideration, the developed system improves the wavelength tunability with a repetition rate up to 1 MHz. For

laser systems having fiber components, pulse energies of this system are higher from PCF coupled supercontinuum cases and comparable to the outputs of special tapered PCF designs. The system

also offers all-fiber integrity and higher PRF by means of custom developed FPGA electronics that controls laser diode. Pulse energies of SRS peaks can be surpassed at near-infrared region

with same bandwidth, at visible region only by using filters with wider bandwidths. This paper presents the potential of a tunable fiber laser system in MOPA configuration for

multiwavelength OR-PAM. We believe that the system may provide the means of spectroscopic photoacoustic microscopy applications via widely tunable fiber laser technologies. METHODS For

photoacoustic microscopy system, a widely tunable fiber laser system is designed in MOPA configuration. Figure 3 shows the general scheme of the laser system. The output of MOPA

configuration seeds two arms; the first one is used for supercontinuum generation via spectrum broadening and the second is for harmonic generation through nonlinear crystals. Pulses with

sufficiently narrow bandwidths are required for harmonic generation (second, third, and fourth)71 through nonlinear crystals. The increase in the length of the crystal results in more

efficient wavelength conversion; yet, longer crystals bring along phase shifts proportional to the bandwidth of the laser, and decrease the efficiency72. For this reason, a 1064 nm

fiber-coupled diode laser (I-IV Laser Enterprise) with a very narrow bandwidth (0.3 nm) is used and driven by a nanosecond diode driver (PicoLas, LDP – V03–100 UF V3). Pulse width of the

laser diode is adjusted through a field programmable gate array (FPGA) card (BASYS2, Xilinx). 15 ns long pulses at 65 kHz repetition rate are generated and sent to Yb-doped gain fiber after

passing through an isolator and an amplified spontaneous emission (ASE) filter. As a pump source, a 976 nm laser diode (II-VI Laser Enterprise) delivering a maximum power of 540 mW is used.

The pump is first passed through a pump protection filter with a maximum power handling of 300 mW, followed by a 30:70 coupler allotting two stages of preamplifier. In the first stage,

Yb-doped fiber is backward-pumped by 30% of the output of the laser diode, then combined with the signal through a wavelength division multiplexer (WDM). Backward pumping is crucial for

decreasing ASE generation rate, and hence preventing possible damage to the pump diodes and other fiber components. Another ASE filter is used between pre-amplifier stages to prevent the

first from ASE that may be produced in the second. A WDM is used to combine 70% of the output of the laser diode pump and the first stage of the preamplifier. For amplification, an Yb-doped

fiber is used and the output power is measured as 170 mW at 65 kHz repetition rate. The last component of the second preamplifier is an isolator with a maximum power handling of 2 W in order

to protect it from back reflections. At the end of the preamplifier, a 30:70 coupler separates the signal, 30% is utilized for supercontinuum and 70% is for harmonic generation.

Polarization of light is crucial for frequency multiplication; thus, 70% of the allocated signal is passed through a polarizer and all fiber components beyond this point are polarization

maintaining. A 976 nm diode laser is used and a multi-mode pump combiner (MPC) combines the pump and signal. A polarization maintaining double cladding Yb-doped (PM-DC-Yb) fiber is spliced

to the end of the MPC for amplification of the signal and pulses with 8 ns duration with an average power of 3 W at 65 kHz repetition rate are acquired. Figure 4a shows the optical spectrum

and Fig. 4b shows the temporal profile of a pulse at the end of the amplification. In the temporal profile, the leading edge of the pulse is sharpened, or self-steepened, as the gain is

partially saturated by each individual pulse and consequently less gain is available for the trailing edge. The temporal structure in the trailing edge is a static structure, which does not

vary from pulse to pulse, originating primarily from the dynamically varying impedance of the semiconductor diode that seeds the system. Besides, 30% of the signal having an average power of

45 mW is firstly amplified for supercontinuum generation, a 15 m long PCF (SC 5.0–1040, NKT) with 5 μm core size is spliced to the end of Yb-doped fiber (Yb-1200 20/125 PM, nLight Liekki).

The core size of the Yb-doped fiber is 20 μm which is larger than the core size of the PCF. For this reason, a special splice is used in between the Yb-doped fiber and PCF by a suitable

splicer (FSM-100M, Fujikura). Figure 5a and b show the photograph of the output of supercontinuum and harmonic generation units, respectively. Optical spectrum of the supercontinuum is

measured by two optical spectrum analyzers (OSA) with different wavelength ranges; OSA 1 (Avaspec-3648-VIS, Avantes) and OSA 2 (QE65 Pro, Ocean Optics). The acquired spectra are digitally

combined in a single figure (Fig. 6a). In the first spectrum, the intensity of near infrared region appears lower than its actual level due to the decrease in the response of the analyzer

while approaching to the edges of the measurable spectra region. It may also be caused by the difficulty of collecting all the beam with broad spectrum which is collimated by a single lens.

Although the lens is an achromatic lens, it may still not be enough to eliminate slight divergence for different wavelengths and thus amplitude measurement variation throughout this broad

spectrum range. In the second one, the intensity of the region between 450 to 650 nm lowered to noise level as a result of using neutral density filters in order to prevent saturation of the

detector for the remaining spectrum. For frequency multiplication process, a half wave plate is employed to match the polarization between the isolator and crystals. An anti-reflection

coated (for 1064 nm wavelength) lens with a focal length of 30 mm is used to focus light into crystal. For SHG, a 20 mm long Lithium Triborate (LBO) crystal (Eksma, LBO-405) is used. For

non-critical phase matching (NCPM), a crystal oven and a proportional–integral (PI) controller is added to maintain the temperature at 150.8 °C that results in maximum power. The light is

passed through an anti-reflection coated (for 532/1064 nm) lens for collimation. Two dichroic mirrors separate the generated SHG beam (532 nm light) from the 1064 nm beam. Here, the output

power is measured as 500 mW for 532 nm light. A mirror hold including a dichroic mirror reflecting 532 nm wavelength is added to the system. When the mirror is flopped, beam including 532

and 1064 nm wavelengths pass through a lens to enter a crystal (Eksma LBO-407) for THG. The crystal is maintained at 40 °C for NCPM. The output power is around 3 mW for 355 nm light. Another

flip mirror mount with a dichroic mirror that is transmitting 1064 nm and reflecting 532 nm beam is added to direct the beam toward a lens with a focal distance of 30 mm. This lens focuses

the beam into a Barium Borate (BBO) crystal (Eksma BBO-700, thickness = 6 mm) that generates second harmonic of the 532 nm beam (fourth harmonic generation), which results in around 10 mW of

266 nm light. The output of the crystal is filtered via a dichroic mirror reflecting 266 nm light and collimated by using a UV-coated lens with a focal distance of 50 mm. The optical

spectrum of SHG is shown in Fig. 6b and of THG in Fig. 6c. The spectra are acquired with OSA 2 and OSA 1, respectively. Figure 3 shows the schematics of fiber laser in MOPA configuration,

all-fiber supercontinuum, and free-space harmonic generation units. The schematics of experimental setup for transmission mode OR-PAM system by using the irradiation source explained

previously is shown in Fig. 7. Pulse duration of the laser is 8-ns for harmonics generation output and 10 ns for supercontinuum port. ADDITIONAL INFORMATION HOW TO CITE THIS ARTICLE:

Aytac-Kipergil, E. _et al_. Development of a Fiber Laser with Independently Adjustable Properties for Optical Resolution Photoacoustic Microscopy. _Sci. Rep._ 6, 38674; doi:

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references ACKNOWLEDGEMENTS This research was supported in part by TUBITAK Grant No. 213E033, Bogazici University Research funding Grant No. BAP 15B03TUG3, and the European Research Council

(ERC) Consolidator Grant ERC-617521 NLL. AUTHOR INFORMATION Author notes * Aytac-Kipergil Esra, Demirkiran Aytac, Uluc Nasire and Yavas Seydi contributed equally to this work. AUTHORS AND

AFFILIATIONS * Department of Physics, Bogazici University, Istanbul, 34342, Turkey Esra Aytac-Kipergil, Aytac Demirkiran, Nasire Uluc, Tunc Kayikcioglu & Mehmet Burcin Unlu * Institute

of Materials Science and Nanotechnology, Bilkent University, Ankara, 06800, Turkey Seydi Yavas * FiberLAST, Inc., Ankara, 06800, Turkey Seydi Yavas & Sarper Salman * Department of

Electrical and Electronics Engineering, Bilkent University, Ankara, 06800, Turkey Sohret Gorkem Karamuk & Fatih Omer Ilday * Lumos Laser, Ltd., Ankara, 06500, Turkey Sohret Gorkem

Karamuk * Department of Physics, Bilkent University, Ultrafast Optics and Lasers Group, Ankara, 06800, Turkey Fatih Omer Ilday Authors * Esra Aytac-Kipergil View author publications You can

also search for this author inPubMed Google Scholar * Aytac Demirkiran View author publications You can also search for this author inPubMed Google Scholar * Nasire Uluc View author

publications You can also search for this author inPubMed Google Scholar * Seydi Yavas View author publications You can also search for this author inPubMed Google Scholar * Tunc Kayikcioglu

View author publications You can also search for this author inPubMed Google Scholar * Sarper Salman View author publications You can also search for this author inPubMed Google Scholar *

Sohret Gorkem Karamuk View author publications You can also search for this author inPubMed Google Scholar * Fatih Omer Ilday View author publications You can also search for this author

inPubMed Google Scholar * Mehmet Burcin Unlu View author publications You can also search for this author inPubMed Google Scholar CONTRIBUTIONS M.B.U. initiated the study, along with F.O.I.

directed the study and revised the manuscript. E.A.K., A.D., N.U., S.Y. and S.S. carried out the experiments. T.K. and E.A.K. developed the data acquisition scheme. S.K. contributed to

software development of FPGA. E.A.K., A.D., N.U. wrote the manuscript with comments from all authors. ETHICS DECLARATIONS COMPETING INTERESTS The authors declare no competing financial

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CITE THIS ARTICLE Aytac-Kipergil, E., Demirkiran, A., Uluc, N. _et al._ Development of a Fiber Laser with Independently Adjustable Properties for Optical Resolution Photoacoustic Microscopy.

_Sci Rep_ 6, 38674 (2016). https://doi.org/10.1038/srep38674 Download citation * Received: 02 August 2016 * Accepted: 11 November 2016 * Published: 08 December 2016 * DOI:

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