Photoacoustic Imaging using Diode Laser for Soft Tissue Visualization

This study aimed to develop a photoacoustic imaging system using a diode laser as a radiation source, and a condenser microphone as a soft-tissue image detector. These tools were set in a static position and combined with the mechanic component to move the object table in the X-Y direction and were controlled by a computer with a LabView program. The result showed the built photoacoustic imaging system was used to analyze soft tissue using a diode laser in the frequency range of over 15 kHz with a 24%-47% duty cycle modulation. Furthermore, the minimum shift of the mechanic component X-Y stage was 0.02 mm, the sampling data rate was 1 second/pixel, and the image showed soft tissue intensity is generally higher than non-biological materials.


Introduction
Photoacoustic imaging (PAI) is a non-invasive optical and acoustic imaging technique that provides high-resolution optical images of biological tissue [1].The development of PAI systems is a significant modality in biomedical optics and has grabbed the interest of numerous research communities over the past decade, particularly in the field of biomedical imaging [2].
In 1880, Alexander Graham Bell discovered the photo-acoustic effect.When solid materials are exposed to a pulsed light source, Bell discovered that sound waves can be generated.In addition, Theodore Bowen was the first individual to propose the phenomenon of imaging soft tissue using photoacoustic technology in 1981.In 1993, an article reporting one of the first in -vivo photoacoustic experiments performed on the human finger was published as a result of this preliminary research [2].Since then, significant progress has been made in instrumentation, image reconstruction algorithms, functional imaging, and molecular imaging capabilities.
The photoacoustic effect has been applied to an imaging technique known as the photoacoustic imaging (PAI) system.Generally, this system uses an ultrasonic transducer as a signal detector combined with a Q-switched Nd YAG laser as the radiation source [3], which is complex, expensive, and time-consuming due to the pumping process.Therefore, researches are looking for alternative radiation sources as well as an acoustic detector to develop a simpler PAI system.
The photoacoustic imaging used a diode laser as the radiation source combined with a condenser microphone as a detector.The laser is inexpensive, able to produce a high-frequency signal, and spontaneously emits light with pumping.Furthermore, the condenser microphone can detect acoustic signals with frequencies up to 20 kHz.High-frequency detection is needed in the PAI system to avoid background noise, which is generally detected below 15 kHz.Therefore, to produce a high -quality image, the signals were set above 15 kHz.This is different from a closed-cell system, which is usually used for gas detection that places the sample inside the photoacoustic cell [4].However, the PAI in this study was an open cell.The image was reconstructed from the signal using LabView software, and the contrast was determined by the intensity [5].The ability of the system has been tested to distinguish the image of the rat's soft tissue of the tongue from the surrounding image of plasticine.This proves the system has the potential to be used as an early detection tool for oral cancer.Because about 95% of all oral cancers are squamous cell carcinomas (cancers originating from the epidermal layer), most oral diseases will be diagnosed from lesions on the mucosal surface [6].This means that oral cancer is only on the surface [7], making it possible to detect it even at an early stage [8].The imaging technique is one of the methods in screening [9].Screening methods can help detect oral cancer also though no symptoms have appeared.To enable early diagnosis and treatment of patients with oral cancer [10].Therefore, an open cell photoacoustic system was developed using a low-cost system, condenser microphone, and laser diode, in order to generate highquality acoustic images from soft tissue.

The Photoacoustic Signal Detection
The waves were generated by the thermoelastic effect with the adiabatic process when a short laser pulse was absorbed by the medium [11].When they interact, a small amount of energy is absorbed and converted to heat.Also, thermal expansion generates a pressure wave that spread on the medium [12].The diode laser-induced photoacoustic signal generation process is described in   The signal production mechanism starts from laser pulses generation, and light distribution in tissues can be approximated by exponential decay, which depends on depth ( > 0) and the laser radial width ().The light intensity distribution [12] can be written as follow (, ) ≅  0 () (−  )  (1)  0 in equation ( 1) is the surface intensity,   is an effective attenuation coefficient, and (, ) is the intensity in a cylindrical coordinate.
In tissue, the light was absorbed, and some were scattered.In the three-dimensional coordinate space, temperature spread [12] can be approximated by the equation ( 2) and is the heat capacity at constant volume,  is laser energy flux unity extents,   is the sample absorption coefficient, and  is sample density.Pressure wave equations that periodically produce heat [13] can be expressed as (, ) is heat function, which is defined as the energy supplied by electromagnetic radiation per unit time and volume.The acoustic waves then propagate by gas volume or ambient air to the microphone [14], which was obtained by recording the waves in the function of time.
This recording was done using the ECM Behringer 8000 condenser microphone connected to the audio soundcard.Furthermore, voltage conversion resulting from capacitance changes due to the pressure induced by the waves and was detected by the transducer in a condenser microphone.This process is described in Figure.The total charge () accumulated in the diaphragm and the backplate [15] can be written as follows  =  0  0 (5) () is resistor,  0 is diaphragm capacitance with backplate,  0 is the voltage (V).The output voltage () in the time function generated from the condenser microphone in DC polarizing method [15] is () is capacitance changes due to the distance between the diaphragm and a backplate.The signal can then be amplified using an audio soundcard as an amplifier.

The PAI Mechanic System
In producing a photoacoustic image, a data sequence has to be reconstructed from an X-Y direction for 2D, or an XYZ for 3D image [16].Assuming the surface scattering and the laser pulse energy variations are equal at every point on tissue samples [17], the scan point data were obtained from the signal intensity, which is captured at a frequency range (or wavelength) ideally located at the pulsation interval or modulation [17].Furthermore, intensity data retrieval was performed at the same frequency as the laser.
The scan method generally uses cartesian or polar coordinates [18].However, this study used a Cartesian system in the scanning process, and the sample on the table was moved in X-Y direction driven by NEMA 17 stepper motor connected with a driver TB6560, which is controlled by the microcontroller.

The Photoacoustic Image Reconstruction
In the scanning process, the PAI system search for photoacoustic intensity peak at every scan point.This peak was detected from the signal resulting from the Fourier transform of the acoustic pressure wave in the form of time function array (s) to frequency (Hz).The Fourier transform was done using a digital computer, in the form of discrete equations [19] which is being written as: is the frequency with index , ( +1 −   ) is a sampling interval,  is a lot of sampling data from the function (  ) to know the amplitudes of each forming sinusoidal wave (  ), then the time required to calculate it is proportional to  2 [19].
The intensity was stored in a 2D with an array resolution that was determined by the number of X and Y points.This is represented as the contrast in the image reconstruction process, and when the intensity has more differences, it produces a high contrast image and vice versa.

PAI Mechanic Component
The mechanic component is connected to a microcontroller.It is controlled using an interface program on the computer.The PAI system used in this study is identical to the system used in previous research [20], which includes a green laser with a wavelength of 532 nm as the light source and a condenser microphone as the acoustic signal detector.The PAI system was mounted perpendicularly between the X and Y axis for the sample table to be shifted in two directions in the scanning process.
During the scanning process, the component moves in a repeated S line direction, and all the systems are controlled by an integrated interface on the computer, i.e., the LabView program.The rotational motion of the motor screw driven by stepper was converted into a translational (motion manipulator) using a ball screw, also called a linear actuator.Inside the screw component, there should be no backlash delay, as it causes reduced accuracy [21,22].The accuracy of the X-Y stage has been measured, and Figure.3 shows the object shift needs 0.02 mm for one step.This is important in the scanning process for image resolution reconstruction.

Photoacoustic Signal Detection
The signal recorded by the microphone was still in time function and needs to be transformed into a frequency function of the signal peak intensity.This transformation should be done using a digital computing process (Eq.7).Thus, the intensity frequency was corresponding with that of diode laser modulation, and the signal acquired from FFT is shown in Figure . 4.
Figure .4 showed a photoacoustic intensity when the material was subjected to energy in the form of non-stationary or modulated laser light.It also shows that at low frequencies, there are signals with high intensity called background noise.This is usually in the range of 0-6 kHz, but it was determined in this study at less than 15 kHz.The photoacoustic signal intensity had the same frequency as that of laser modulation.However, when this modulation frequency is changed, that of the photoacoustic signal will also change, as shown in

Figure.4.
A diode laser is a major component in the PAI system, and its beam produces energy in the form of radiation to be absorbed by the tissue or material.Therefore, the power emitted (Js -1 ) should be known.Figure.5 shows the laser power is not linear to the duty cycle modulation, which is the percentage of the on-off phase in one period.The built PAI used an open-cell system for the energy to be absorbed by gases in the air before heading to the sample.Also, a laser with low modulation produces lower power because almost all is absorbed by air gases.Based on the results in Figure .5, the optimal duty cycle value for modulation in the PAI system is 20-60%.By increasing the duty cycle, the laser output energy will also increase, resulting in a greater number of interactions between specific chromophores within the tissue.The large number of interactions that occur can increase the acoustic signal's intensity.To determine the optimum laser modulation, which regarding soft tissue, the image was produced using 16-47% duty cycle, as shown in Figure .6, a low cycle was unable to produce an optimal image (Figure .6 b).Also, the soft tissue can be clearly differentiated from non-biological material (plasticine) in photoacoustic images produced using 16-47% duty cycle.In Figure 6 c, soft tissue and non-biomedical materials are recognizable but the image is ineffective.At duty cycles between 32% (Figure 6 d) and 47% (Figure 6 f), the image quality is acceptable, but there is no noticeable difference.This indicates that the maximal interaction results that occur during the duty cycle range (30% to 50%) are displayed.Therefore, if the duty cycle is increased further within this range, there will be no image difference.In addition, the sample tends to burn when irradiated by a modulated diode laser with a duty cycle of more than 50%.The sample will burn because it has attained the maximum amount of energy that sample can tolerate.

Photoacoustic Image Reconstruction
The image is reconstructed from the signal intensity, which is generated by the sample plotted in a 2D array.Hence, the contrast was determined by the intensity differences.The photoacoustic signal in high intensity was plotted in red color, medium in yellow, and low in blue.This color order was used to facilitate the visual distinction of the image.To obtain the soft tissue image, the sample was placed in a plasticine, and then scanned with the PAI system.Figure.8 shows the plasticine with soft tissue has higher intensity.The imaging system was able to detect the difference between the tissue and non-biological materials.Nevertheless, there is noise in the form of dots in the image, which is related to less laser stabilization and acoustic background outside the system.Furthermore, the system needs to be developed both in hardware and software components for noise to be reduced, as well as improve the image quality.

Conclusion
A photoacoustic imaging system was built, which has a maximum laser power of 144mW and a minimum X-Y stage shift of 0.0200 ± 0.0003 mm.The system was capable of producing a minimum image scale of 1 × 1 mm 2 , with a sampling data rate of 1 second/pixel.Therefore, it takes 100 seconds to produce an

2 .
The voltage was also used in time sampling on the LabView program.

Figure. 2 .
Figure. 2. The voltage conversion circuits in a condenser microphone.

Figure. 3 .
Figure. 3. The calibration factor of the motor stepper has a constant shift with error correction of 0.0006 mm.3.2Photoacoustic Signal Detection

Figure. 4 .
The FFT (Fast Fourier Transform) feature changes the acoustic signal in the time function (a) into the acoustic signal in the frequency function (b).

Figure. 5 .
Figure. 5.The effect of the duty cycle in laser modulation to the laser output power.

14th
International Symposium on Modern Optics and Its Applications Journal of Physics: Conference Series 2696 (2024) 012016 The reconstruction and interpolation program was made in the block diagram inside the LabView panel, as shown in Figure.7.

Figure. 7 .
Figure. 7. Block diagram of photoacoustic image reconstruction and interpolation in LabView program.

Figure. 8 .
Soft tissues as a sample in this study (a) and the photoacoustic image (b) produced from the sample.The tissue area showed a high intensity that contrasts with non-biological material surrounding them.
14th International Symposium on Modern Optics and Its Applications Journal of Physics: Conference Series 2696 (2024) 012016