Depth resolution limitation is a big problem of pure optical imaging techniques e.g. Optical Coherence Tomography (OCT). Photoacoustic imaging techniques gives the opportunity to overcome this problem. It is based on the photoacoustic effect, which describes the emission of thermoelastic pressure waves, generated by absorbers subsequent to short-pulsed optical excitation (Figure 1). The high contrast is caused by the wavelength-dependent optical absorption of chromophores inside a tissue.
After the pulsed light has reached the tissue surface, it is distributed depending on scattering and absorption. Due to absorption by the desired absorbing structure, its temperature increases followed by a rapidly rising pressure. Thus, the absorber expands thermoelastically and pressure transients are emitted. As a consequence of their low refraction and scattering, pressure waves can propagate large distances in tissue. This is a significant advantage in comparison to optical imaging techniques.
There are various methods to detect these pressure transients at the tissue surface. Commonly pressure measuring sensors e.g. piezoelectric transducers are used that require acoustic contact . However, in some imaging applications a sterile non-contact approach can be beneficial.
For this reason, a novel holographic off-axis detection method was developed. Compared to classical piezoelectric detection, the principal difference is that surface displacements induced by the pressure transient are measured in order to reconstruct the absorbing structures.
The first proof of concept of this novel detection method was published in . The main drawbacks of the used setup based on hardware limitations. Furthermore, simple tissue phantoms were used, where the optical properties have been neglected.
To overcome these drawbacks, a redesigned and enhanced setup with appropriate hardware components was developed and evaluated. In addition, a new type of tissue phantoms was developed, that takes the optical properties of dermal skin into account. One example will be presented as part of this papers.
2 Material and methods
To generate an emission of acoustic waves under stress confinement, the detection system is combined with the excitation system. It consists of an Nd:YAG laser (Edge-wave, BX60-2-G) which has a wavelength of 1064 nm and a pulse duration under 10 ns. The beam profile has a shape of a top-hat with an area size of 16 mm2. The beam is expanded to an area of 36 mm2. The radiant exposure of the excitation is close to the maximum permissible exposure of 20 mJ/cm2.
Figure 2 illustrates schematically the enhanced speckle based holographic off-axis detection method, which is generally based on a Mach-Zehnder interferometer. It is used to measure surface displacements of an observed tissue specimen.For the data acquisition over time, the light of an Nd:YAG laser (Newport Spectra Physics Explorer) with a wavelength of 532 nm is split into an object wave, to illuminate the specimen, and a reference wave. Mutual interfering of the backscattered object light results random intensity distribution called speckle pattern, which depends on the individual specimen topography. This speckle pattern is filtered using a spatial frequency filter and subsequently superimposed with the reference light, which incidents with a small angle greater 0° onto the CMOS-chip of a high-speed camera (Photron, SA3). If a point at the specimen surface experiences small axial deformations caused by photoacoustic pressure waves, the optical path of the signal channel alters and hence the measured phases of the speckle pattern change. In this context, the off-axis holography allows the measurement of the magnitude and direction of the phase differences, or rather displacements. Regarding the axial deformation, the detection has a sensitivity of ±1 nm. The lateral resolution is 35 µm and the temporal resolution 25 ns. This results in a sampling frequency of 40 MHz.
All setup components are controlled by a Counter/Timer-module (National Instruments NI PCI-6602) in combination with a specific developed software.
2.1 Repetitive measurement of surface displacements
To measure the varying surface displacement of the specimen over time, a novel triple pulse detection (Figure 4) was developed.
Basically, the recorded camera frames are split into two groups. Each recorded camera frame with an odd index is related to an interferogram of the undisturbed surface. Each frame with an even index is related to an inter-ferogram of the displaced surface, which is captured with a varying time delay of multiple t after the excitation pulse.
This repetitive procedure is carried out until the required maximum time delay corresponding to a certain imaging depth is reached. From each interferogram, the absolute object phase is calculated using the Fourier transformation. In order to obtain the phase difference images, the respective pairs of reference and deformation phase images are subtracted. This leads to a time-dependent displacement dataset. Thus, the measuring period is for example 400 ms at a physical repetition rate of 2 kHz for a maximum depth of 1 cm at a sampling frequency of 40 MHz.
One non-negligible problem is the random phase of non-speckle areas of the interferogram which leads to a noise with a high frequency that is superimposed with the displacement signal. To suppress such noise, the displace-ment dataset is filtered by a low pass filter. The choice of the cut-off frequency is made in compliance with the highest expected frequency of the displacement signal. Based on these data, the reconstruction is performed.
2.2 Reconstruction technique
The tomographic reconstructions are performed in three dimensional space and have been carried out using the delay and sum algorithm as introduced in Table 1, which is similar to the Synthetic Aperture Focusing Technique (SAFT) .
The result of the reconstruction algorithm is a probability distribution of the sources of photoacoustic signals. Details of the reconstruction are published in .
2.3 Measurement of tissue phantoms
The main material for the construction of tissue phantoms was a two-component transparent silicone (Wacker, Elastosil RT 604 A/B). To adjust the optical properties of the absorber surrounding silicon carrier matrix similar to dermal tissue, barium sulfate (BaSO4) was added in a specific concentration. This leads to an optical scattering coefficient µs close to human skin µs = 10 cm−1 at a wavelength of 1064 nm . To determine the resulting optical scattering coefficient, the adding doubling method was used . In addition to the scattering properties Barium sulfate was chosen because it is a heavy metal and therefor generates an X-ray contrast. This enables a co-evaluation of the reconstruction results with the three dimensional reconstruction dataset from a µCT (GE imagination at work, phoenix nanotom m, nanoCT®) which has a voxel resolution of 25 µm.
In addition to the optical scattering coefficient, the optical absorption coefficient µa plays a crucial role for the absorbing target structures. Caused by the purpose conditions it should be in the range of oxy-haemoglobin, µa = 3 cm−1 at 1064 nm , which is often used as contrast agent in human tissue. Preliminary investigations yield no quantifiable displacement with µa = 3 cm−1. For this reason, the value was increased to 20 cm−1 where conclusive results were measured.
To simulate a blood vessel, a thin silicone tube (Silastic® BioMedical Grade ETR Elastomer Q7-4750) with an inner diameter of 640 µm was used. The tube contains the liquid silicone (Wacker, Elastosil RT 604 A) absorber as a blood substitute. It was stained with black pigment paste in order to obtain the desired optical absorption coefficient. Figure 3 represents a developed tissue phantom schematically. Two tubes were positioned in an x-assembly at a depth from approximately 0.6 mm to 3 mm below the surface.
3 Results and discussion
Figure 5 presents the surface of the phantom at four different time points after optical excitation. The arising surface displacements and their temporal profile are clearly delineated. In Figure 5b), the surface deformation appears by the incident pressure wave of the upper tube. 1 µs later, the second surface deformation appears by the incoming pressure wave of the lower tube (Figure 5c)). Its amplitude is significantly weaker due to the attenuation during its propagation. The displacement signals migrate from its source to the edges of the detection area, where both remain clearly visible. The results demonstrate the high quality of developed detection approach.
The reconstruction results of the measured PAT data with a voxel resolution of 100 µm are illustrated in Figure 6a) and c). The tubes have been reconstructed only in the center of the field of view. This is a crucial disadvantage which based on the detector geometry and is known in literature as “Limited view problem” . Despite these circumstances, the shape of the tubes can be identified quite well. For the comparison with the µCT results (see Figure 6b) and d)) only the center region of the respective field of views and the center of the cross section of the tubes were analysed and summarised in Table 1.
The results show a great agreement of the measured depth positions. The slight differences can be explained by inaccuracies in the determination of the speed of sound, which has a significant influence on the reconstruction results of the PAT data.
In addition to the depth position, the angle of both tubes to each other can be calculated. The µCT and the PAT results obtain an angle of 85.7 and 84.6 degree. These values are very close to each other, which confirms the quality of the reconstructed PAT results.
4 Conclusion and outlook
The presented results could validate that non-contact displacement measurement is a suitable approach for photoacoustic detection. In addition, the reconstructed results agree well with the results determined by the µCT measurement.
Further investigations will be carried out towards a more detailed analysis of the potential and limitations of the approach.
This publication is a result of the ongoing research within the LUMEN research group, which is funded by the German Federal Ministry of Education and Research (BMBF, FKZ 13EZ1140A/B). LUMEN is a joint research project of Lübeck University of Applied Sciences and Universität zu Lübeck and represents an own branch of the Graduate School for Computing in Medicine and Life Sciences of Universität zu Lübeck.
Partially funded by the German Federal Ministry of Education and Research (BMBF) (FKZ 03FH015IN3).
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About the article
Published Online: 2015-09-12
Published in Print: 2015-09-01
Conflict of interest: Authors state no conflict of interest. Material and Methods: Informed consent: Informed consent has been obtained from all individuals included in this study. Ethical approval: The research related to human use has been complied with all the relevant national regulations, institutional policies and in accordance the tenets of the Helsinki Declaration, and has been approved by the authors’ institutional review board or equivalent committee.