Development of a blood oxygenation phantom for photoacoustic tomography combined with online pO2 detection and flow spectrometry

Abstract

Photoacoustic tomography (PAT) is intrinsically sensitive to blood oxygen saturation (sO2) in vivo. However, making accurate sO2 measurements without knowledge of tissue- and instrumentation-related correction factors is extremely challenging. We have developed a low-cost flow phantom to facilitate validation of PAT systems. The phantom is composed of a flow circuit of tubing partially embedded within a tissue-mimicking material, with independent sensors providing online monitoring of the optical absorption spectrum and partial pressure of oxygen in the tube. We first test the flow phantom using two small molecule dyes that are frequently used for photoacoustic imaging: methylene blue and indocyanine green. We then demonstrate the potential of the phantom for evaluating sO2 using chemical oxygenation and deoxygenation of blood in the circuit. Using this dynamic assessment of the photoacoustic sO2 measurement in phantoms in relation to a ground truth, we explore the influence of multispectral processing and spectral coloring on accurate assessment of sO2. Future studies could exploit this low-cost dynamic flow phantom to validate fluence correction algorithms and explore additional blood parameters such as pH and also absorptive and other properties of different fluids.

Keywords: blood oxygenation; flow; phantom; photoacoustic tomography.

Fig. 1

Overview of the flow system. (1) Injection site for introducing oxygenated blood (or other fluids) into the flow system, and for subsequently deoxygenating the blood using sodium hydrosulfite delivered via the syringe driver (MKCB2159V, Harvard); (2) online spectra are recorded using a light source (Avalight-HAL-S-Mini, Avantes) and spectrometer (AvaSpec-ULS2048-USB2-VA-50, Avantes) as the blood passes through a flow cell (170700-0.5-40, Hellma Analytics); (3) needle probes (NX-BF/O/E, Oxford Optronix) measure the temperature and partial pressure of oxygen (${\mathrm{pO}}_2$) before (3a) and after (3b) the blood passes through an agar phantom immersed in the photoacoustic imaging system (MSOT inVision 256-TF, iThera Medical) (4); (5) a touch-screen monitor (OxyLite Pro, Oxford Optronix) displays temperature and oxygen data; (6) these data are downloaded via an Arduino UNO and read in MATLAB on a laptop, which also records the spectrometer readings via AvaSoft software; (7) a peristaltic pump (CTP100, Fisher Scientific) provides blood circulation.

Fig. 2

Assessment of the effect of tube type on the quality of photoacoustic images. (a) PAT images (at 775 nm) of four tubes with inner and outer diameters (I.D.-O.D.) in $\mu \mathrm{m}$ of 630 to 1190 (silicone), 667 to 1000 (PMMA), 1500 to 2100 (PVC), and 1570 to 2410 (silicone), filled with a solution of ICG and embedded within a scattering agar cylinder (not shown). (b) Mean photoacoustic signal intensity inside the tube relative to the signal outside (SBR) for 11 different tubes. The tube with I.D.-O.D. 1500 to $2100\mu \mathrm{m}$ was selected due to its high SBR, flexibility, and low cost. The four tubes illustrated in (a) are marked with symbols.

Fig. 3

Comparison of dye spectra measured using PAT and online flow spectrometry under closed conditions. (a), (b) PA signal intensities (with smoothed spline) and (c), (d) online flow spectrometer absorbance values measured for different concentrations of MB [panels (a) and (c)] and ICG [panels (b) and (d)]. Gray box indicates wavelengths outside the PAT spectral range.

Fig. 4

Comparison of endmember spectra used for spectral unmixing. (a), (b) Literature spectra, spectra acquired using (c), (d) the online flow spectrometer and (e), (f) the offline flow spectrometer for MB [panels (a), (c), and (e)] and ICG [panels (b), (d), and (f)].

Fig. 5

Spectrally unmixed PA signal intensities for a range of dye concentrations. (a) MB and (b) ICG concentrations obtained by unmixing with literature spectra and those measured using the online and offline spectrometers. The LM was used to calculate concentrations of (c) MB and (d) ICG during evolution of dye concentrations within the flow circuit due to continual dye injection. The starting points were forced to zero for ease of comparison.

Fig. 6

Assessment of blood oxygenation within the flow circuit under closed conditions. Blood oxygen saturation (${\mathrm{sO}}_2$) was calculated using the three independent methods: the ${\mathrm{pO}}_2$ probe measurements [using Eq. (2)]; unmixing of the spectra measured using the online flow spectrometer (with experimentally measured spectra as end-members); unmixing of the mean pixel intensities in the PA images (with literature spectra as end-members) acquired while blood circulated in the flow system for 200 s.

Fig. 7

Dynamic deoxygenation of the circulating blood. (a) Change in blood oxygen saturation (${\mathrm{sO}}_2$) measured using the ${\mathrm{pO}}_2$ probe, online spectrometer, and PAT system while injecting 3% w/v sodium hydrosulfite in PBS over a period of 9 min. The anomalous data for the online spectrometer around 100 s may be due to a small bubble passing through the circuit. (b) Literature spectra for oxy- and deoxyhemoglobin used for spectral unmixing, compared to spectra obtained from the online spectrometer at the start and end of dynamic deoxygenation.

Fig. 8

Investigation of spectral coloring. (a) Spatial profiles of blood oxygen saturation (${\mathrm{sO}}_2$) as a function of distance from the center of the flow circuit tube by PA unmixing with spectra measured using the online spectrometer during dynamic blood oxygenation (Fig. 7). (b) Impact of correction for the absorption spectrum of the nigrosin dye included in the background of the tissue-mimicking phantom as a function of dye concentration. For absorption coefficients of $>0{\mathrm{cm}}^{-1}$, the ${\mathrm{sO}}_2$ (gray bars) deviated from the ground truth value (dotted line); accurate values were restored by recalculating ${\mathrm{sO}}_2$ (orange bars) after dividing the photoacoustic images by the relevant nigrosin spectrum as described in the text.