Polyacrylamide-based phantoms of human skin for fluorescence optical measurements
Current methods of optical medical diagnosis, in particular fluorescence analysis methods, allow for a high degree of accuracy in differentiating between areas of tumor transformations. Fluorescence imaging demonstrates considerable sensitivity in detecting the presence of pathological changes, including malignant changes, in biological tissues. However, the calibration and verification of such imaging systems requires the development of optical phantoms of biological tissues with known and quantified optical and mechanical properties. Thus, the study proposes the idea of manufacturing a solid matrix phantom base using polyacrylamide (PAA) as a bonding agent with stable thermal and chemical characteristics and gelatin to reproduce the fluorescent properties of collagen. Protoporphyrin IX, contained in human biotissue, is used as a fluorescent agent.
The phantom matrix base was a homogeneous solution obtained by mixing 0.2 g of powdered gelatin in 20 ml of distilled water at 40 °C. To give phantom elasticity, 6 g of acrylamide (AA) and 0.16 g of bisacrylamide (BAA) were added to the gelatin solution, followed by the addition of ammonium persulfate and TEMED, initiating a polymerization reaction to form a gel, a structure in which polyacrylamide chains are cross-linked by bisacrylamide molecules. To simulate the scattering properties of biotissues, 0.03 g of dispersed ZnO, with actively scattering properties, was added to the manufactured polymer structure. To simulate the fluorescence of biotissue, protoporphyrin IX powder at a concentration of 150 µmol/L was added to the manufactured structure.
The spectral characteristics of the developed phantom were determined using a spectrophotometer with an integrating sphere (Shimadzu, Japan). The absorption coefficient (μ_a) and reduced scattering coefficient (〖μ'〗_s) in the range 400-1000 nm were calculated using the inverse addition-doubling method for the fabricated solid base with added fluorophore. The fluorescence properties of the developed phantom were determined using the developed experimental hyperspectral fluorescence imaging system. This system includes a 450 nm M450LP1 LED light source (Thorlabs, USA) with an MF445-45 bandpass filter (Thorlabs, USA), an MD416 dichroic mirror (Thorlabs, USA), a hyperspectral camera (SPECIM Spectral Imaging Ltd, Finland) with a 500 nm FELH0500 cut filter (Thorlabs, USA).
The obtained results confirm the optical characteristics of the developed optical phantom with the addition of protoporphyrin IX, whose values are close to the real values in human tissues. The main advantage of the manufactured optical phantoms is that the polymerization conditions do not affect the fluorescence properties of the added fluorophores. The resulting elastic phantom matrix satisfies the stationarity condition, which is a necessary requirement for calibration measurements. The application of the developed phantom technology will make it possible to test, standardize, and calibrate fluorescence imaging systems. As further work, the reproduction of tumor transformations in tissues, by varying concentrations of protoporphyrin IX, is considered.
The work was supported by the Russian Science Foundation (the research project 20-75-00123). A.D. and I.M. thank the Academy of Finland for the support of Project №326204. V.D. kindly acknowledges personal support from the European Union’s Horizon 2020 research and innovation program under the Marie Skłodowska-Curie grant agreement No. 839888.
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Shupletsov Valery Vitalevich
Research & Development Center of Biomedical Photonics, Orel State University, Orel, Russia
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