# Surface Acoustic Wave Sensor for C-Reactive Protein Detection
## 3 Results and Discussion
[40] reported that the mass response was only a small fraction of the total velocity shift response for a polymer-coated SAW device. In addition, [PERSON] et al. [34] reported that the resonance frequency deviates from its stable value if the surface liquid residues are not fully removed. The final possible reason is that the frequency shift is too small to be detectable in our measurement, because a very small frequency shift was observed at a very high CRP concentration (1 mg/mL). Figure 6 demonstrates the correlation of amplitude attenuation and CRP concentrations of SAW sensors. It was clearly observed that the CRP concentration of 0.1 \(\upmu\)g/mL can be detected in the proposed SAW sensor. [PERSON] et al. reported that the median CRP value of 1.92 \(\upmu\)g/mL (interquartile range 0.97-3.41) was measured [20]. In the proposed SAW sensor, the detectable CRP concentration of 0.1 \(\upmu\)g/mL is high enough for clinical diagnosis because the clinically relevant range for detecting CRP concentrations is generally from 1 to 200 \(\upmu\)g/mL [18]. The CRP sensitivity of a proposed SAW sensor should at least be 0.1 \(\upmu\)g/mL based on the three trials for each concentration. A good linear relationship between the detected insertion loss (amplitude) and the concentrations of CRP from 0.1 \(\upmu\)g/mL to 1 mg/mL was observed. The dark points in Figure 6 show the mean value of different amplitude (dB) for each CRP concentration. It was observed that the amplitude decreases with increasing CRP concentration due to the mass-loading effect. The mean value for each concentration can be fitted by a linear regression line. This regression line is useful to estimate the CRP concentration by using the proposed SAW sensor. Figure 7 showed a large variation in the three measurements for each CRP concentration. One possible reason for a large variation is that the surface liquid residue is not completely dried by blowing a small amount of nitrogen gas for 30 s. It is known that it is difficult to control the same treatment for every experiment. It was noted in this study that the amplitude response of SAW sensor was severely attenuated due to the radiation of the acoustic energy into the liquid if the surface liquid residues were not completely removed. Accordingly, a small amount of liquid residue variation may cause considerable attenuation changes. We were unable to perform real-time detection for the proposed SAW sensor. Therefore, we could not estimate the response time for the proposed SAW sensor. Real-time measurement and the estimation of the response time can be made if we change the substrate  
Figure 6: The correlation of amplitude attenuation and CRP concentrations of SAW sensors. of 128-degree Y-cut LiNbO\({}_{3}\) to the 64-degree Y-cut LiNbO\({}_{3}\), owing to the in-plane polarization of the wave (love wave). The detailed characterization of the biomolecular membrane modification during the preparation and the following CRP/anti-CRP interactions were also examined, as shown in Figure 8. It is worth noting that wafer-drying had been done before all S-parameter measurements. The insertion loss of \(-\)33.24 dB was observed in the bare SAW sensor with an Au-coated layer on top of SiO\({}_{2}\) guiding layer. It decreased to \(-\)52.67 dB after the Au film was immobilized with the 11-MUA containing thiol groups. After activation (EDC/NHS) and the antibody fixation (anti-CRP antibodies) on the self-assembled monolayer (SAM) film, we found that the value of the insertion loss was attenuated to \(-\)54.46 dB, as the anti-CRP was linked to the Au surface. A new equilibrium plateau of \(-\)42.54 dB was achieved as the SAW sensor was immersed in Ethanolamine-HCl solution and cleaned by PBS. Several factors, such as the improper pH value of the PBS or the residual charge and extension of the polymer brushes, may be responsible for the observation of an apparent increase in the measured insertion loss as the deactivation step was introduced [41; 42].