Comparison Between Performances of In2O3 and In2TiO5-Based EIS Biosensors Using Post Plasma CF4 Treatment Applied in Glucose and Urea Sensing

In this study, the effect of post-deposition tetrafluoromethane (CF4) plasma treatment on the physical and electrical characteristics of an In2TiO5 based electrolyte-insulator-semiconductor (EIS) sensor was investigated. Post-deposition CF4 plasma treatment typically improved the crystalline structure and repaired dangling bonds at the grain boundaries. We used the newly fabricated device to detect several ions, such as sodium and potassium, which are essential for many biological processes. The as-deposited and CF4 plasma treated In2TiO5 sensing window with an EIS structure was also able to detect the pH of a solution, different alkali ions (Na+ and K+), glucose, and urea. The sensing membrane after a 60-sec CF4 plasma treatment displayed improved biosensing characteristics, such as higher sensitivity (59.64 mV/pH), better drift rate, and a smaller hysteresis voltage of approximately 0.424 mV/h. The In2TiO5 sensing membrane treated with CF4 plasma is a promising material for use in EIS biosensing applications.

Over the last few decades, major advances have been made in the field of biosensors for the monitoring and control of many biochemical activities. Biosensors are extensively used for monitoring food and soil quality, as well as human biochemistry and other medical applications 1 . Ion-sensitive field-effect transistors (ISFETs) are considered excellent transducers for sensing biochemical reactions due to their small size, fast response, and reliability. Electrolyte-insulator-semiconductor (EIS) materials operate with the same working principle as ISFETs, but their fabrication process is relatively simple and inexpensive as compared to that of ISFETs 2 . The surface potential of the exposed insulator sensing area changes in response to the H + concentration of the solution 3 . Therefore, selecting a proper insulating material with suitable characteristics is important to achieve high sensitivity and long-term reliability of the device.
Several metal oxides like indium-gallium-zinc oxide (InGaZnO, IGZO), zinc oxide (ZnO), and indium tin oxide (InSnO, ITO) have been developed as sensor membranes owing to their excellent sensing characteristics [4][5][6] . Indium oxide (In 2 O 3 ) thin films have attracted significant attention as a sensing membrane because of its higher mobility (160 cm 2 /(V·s)) 7 , higher melting point (1910 °C), larger band gap (3.5-3.7 eV) 8 , and insolubility in water. In 2 O 3 films can easily be reduced to create an oxygen deficiency, where insufficient oxygen atoms are contained in the crystal structure 9 . This results in the formation of a nonstoichiometric In 2 O 3−x film with an increased number of defects. Thus, the main free carriers in In 2 O 3−x are the internal oxygen vacancies of the film, which also affect the chemical stability of the film. As a potential solution, CF 4 plasma treatment of the In 2 O 3 and In 2 TiO 5 film can enhance the In-O bonding. In 2 O 3 is widely used for gas sensing 10 , transparent conducting oxides 11 , and thin film transistors 12 because of its excellent material characteristics.
The performance of a biosensor is conferred by doping with a transition metal, such as titanium (Ti). Ti affects the sensing membrane through the passivation of the defects in the bulk to the sensing performance and  www.nature.com/scientificreports www.nature.com/scientificreports/ production of dangling bonds at the oxide interface 13,14 . Ti internalization also enhances device capacitance and reduces the reactivity with the surrounding moisture 15,16 .
Recent evidence demonstrated CF 4 plasma treatment that improves the sensing performance of oxide materials for biosensing applications 17 . Fluorine incorporation improves the dielectric properties of the material by fixing the dangling bonds and replacing the weak bonds at the grain boundaries 18 . Fluorine introduces a net negative surface charge owing to its high electronegativity and improves the ability to capture positively charged H + ions 19 . Moreover, pre-and post-deposition CF 4 plasma treatment reduces metal-silicate formation at the oxide/ silicon (Si) interface 20 .
Here, we used a post-deposition CF 4 plasma treatment on In 2 O 3 and In 2 TiO 5 thin films to improve their electrical and physical characteristics. We describe the consequence of the post-deposition CF 4 plasma treatment on the sensing and physical characteristics of a deposited In 2 TiO 5 thin film as a sensing membrane. The detailed process flow of the EIS structure fabrication is shown in Fig. 1. After fabricating the device, sensing characteristics including sensitivity, hysteresis, and drift of the device were measured. To determine the physical characteristics, X-ray diffraction (XRD) atomic force microscopy (AFM), and secondary ion mass spectroscopy (SIMS) analyses were done.

Results and Discussion
physical characteristics. XRD was used to probe the crystalline structure of the materials. Figure 2(a,b) depicts XRD patterns of as-deposited and post-deposition CF 4 plasma treated In 2 O 3 and In 2 TiO 5 membranes, respectively. The diffraction angles (2θ) ranged from 20 to 60°. All the In 2 O 3 membrane samples displayed a cubic crystalline phase with two diffraction peaks, (222) and (321), at a diffraction angle of 30.5° and 33.10°, respectively 21 . A strong In 2 O 3 (321) peak was observed for the sample annealed at 60 sec. In 2 TiO 5 displayed a strong peak at (222) at a 2θ of 30.5. The major phase changed from (321) to (222) in XRD pattern of In 2 TiO 5 . The intensities of these three peaks were enhanced by the CF 4 plasma treatment and further increased by increasing www.nature.com/scientificreports www.nature.com/scientificreports/ the plasma treatment time from 15 to 60 sec due to the improvement in the crystalline structure. The crystallinity improvement arises from the formation of a stronger fluorinated bond between indium and oxygen 18 . Therefore, after CF 4 plasma treatment for 60 sec the In 2 O 3 and In 2 TiO 5 membranes would display higher intensity peaks than those prepared under other conditions. The XRD data indicated that internalization of Ti upgrades the thin film stoichiometry because it minimizes the number of dangling bonds on the dielectric surface and improves the temperature stability compared with In 2 O 3 thin film 22,23 .
AFM was used to study the surface morphology of the devices treated with post-deposition plasma. Figure 3(a-d) show two-dimensional (2D) AFM images of the as-deposited and post-deposition CF 4 plasma treated In 2 O 3 and In 2 TiO 5 samples. The AFM images of as-deposited In 2 O 3 and In 2 TiO 5 membranes revealed surface roughness of 0.557 nm and 0.628 nm, respectively. The roughness increased with In 2 O 3 and In 2 TiO 5 samples with increased CF 4 plasma treatment time from 15 to 60 sec, as shown in Fig. 3(e). Samples doped with Ti achieve better surface roughness compared to bare In 2 O 3 . Ti has a higher affinity toward oxygen, and thus provides a larger grain size. This produces a rougher surface 23,24 . CF 4 plasma treatment increases the interior grain size and causes plasma-induced morphological changes, which increase surface roughness and the number of surface sites, which in turn increase the sensitivity and linearity 25 .  www.nature.com/scientificreports www.nature.com/scientificreports/ Secondary ion mass spectrometry analysis data of the In 2 O 3 and In 2 TiO 5 thin films with and without CF 4 plasma treatment are shown in Fig. 4. The as-deposited sample displayed no trace of fluorine atoms. For all the samples with CF 4 plasma treatment, fluorine atoms were evenly distributed throughout the entire oxide bulk areas. Fluorine content increased as the CF 4 plasma treatment time increased from 15 to 60 sec. Ti-doped In 2 O 3 membranes have less excess etching of the sensing membrane. Fluorine incorporation reduces interfacial defect states at the oxide/silicon interface and increases the overall quality of the deposited oxide layer 26 . sensing characterization. The most important parameter of EIS structure is its flat-band voltage, which is defined as the voltage applied to produce a silicon surface potential of zero. The flat-band voltage of EIS is calculated as 27 : where E ref is the reference electrode potential with respect to vacuum; q 1 ф Si is the work function of Si, which is equal to 4.7 V; Ψ 0 is the potential drop in the electrolyte at the insulator-electrolyte interface; C i and Q i are the insulator capacitance and effective charge per unit area, respectively, χ Sol is the surface dipole potential of the solvent; and δ χ is the number of variations of χ potentials. The surface potential Ψ 0 changes by changing the pH of the solutions, which causes a change in flat-band voltage at the different pHs. The surface potential Ψ 0 is calculated as 25,28 : where pH pzc is the value of pH for which the oxide surface is electrically neutral and β is the parameter that depends on the sensitivity of the sensing surface. The site-binding model can also be explained using the equation for capacitance, Q = CV, where Q is the surface charge, C is the double layer capacitance at the insulator interface, and V is the surface potential (Ψ 0 ). The surface potential can be expressed as: www.nature.com/scientificreports www.nature.com/scientificreports/ where α is the dimension-less sensitivity parameter consisting of the differential double layer capacitance. The ability of the oxide surface to protonate and deprotonate can be represented by the symbol β. Figure 5(a-d) represent the capacitance versus substrate bias (C-V) curves of the In 2 O 3 and In 2 TiO 5 sensing membranes with and without CF 4 plasma treatment. The inset of the figures shows the pH sensing properties of the films extracted from the C-V curves. The as-deposited In 2 O 3 sample displayed a very low sensitivity of 21.67 mV/pH and linearity of 91.66%. The In 2 O 3 sample treated with CF 4 plasma for 15 sec displayed higher sensitivity and linearity of 47.03 mV/pH and 98.76%, respectively. The sensitivity and linearity increased to 52.43 mV/pH and 98.83% for samples treated for 30 sec. When samples were treated for 60 sec, the sensitivity and linearity further increased to 56.15 mV/pH and 98.18%, respectively. The as-deposited In 2 TiO 5 sample displayed a decent sensitivity of 36.34 mV/pH and linearity of 97.65%. For CF 4 plasma treatments of 15 and 30 sec, the sensitivity and linearity increased to 49.98 mV/pH and 94.23% at 15 sec, respectively, and 55.06 mV/pH and 99.67% at 30 sec, respectively. With a 60-sec CF 4 plasma treatment, In 2 TiO 5 samples displayed highest sensitivity and linearity values of 59.64 mV/pH and 99.68%, respectively. The CF 4 plasma treatment increased the overall sensing surface area by increasing the surface roughness. Thus, the larger sensing surface area will lead to higher sensitivity with more surface sites being present and capable of binding ions present in the electrolyte solution 18 . Therefore, pH sensing membrane treated with CF 4 plasma for 60 sec can possess high sensitivity and excellent linearity. These properties can be attributed to the formation of a greater number of fluorinated bonds (F-In bonds) on the In 2 O 3 and In 2 TiO 5 film surface due to CF 4 plasma treatment. The deviation in surface potential of the sensing surface after the surface is dipped into solutions with different pH values was determined by the C-V shift. H + and OH − attachment at the corresponding binding sites at the sensing surface can alter the surface potential 29 . The present results demonstrate that Ti doping increases the numbers of binding sites on the sensing surface. The grain size becomes larger with higher surface roughness due to incorporation of Ti. Correspondingly the number of binding sites increases with the larger surface area 23 . Therefore, the sensitivity and linearity of In 2 TiO 5 membranes that are plasma-treated for 60 sec is much higher than those aspects of In 2 O 3 membranes.
To investigate the hysteresis effects of the membrane, the samples were immersed in buffer solutions with different pH values in an alternating cycle (pH 7 → 4 → 7 → 10 → 7). Moreover, to test the long-term reliability of the devices, the drift effect of the In 2 O 3 with CF 4 plasma treated sensing membrane was measured by a C-V curve in a pH 7 buffer solution for 12 h. The hysteresis voltage and drift rate under the different CF 4 plasma treatment conditions reveal that the 60-sec In 2 O 3 film plasma treatment produced the lowest hysteresis voltage and lowest drift rate (4.33 mV and 0.95 mV/h, respectively ( Fig. 6(a-d)). In 2 TiO 5 CF 4 plasma-treated sensing membrane displayed a hysteresis voltage 25.75, 9.32, 6.85 and 2.72 mV in the as-deposited condition and following 15, 30, and 60 sec of annealing, respectively. Figure 6(d) shows the drift rate for In 2 TiO 5 devices with CF 4 plasma treatment.
The drift rate of the as-deposited sample was 9.23 mV/h, whereas drift rates of 4.92, 1.09, and 0.42 mV/h were www.nature.com/scientificreports www.nature.com/scientificreports/ obtained following annealing for 15, 30, and 60 sec, respectively. This was likely because fluorine compensated for the dangling bonds and replaced weaker bonds in the grain boundaries. Thus, the CF 4 plasma treatment can reduce the trap states in the oxide/silicon interface and improve the hysteresis and drift rate characteristics of the films 18 . The dangling bonds are negated by Ti ions and the defects located underneath of the dielectric surface are compensated 16,23 .
To analyse the sensing performance of our sensing membrane in the presence of potassium and sodium, we prepared a solution using a micropipette to control the concentrations of sodium and potassium ions ranging between 10 −5 and 10 −1 M by injecting 1 M NaCl/Tris-HCl and 1 M KCl/Tris-HCl into buffer electrolyte. Comparisons of H + , Na + , and K + sensing performances of the as-deposited In 2 O 3 and In 2 TiO 5 membrane, and the membrane treated with CF 4 plasma for 60 sec are presented in Fig. 7(a-d). In 2 TiO 5 sample treated with CF 4 plasma for 60 sec had best sensitivities (18.23 mV/pNa and 14.13 mV/pK) compared to the In 2 TiO 5 as-deposited sample (9.93 mV/pNa and 7.98 mV/pK) and 60-sec CF 4 plasma treated In 2 O 3 sample (13.77 mV/pNa and 12.78 mV/pK). For sodium and potassium sensing, considerably lower sensitivity was obtained compared to H + sensing. This can be explained by the heavier and larger Na + and K + ions compared to H + ions.
We also measured the essential human biochemistry data of glucose and urea. They were detected by incorporating appropriate enzymes on the sensing oxide surface. The chemical equations for the reactions that can be used for glucose detection are 30,31 : The glucose sensing properties of the In 2 TiO 5 sensing membrane on the EIS structure with and without the 60-sec CF 4 plasma treatment are presented in Fig. 8(a,b). The glucose concentration was controlled over the range of 2 to 7 mM. The as-deposited In 2 TiO 5 sensing membrane exhibited a low glucose sensitivity of 2.85 mV/mM and linearity of 85.23%. After Ti doping and 60-sec CF 4 plasma treatment, the sensitivity increased to 6.63 mV/mM and the linearity increased to 92.35%. Figure 8. Glucose sensing properties of (a) as-deposited In 2 TiO 5 membrane and (b) membrane treated with CF 4 plasma for 60 sec. www.nature.com/scientificreports/ The urea concentration was controlled in a range between 5 and 40 mM ( Fig. 9(a,b)). The sensitivity values of the In 2 TiO 5 sensing membrane above as-deposited and CF 4 plasma treatment in samples treated for 60 sec were 1.55 and 2.69 mV/mM, respectively. Therefore, the 60-sec CF 4 plasma treated In 2 TiO 5 sensing membrane has better sensitivity and linearity for the detection of urea. These results indicate that the CF 4 plasma treatment can increase the interior grain size and increase the number of surface sites, resulting in better sensing performance for the detection of H + , Na + , K + , glucose, and urea.
The test sample of commercially available glucose devices, such as the ACCU-CHECK glucometer, uses a precious metal (Ag/AgCl) 32 . In this study, we used In and Ti as the sensing film. Their use can reduce the purchase price and enable reuse of material. These advantages also include a sensing membrane with high glucose sensitivity and the capability to accurately and sensitively measure other molecules.  38 . The pH sensitivity, hysteresis, drift rate, glucose and urea sensing of the EIS device prepared with In 2 TiO 5 and incorporating a CF 4 plasma sensing membrane was superior. CF 4 incorporated into In 2 TiO 5 film can enhance the In-O bonding at the In 2 TiO 5 sensing membrane, which improves the sensing characteristics.   www.nature.com/scientificreports www.nature.com/scientificreports/ Conclusion A CF 4 plasma treated In 2 TiO 5 -based EIS sensor was fabricated for H + , Na + , K + , and glucose sensing applications. The effect of the post-deposition CF 4 plasma treatment on the physical and electrical characteristics of the sensors was studied. The In 2 TiO 5 EIS sensor with a 60-sec post-deposition plasma treatment exhibited good sensitivity of 59.64 mV/pH and linearity of 99.68%. Post-deposition CF 4 plasma treatment improves the crystalline structure and repairs the dangling bonds at the grain boundaries. Furthermore, the 60-sec plasma treatment produced the best material and electrical properties, and achieved ideal sensing capabilities, likely due to defect passivation by fluorine. Furthermore, the Ti-doped In 2 O 3 -based EIS sensor treated with CF 4 plasma was more responsive to H + compared to Na + and K + . The In 2 TiO 5 sensing membrane with 60-sec CF 4 plasma treatment also had better sensitivity and linearity than the as-deposited sample for glucose and urea detection.

Methods
The EIS incorporated In 2 O 3 and In 2 TiO 5 sensing membranes were fabricated on 4-inch n-type (100) silicon wafers with a resistivity of 5-10 Ω-cm. After a standard RCA cleaning process, the samples were dipped into a 1% hydrofluoric acid solution to remove native oxide from the surface. A 50 nm-thick SiO 2 substrate was thermally oxidized on the silicon wafer. Then, (a) a 50 nm-thick In 2 O 3 film was deposited on the SiO 2 /Si substrate by reactive radio frequency sputtering. (b) Both In 2 O 3 and Ti were used to co-sputter In 2 TiO 5 film on SiO 2 /Si stacks in a diluted argon flow (Ar/O 2 = 20 sccm/5 sccm) with a radio frequency power and process pressure of 100 W and 20 mTorr, respectively. After deposition, the In 2 O 3 and the In 2 TiO 5 films were subjected to a post-deposition CF 4 plasma treatment in a plasma-enhanced chemical vapor deposition system with a radio frequency power of 30 W and a pressure of 500 mTorr for 15, 30, and 60 sec. The back-contact was made by depositing a 300 nm-thick aluminium film on the back side of the Si wafer. The sensing area of the deposited In 2 O 3 films was defined by an automatic robot dispenser with an adhesive silicone gel to build the final EIS structure on the printed circuit board using a silver gel to form conductive lines. An epoxy package was used to separate the EIS structure and the copper line. The detailed process flow of the EIS structure fabrication is shown in Fig. 1.
Morphological analyses of the In 2 O 3 and In 2 TiO 5 sensing membranes included XRD, AFM, and SIMS. These analyses were done to investigate the link between structural and electrical characteristics properties. SIMS instruments use an internally yielded beam of either positive or negative ions focused on a sample surface to generate secondary ions. The generated ions are then transferred into a mass spectrometer across a high electrostatic potential. The depth profiling of elemental and molecular species, as well as isotopic ratios of compounds was evaluated by SIMS analysis. The surface morphologies of the In 2 O 3 and In 2 TiO 5 nano-layers were observed using AFM in Bruker Dimension Icon modes with intermittent contact using a silicon tip with a 10 pN/nm spring constant. A sample area of 3 × 3 μm was scanned with actuation rates up to 8 kHz in air and fluid. For XRD analysis of the samples, grazing incidence of X-ray beam CuKa (k = 1.542 Å) radiation was used with an incidence angle step of 0.5° in the diffraction angle range (2θ) from 20° to 60°.