Shigeki Kuroiwa

Fabrication of stable antibody-modified field effect transistors using electrical activation of Schiff base cross-linkages for tumor marker detection

In this paper, we present a method of fabricating a rigid antibody-immobilized surface using electric activation of a glutaraldehyde (GA)-modified aminopropylsilyl surface for stable antibody-modified field effect transistors (FETs). Electric activation of the GA-modified gate surface of the FET reduces Schiff bases, which are easily hydrolyzed and collapsed, formed between GA and 3-aminopropyltriethoxysilane, resulting in preventing the immobilized antibodies from desorbing from the surface. The lack of Raman peaks that could be assigned to a Schiff base after the electrical activation of the GA-modified surface indicated that the electric activation had reduced the Schiff base. The use of the antibody-modified FETs has three advantages for the detection of antigens: increased sensitivity, distinct recognition ability, and improved reproducibility. A tumor marker, alpha-fetoprotein (AFP), was quantitatively detected up to a concentration of 10 ng/mL using the antibody-modified FET. The detection ability of the FET accomplished a cutoff value of hepatic cancer. The quantitative detection of AFP in a solution with contaminating proteins was also demonstrated. This electric activation method is applicable to other antibody-modified FETs.

Sensitive electrical detection of human prion proteins using field effect transistor biosensor with dual-ligand binding amplification

Simple and accurate detection of prion proteins in biological samples is of utmost importance in recent years. In this study, we developed a label-free electrical detection-based field effect transistor (FET) biosensor using thiamine as a probe molecule for a non-invasive and specific test of human prion protein detection. We found that thiamine-immobilized FETs can be used to observe the prion protein oligomer, and might be a significant test for the early diagnosis of prion-related diseases. The thiamine-immobilized FET was also demonstrated for the detection of prion proteins in blood serum without any complex pre-treatments. Furthermore, we designed a dual-ligand binding approach by the addition of metal ions as a second ligand to bind with the adsorbed prion protein on the thiamine-immobilized surface. When the prion attached to metal ions, the additional positive charge was induced on the gate surface of the FET. This approach was capable of amplifying the magnitude of the FET response and of enhancing the sensitivity of the FET biosensor. Detection of prion proteins has achieved the required concentration for clinical diagnosis in blood serum, which is less than 2 nM. In summary, this FET biosensor was successfully applied to prion detection and proved useful as a simple, fast, sensitive and low-cost method towards a mass-scale and routine blood screening-based test.

Field Effect Transistor Biosensor Using Antigen Binding Fragment for Detecting Tumor Marker in Human Serum

Detection of tumor markers is important for cancer diagnosis. Field-effect transistors (FETs) are a promising method for the label-free detection of trace amounts of biomolecules. However, detection of electrically charged proteins using antibody-immobilized FETs is limited by ionic screening by the large probe molecules adsorbed to the transistor gate surface, reducing sensor responsiveness. Here, we investigated the effect of probe molecule size on the detection of a tumor marker, α-fetoprotein (AFP) using a FET biosensor. We demonstrated that the small receptor antigen binding fragment (Fab), immobilized on a sensing surface as small as 2–3 nm, offers a higher degree of sensitivity and a wider concentration range (100 pg/mL–1 μg/mL) for the FET detection of AFP in buffer solution, compared to the whole antibody. Therefore, the use of a small Fab probe molecule instead of a whole antibody is shown to be effective for improving the sensitivity of AFP detection in FET biosensors. Furthermore, we also demonstrated that a Fab-immobilized FET subjected to a blocking treatment, to avoid non-specific interactions, could sensitively and selectively detect AFP in human serum.

A label-free electrical assay of fibrous amyloid β based on semiconductor biosensing

We propose, as an alternative to conventional spectroscopic assays, a simple method for discriminating fibrous amyloid proteins by using a label-free semiconductor-based biosensor. The highly sensitive assay is expected to be useful for accelerating amyloid related research.

Label-free detection of Cu(II) in a human serum sample by using a prion protein-immobilized FET sensor

We have developed a field effect transistor (FET) sensor to sensitively detect copper ions (Cu(2+)) in a human serum (HS) sample for promising health-care diagnosis. By utilizing a Cu(2+)-binding prion protein that was immobilized on the FET gate surface, such an FET sensor can provide a simple, label free and highly selective performance, even in HS samples. We demonstrated the sensitivity of the sensor at the nanomolar level, 0-100 nM, which is very useful for the detection range of Cu(2+) deficiency in practical applications.

Effect of Surface Morphology of Reference Field Effect Transistor Modified by Octadecyltrimethoxysilane on Ionic Responses

For completion of on-chip field effect transistor (FET) biosensors, it is essential to produce a reference FET instead of a reference electrode that uses an inner electrolyte. In this study, a light-addressable potentiometric sensor (LAPS) modified. by a self-assembled monolayer (SAM) of octadecylsilane (ODS) was pH-insensitive (-1.5 mV/pH), but it was sensitive to ionic strength as was reported in polymer-gate FETs. Our purpose was to make clear the problems of the evaluation methods of ODS-SAM FETs concerning pH and ionic responses and the effect of the surface morphology to improve a reference SAM-FET. An ODS-SAM LAPS and FET were deposited under different conditions. pH and ionic responses, roughness measured by atomic force microscope, and contact angle (CA) were greatly changed by the difference in morphology of the ODS-SAM. ODS coagulation raised the CA. It is difficult to distinguish the ODS-SAM with and without ODS coagulation only by CA, but CA is utilizable for observing the state of hydroxyl groups on the surface without ODS coagulation. ODS coagulation caused the pH response with the ODS-SAM but decreased the ionic response of the LAPS. Controlling the surface morphology of the ODS-SAM is important to suppress the pH and ionic sensitivities for an in vivo applicable reference FET.