Teruko Yuhi

Gold nanoparticle-based novel enhancement method for the development of highly sensitive immunochromatographic test strips

Abstract The immunochromatographic assay that is in widespread use for pregnancy diagnosis is a method for easy visual judging of the antigen–antibody reaction using gold nanoparticle. The rapid observation of results directly by the naked eye ensures the convenience of performing bioassays on-field. Therefore, gold nanoparticle-based immunochromatographic assays have provided attractive means for developing biosensors without the handling of toxic reagents, while allowing an easy and rapid procedure. However, the detection limit of this method is higher than the conventional method, enzyme-linked immunosorbent assay (ELISA). In this report, we developed a highly sensitive immunochromatographic assay for the detection of human chorionic gonadotropin hormone (hCG) as the model case. In this research, we are reporting the application of a new ‘sensitizer’ that contains gold nanoparticle conjugated primary antibody and the antigen. The sensitizer was added to the membrane after finishing the application of the normal method. The antigen of the sensitizer was captured by the secondary antibodies at the test line on the strip. As a result, the accumulation of the gold nanoparticle increased at the test line, and the sensitivity was higher. The sensitivity of our method could be enhanced by the sensitizer to almost the same level of ELISA assay. The test line intensity of hCG at 25 pg/ml treated with sensitizer was almost equal to the density that we observed at 1.0 ng/ml with the normal method. We also tested the performance of the sensitizer by using the surface plasmon resonance (SPR) technology of BIACORETM. The sensitizer using immunochromatographic assay is a promising candidate for decentralized diagnosis in clinically important fields such as the sensitive detection of cancer markers.

Gold nanoparticle based immunochromatography using a resin modified micropipette tip for rapid and simple detection of human chorionic gonadotropin hormone and prostate-specific antigen

Abstract A novel bioanalysis system based on immunochromatography was developed in connection with a nitrocellulose resin-modified micropipette tip, namely as ZipTip®. The sandwich-type immunoassay was applied to our bioananalysis system. The first antibodies that were aspirated by the micropipette were immobilized on the immunochromatographic resin at the edge of micropipette tip. The blocking solution was also aspirated in the same fashion. The measurement operation was performed by aspirating the sample solution, and then the gold colloidal nanoparticles (Au naps) conjugated secondary antibody solution. Since this bioanalysis system utilizes a micropipette, it is possible to increase the sample volume, which would enable the detection of antigen at low concentrations. In addition, the washing procedure can also be performed easily to reduce the background level. After the antigen-antibody reaction, the color intensity of Au naps could be observed by the naked eye. For analytical evaluation, the color intensity was captured by a scanner, and processed by analysis software. We have achieved the detection of human chorionic gonadotropin hormone (hCG) and total prostate-specific antigen (TPSA), which are well-known as fundamental indicators of pregnancy and cancer. The limit of detection (LOD) for hCG was 8 ng/ml (0.8 ng/tip), which is comparable to that of other conventional systems based on immunochromatography. Moreover, the LOD for TPSA was improved over the existing systems with the application of different sample volumes, such as 5 ng/μl (1 ng/tip) in 200 μl sample volume, and 2 ng/ml (0.6 ng/tip) in 300 ml sample volume. Since our bioanalysis system requires a small amount of immobilized antigen, it would be greatly useful in basic research for screening the antigen–antibody reactions. Besides, our bioanalysis system can be applied to on-field screening, since its operation is simple, and the visual results can be obtained rapidly.

A sensitive immunochromatographic assay using gold nanoparticles for the semiquantitative detection of prostate-specific antigen in serum

In prostate cancer screening, prostate-specific antigen (PSA) has been utilized as a valuable biomarker. There are routinely used procedures based on enzyme-linked immunosorbent assay (ELISA) for PSA detection. The procedures based on ELISA, however, are time consuming, complicated, and costly. We have developed a rapid, very simple, cost effective and sensitive immunochromatographic assay using gold nanoparticles and evaluated its applications for first screening of prostate cancer in serum samples. The sensitive immunochromatographic assay requires only 40 μL of the serum sample. The assay used is rapid and simple, that it totally takes approx 15 min to complete. The method for sensitive immunochromatographic assay has the other advantage of decreasing the antibody concentration that is used for the test line. In this study, we show the advantage to decrease the antibody concentration and the evaluation of our sensitive immunochromatographic assay for the semiquantitative detection of PSA in serum. The results obtained from 163 serum samples using sensitive immunochromatographic assay are compared with the results obtained using the chemiluminescent enzyme immunoassay (CLEIA) and normal immunochromatographic assay. The results obtained in the sensitive immunochromatographic assay correlated well with the values obtained in CLEIA. We concluded that our sensitive immunochromatographic assay is applicable to the first screening test for the diagnosis of prostate cancer. Our developed sensitive immunochromatographic assay is a promising candidate for diagnosis or research use, which may become commercially available in the near future.

Immunochromatographic assay using gold nanoparticles for measuring salivary secretory IgA in dogs as a stress marker

Abstract The concentration of salivary secretory immunoglobulin A (sIgA) is a well-known stress marker for humans. The concentration of salivary sIgA in dogs has also been reported as a useful stress marker. In addition, salivary sIgA in dogs has been used to determine the adaptive ability of dogs for further training. There are conventional procedures based on enzyme-linked immunosorbent assay (ELISA) for measuring salivary sIgA in dogs. However, ELISA requires long assay time, complicated operations and is costly. In the present study, we developed an immunochromatographic assay for measuring salivary sIgA in dogs using a dilution buffer containing a non-ionic surfactant. We determined 2500-fold dilution as the optimum condition for dog saliva using a phosphate buffer (50 mM, pH 7.2) containing non-ionic surfactant (3 wt% Tween 20). The results obtained from the saliva samples of three dogs using immunochromatographic assay were compared with those obtained from ELISA. It was found that the immunochromatographic assay is applicable to judge the change in salivary sIgA in each dog. The immunochromatographic assay for salivary sIgA in dogs is a promising tool, which should soon become commercially available for predicting a dogs psychological condition and estimating adaptive ability for training as guide or police dogs.