Kubitschko S, Spinke J, Brückner T, Pohl S, Oranth N
Diagnostics Division, F. Hoffmann-La Roche Ltd., Basel, CH-4070, Switzerland.
Anal Biochem. 1997 Nov 1;253(1):112-22. doi: 10.1006/abio.1997.2337.
In recent years, several optical sensor techniques have been developed for the direct monitoring of biomolecular recognition processes at the surface of a sensor chip. Applications of these immunosensors for the determination of substances in serum could be demonstrated only for a few analytes due to the lack of sensitivity. Beside nonspecific binding of serum components to the sensor surface, the analytical sensitivity of these sensors is limited by the molecular weight of the analyte, so that smaller analyte molecules give only a moderate sensor response. In order to enhance the sensor signal, the use of mass labels, such as latex particles, was proposed in the literature. However, detection limits comparable to those of conventional ELISA techniques could not be realized so far. We demonstrate the optimization of a "nanoparticle enhanced immunosensor assay" for the detection of thyroid stimulating hormone, with respect to the particle coating, size, and nonspecific binding. The developed prototype assay requires a sample volume of 225 microL and has a measuring range up to 35 mIU/L. For the first time, we obtained a detection limit of 0.03 mIU/L (0.1 pm), which is fully competitive to conventional ELISA techniques. The assay allows serum samples to be measured with good precision and dilution linearity. The sensor can be reused several times and shows an excellent correlation to a commercial enzyme immunoassay.
近年来，已开发出几种光学传感器技术，用于直接监测传感器芯片表面的生物分子识别过程。由于缺乏灵敏度，这些免疫传感器仅能用于少数分析物的血清中物质测定。除血清成分与传感器表面的非特异性结合外，这些传感器的分析灵敏度还受分析物分子量的限制，因此较小的分析物分子只能产生适度的传感器响应。为增强传感器信号，文献中提出使用质量标签，如乳胶颗粒。然而，目前尚未实现与传统酶联免疫吸附测定（ELISA）技术相当的检测限。我们展示了针对甲状腺刺激激素检测的“纳米颗粒增强免疫传感器测定法”在颗粒包被、尺寸和非特异性结合方面的优化。所开发的原型测定法需要225微升的样品体积，测量范围高达35 mIU/L。我们首次获得了0.03 mIU/L（0.1皮摩尔）的检测限，与传统ELISA技术具有充分的竞争力。该测定法能够以良好的精密度和稀释线性对血清样品进行测量。该传感器可重复使用多次，并且与商业酶免疫测定法具有出色的相关性。
