Fiber-optic chemical sensors for competitive binding fluoroimmunoassay.
Fiber-optic chemical sensors for competitive binding fluoroimmunoassay.
复制标题
用于竞争性结合荧光免疫测定的光纤化学传感器。
DOI：
10.1021/ac00135a033
复制
发表时间：
1987
影响因子：
7.4
通讯作者：
Griffin,GD
中科院分区：
文献类型：
--
作者：
Tromberg,BJ;Sepaniak,MJ;Vo-Dinh,T;Griffin,GD
本文介绍了一种基于竞争结合荧光免疫分析原理的光纤化学传感器的研制。兔免疫球蛋白G（IgG）共价固定在石英光纤的远端传感尖端上。传感器暴露于异硫氰酸荧光素（FITC）标记和未标记的抗兔IgG。氩离子激光器的488 nm线提供传感器结合分析物的激发。这导致在光纤的感测尖端处的荧光发射。传感器响应与样品中未标记抗IgG的量成反比。检测限（LOD）随孵育时间、样本量和测量条件而变化。对于10 μ L样品，典型的LOD为20 min孵育期内未标记抗体的25 fmol。这些结果表明，每一个光纤荧光免疫传感器可以被构造为执行一个单一的灵敏，快速，低容量的免疫测定，在原位或benchtop应用。光纤化学传感器（FOCS）已被设计为提供简单，快速，在原位分析的痕量化学品（1-8）。FOCS的特点是在纤维的取样末端有化学选择性的固定化试剂相.该试剂相将这些装置与其选择性较低的物理传感器对应物（9-11）区分开。固相免疫分析的原理可应用于FOCS的设计中，以实现对痕量化学物质的高灵敏度和高特异性的检测。这涉及将受体分子（抗体或抗原）共价固定到单链600 μ m直径石英光纤的远端面。纤维和固定的受体分子形成稳定的、选择性的荧光免疫传感器（FIS）。仔细选择适当的固定程序，通过最大限度地减少受体泄漏来增强FIS稳定性。只要有足够的固定化试剂用于合理的灵敏度，这种通过有机硅烷化试剂直接连接抗体或抗原的方法优于那些利用膜或凝胶包埋试剂相的技术。这是由于传感器响应时间受到纤维质量传输和免疫化学动力学的限制。尽管它们具有更高负载的潜力，但这些过程在膜或凝胶系统中可能较慢（1，2）。可以进行几种类型的免疫测定。最简单的方法包括原位FIS孵育，然后直接测量天然荧光分析物（12）。对于非荧光材料，原位孵育后在荧光团标记的第二抗体中“显影”。由此产生的“抗体夹心”会产生荧光信号
This paper describes the development of a fiber-optic chemical sensor based on the principle of competitive-binding fluorescence immunoassay. Rabbit immunoglobln G (IgG) is covalently Immobilized on the distal sensing tip of a quartz optical fiber. The sensor Is exposed to fluorescein Isothio-cyanate (FITC) labeled and unlabeled anti-rabbit IgG. The 488-nm line of an argon-ion laser provides excitation of sen-sor-bound analyte. This results in fluorescence emission at the optical fiber’s sensing tip. Sensor response Is inversely proportional to the amount of unlabeled antl-IgG In the sam-ple. Limits of detection (LOD) vary with Incubation time, sample size, and measurement conditions. For10-/xL sam-ples, typical LOD are 25 fmol of unlabeled antibody In a 20-min Incubation period. These results Indicate that each fiber-optic fluorolmmunosensor can be constructed to perform a single sensitive, rapid, low-volume immunoassay, In in situ or benchtop applications.Fiber-optic chemical sensors(FOCSs) have been designed to provide simple, rapid, in situ analyses of trace chemicals (1-8). FOCSs are characterized by their chemically selective immobilized reagent phase atthe fiber’s sampling terminus. This reagent phase distinguishes these devices from their less selective physical sensor counterparts (9-11). In orderto detect trace amounts of chemicals with high sensitivity and specificity, the principles of solid-phase im-munoassay can be applied to FOCSdesign. This involves the covalent immobilization of receptor molecules (antibody or antigen) to the distal face of a single-strand 600-/wn-diameter quartz optical fiber. The fiber and immobilized receptor molecules form a stable, selective fluoroimmunosensor (FIS). Careful selection of the proper immobilization procedure enhances FIS stability by minimizing receptor leakage. Provided there is sufficient immobilized reagent for reasonable sensitivity, this direct attachment of antibody or antigen via organosilanating reagents is preferable to those techniques which utilize membrane or gel-entrapped reagent phases. This is due to the fact that sensor response times are limited by mass transport to the fiber and immunochemical kinetics. Despite their potential for higher loadings, theseprocesses may be slower in membrane or gel systems (1, 2). Several types of immunoassays can be performed. The simplest involves in situ FIS incubation followed by direct measurement of a naturally fluorescentanalyte (12). For nonfluorescent materials, in situ incubation is followed by “development” in fluorophor-labeled second antibody. The resulting “antibody sandwich" produces a fluorescence signal