Flow injection analysis using specific biochemical reaction on latex particles and liposome

为了提高反应效率，提出了一种新的流动注射法（FIA），将酶固定在合成聚合物乳液表面。合成的苯乙烯-丙烯酸共聚物乳液为球形，粒径均匀，约为0.5mm。固定化酶的酶活性与丙烯酸的摩尔分数有关。将乳酸脱氢酶固定在乳胶粒上，两个月或更长时间内酶活仍保持在原酶的50%以上。在FIA中使用悬浮在载体溶液中的胶乳获得的结果与使用酶固定化反应器的常规FIA中的结果相当。另一种新的流动注射脂质体免疫分析法（Flowinjection Liposome Immunoassay，FILIA）是将抗原固定化的脂质体与标记分子结合，在连续流动系统中进行竞争性免疫分析。通过在使用POD缀合抗原的流动注射酶免疫测定中观察到令人满意的结果，证实了固定在反应器壁上的抗体的足够活性。然而，由于体积大的脂质体和反应器的宽壁的空间位阻，在FILIA中不能找到免疫反应的任何明确证据。通过在抗原和脂质体之间或抗体和反应器壁之间插入亲水聚合物间隔物，对于作为样品注射的游离抗原获得S形响应。但对于低浓度的游离分析物的测定，响应不足。

A New Flow injection method (FIA) was proposed in order to improve the reaction efficiency, in which enzyme was immobilized on the surface of synthetic polymer latex. Stirene-acrylic acid copolymer latex synthesized was spherical and had homogeneous size of about 0.5 mm diameter. The enzymatic activity of enzyme-immobilized latex was dependent on the molar fraction of acrylic acid. Lactate dehydrogenase immobilized on the latex particles kept the activity higher than 50% of original one for two months or longer. The results obtained in FIA using latex suspended in the carrier solution were comparable to those in the conventional FIA using enzyme-immobilized reactors. Difficulties in recovering and reusing the used latex were the serious problem.Another new FIA method (flow injection liposome immunoassay : FILIA) was discussed, in which antigen-immobilized liposome having marker molecules in its inner space was used and a competitive immunoassay was carried out in a continuous flow system. The sufficient activity of antibody immobilized on the wall of a reactor was confirmed by observing satisfactory results in a flow injection enzyme immunoassay using POD-conjugated antigen. However, any explicit proof for the immunoreaction could not be found in FILIA because of steric hindrance by bulky liposome and wide wall of the reactor. By inserting a hydrophillic polymer spacer between antigen and liposome or between antibody and the reactor wall, a sigmoid response was obtained for the free antigen injected as a sample. But the response was insufficient for the determination of low concentration of the free analyte.