Reactivity of urinary albumin (microalbumin) assays with fragmented or modified albumin
Reactivity of urinary albumin (microalbumin) assays with fragmented or modified albumin
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DOI：
10.1373/clinchem.2007.092825
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发表时间：
2008-01-01
影响因子：
9.3
通讯作者：
Hortin, Glen L.
中科院分区：
文献类型：
--
作者：
Sviridov, Denis;Drake, Steven K.;Hortin, Glen L.
背景：关于尿中白蛋白结构变异体的发生和测量存在争议。在这项研究中，我们检查了在体外修饰的白蛋白在测定尿白蛋白（微量白蛋白）的交叉反应性。方法：我们分析了试剂，胰蛋白酶，或物理处理或不同的一级序列（动物白蛋白）与免疫比浊法（贝克曼LX 20）使用山羊抗血清和竞争性免疫测定（西门子Immulite）使用单克隆抗体的白蛋白。我们评估发生在尿中的白蛋白片段通过使用Western blotting的24个samples.Results：化学修饰，适度的序列取代（大猩猩白蛋白），或裂解的白蛋白溴化氰（CNBr）在LX 20检测反应性的影响不大。用胰蛋白酶广泛裂解的白蛋白保留部分反应性。Immulite测定通常受白蛋白修饰和序列变化的影响更严重。新鲜尿液标本或储存在-80 ℃的标本的蛋白质印迹显示白蛋白几乎没有碎片，但一些储存在-20 ℃ 3年的标本有广泛的碎片白蛋白，可通过LX 20检测到，但不能通过Immulite检测。在免疫比浊法中，完整白蛋白和CNBr片段的反应性几乎相等，表明抗体在整个过程中具有多个表位的反应性。白蛋白。因此，在这种类型的尿白蛋白测定中难以消除白蛋白的反应性。2种检测试剂盒对白蛋白修饰的灵敏度差异确定了测定尿白蛋白时检测试剂盒不等效性的潜在来源，特别是对于储存在-20 ℃下的标本。(c)2007年美国临床化学协会。
BACKGROUND: Controversy exists regarding occurrence and measurement of structural variants of albumin in urine. In this study, we examined cross -reactivity of in vitro modified albumins in assays for urine albumin (microalbumin).METHODS: We analyzed albumin modified by reagents, trypsin, or physical treatments or differing in primary sequence (animal albumins) with an immunoturbidimetric assay (Beckman LX20) using goat antiserum and a competitive immunoassay (Siemens Immulite) using a monoclonal antibody. We assessed occurrence of albumin fragments in urine by use of Western blotting of 24 specimens.RESULTS: Chemical modification, modest sequence substitution (gorilla albumin), or cleavage of albumin by cyanogen bromide (CNBr) had little effect on reactivity in the LX20 assay. Albumin extensively cleaved with trypsin retained partial reactivity. The Immulite assay generally was affected more severely by albumin modifications and sequence changes. Western blots of fresh urine specimens or specimens stored at -80 degrees C showed little albumin fragmentation, but some specimens stored for 3 years at -20 degrees C had extensively fragmented albumin that was detected by the LX20 but not the Immulite assay.CONCLUSIONS: Nearly equivalent reactivity of intact albumin and CNBr fragments in the immunoturbidimetric assay indicates reactivity of antibodies with multiple epitopes throughout albumin. Therefore, it is difficult to abolish reactivity of albumin in this type of urine albumin assay. Differential sensitivity of 2 assays to albumin modification identifies a potential source of assay nonequivalence in measuring urinary albumin, particularly for specimens stored at -20 degrees C. (c) 2007 American Association for Clinical Chemistry.