Production of scFv-conjugated affinity silk film and its application to a novel enzyme-linked immunosorbent assay.
Production of scFv-conjugated affinity silk film and its application to a novel enzyme-linked immunosorbent assay.
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DOI：
10.1038/srep04080
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发表时间：
2014-02-12
影响因子：
4.6
通讯作者：
Kitani H
中科院分区：
文献类型：
--
作者：
Sato M;Kojima K;Sakuma C;Murakami M;Tamada Y;Kitani H
家蚕丝蛋白作为一种独特的生物材料已被广泛应用于医学领域。为了开发一种新型的亲和丝材料，我们构建了一种转基因家蚕，其丝素蛋白L链与单链可变区(ScFv)相连作为融合蛋白。在此之前，单链抗体结合的“亲和力”丝粉特异性地免疫沉淀其目标蛋白--Wiskott-Aldrich综合征蛋白。为了扩大亲和丝材料的适用性，我们将ScFv偶联的丝蛋白处理成薄膜，将其溶解在溴化锂中，然后在96孔板的孔中干燥。酶联免疫吸附试验证明，Wiskott-Aldrich综合征蛋白作为一种重组蛋白和从小鼠巨噬细胞中提取的天然形式都可以特异性地检测到。这些发现表明，这种单链抗体结合的丝膜可以作为替代免疫检测系统的基础。
Bombyx mori (silkworm) silk proteins have been utilized as unique biomaterials for various medical applications. To develop a novel affinity silk material, we generated a transgenic silkworm that spins silk protein containing the fibroin L-chain linked with the single-chain variable fragment (scFv) as a fusion protein. Previously, the scFv-conjugated “affinity” silk powder specifically immunoprecipitated its target protein, Wiskott-Aldrich syndrome protein. To expand the applicability of affinity silk materials, we processed the scFv-conjugated silk protein into a thin film by dissolving it in lithium bromide, then drying it in the wells of 96-well plates. Enzyme-linked immunosorbent assay demonstrated specific detection of Wiskott-Aldrich syndrome protein, both as a recombinant protein and in its native form extracted from mouse macrophages. These findings suggest that this scFv-conjugated silk film serves as the basis for an alternative immunodetection system.
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