Department of Biomedical Engineering, and ‡Department of Physics, Tufts University , 4 Colby Street, Medford Massachusetts 02155, United States.
Department of Chemical and Biomolecular Engineering, and ∥Polymer Program, University of Connecticut , Storrs, Connecticut 06269, United States.
ACS Appl Mater Interfaces. 2016 Jun 29;8(25):16218-26. doi: 10.1021/acsami.6b02215. Epub 2016 Jun 20.
Regenerated silk fibroin is a high molecular weight protein obtained by purifying the cocoons of the domesticated silkworm, Bombyx mori. This report exploits the aqueous processing and tunable β sheet secondary structure of regenerated silk to produce nanostructures (i.e., inverse opals) that can be used as colorimetric immunosensors. Such sensors would enable direct detection of antigens by changes in reflectance spectra induced by binding events within the nanostructure. Silk inverse opals were prepared by solution casting and annealing in a humidified atmosphere to render the silk insoluble. Next, antigen sensing capabilities were imparted to silk through a three step synthesis: coupling of avidin to silk surfaces, coupling of biotin to antibodies, and lastly antibody attachment to silk through avidin-biotin interactions. Varying the antibody enables detection of different antigens, as demonstrated using different protein antigens: antibodies, red fluorescent protein, and the beta subunit of cholera toxin. Antigen binding to sensors induces a red shift in the opal reflectance spectra, while sensors not exposed to antigen showed either no shift or a slight blue shift. This work constitutes a first step for the design of biopolymer-based optical systems that could directly detect antigens using commercially available reagents and environmentally friendly chemistries.
再生丝素蛋白是一种高分子量蛋白质，通过纯化家蚕（Bombyx mori）的茧获得。本报告利用再生丝的水相加工和可调β片二级结构来制备纳米结构（即反蛋白石），可作为比色免疫传感器。这种传感器可以通过纳米结构内结合事件引起的反射光谱变化来直接检测抗原。通过溶液浇铸和在潮湿气氛中退火来制备丝素反蛋白石，使丝素不溶。接下来，通过三步合成赋予丝素抗原传感能力：丝素表面与亲和素偶联、抗体与生物素偶联，最后通过亲和素-生物素相互作用将抗体附着在丝素上。通过改变抗体，可以检测到不同的抗原，如使用不同的蛋白质抗原：抗体、红色荧光蛋白和霍乱毒素的β亚基。抗原与传感器结合会引起蛋白石反射光谱的红移，而未暴露于抗原的传感器则没有位移或略有蓝移。这项工作是设计基于生物聚合物的光学系统的第一步，该系统可以使用市售试剂和环保化学物质直接检测抗原。
