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著者: Yukihiro Kobayashi, Shoichiro Ishizaki, Yuji Nagashima, Kazuo Shiomi
雑誌名: Parasitol Int. 2008 Sep;57(3):314-9. doi: 10.1016/j.parint.2008.01.005. Epub 2008 Feb 15.
Abstract/Text
Third stage larvae of the nematode Anisakis simplex often infect marine fish and invertebrates. When the larvae are ingested orally via seafood, they can cause IgE-mediated allergic reactions as well as anisakiasis. Of the known A. simplex allergens, Ani s 1 (Kunitz/bovine pancreatic trypsin inhibitor family protein) has been demonstrated to be a major allergen, being expected to be a useful tool for diagnosis of A. simplex allergy. For a diagnostic purpose, sufficient amounts of either natural Ani s 1 (nAni s 1) or recombinant Ani s 1 (rAni s 1) with an IgE-binding capacity should be stably supplied whenever needed. In this study, therefore, we first developed a simple and rapid purification method for Ani s 1 that is based on affinity chromatography using anti-Ani s 1 antibodies as ligands. The method was shown to produce nAni s 1 with a higher yield than the previously reported methods. Then, an attempt was made to express rAni s 1 in Escherichia coli as a His-tagged protein. rAni s 1 obtained as an inclusion body was solubilized in a solvent containing denaturing and reducing reagents and purified by nickel-chelate chromatography. Refolding of rAni s 1 was accomplished by dialysis in the presence of arginine, followed by that in the absence of arginine. Fluorescence ELISA and inhibition ELISA data revealed that rAni s 1 is IgE reactive enough to be used as a diagnostic tool.
PMID 18374626 Parasitol Int. 2008 Sep;57(3):314-9. doi: 10.1016/j.parint.2008.01.005. Epub 2008 Feb 15.
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