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1. 发明申请

WO2010071462A1 PEPTIDES MIMICKING UREASE, METHODS OF MANUFACTURING, APPLICATION IN DIAGNOSTIC TESTS AND THE WAY OF PERFORMANCE THE TEST 审中-公开
标题翻译：胶原蛋白的制备方法，制造方法，诊断测试中的应用和性能测试方法
公开(公告)号：WO2010071462A1
公开(公告)日：2010-06-24
申请号：PCT/PL2009/000106
申请日：2009-12-16
申请人： UNIWERSYTET HUMANISTYCZNO-PRZYRODNICZY , POLITECHNIKA ŁÓDZKA , KACA Wiesław , KAMIŃSKI Zbigniew , KOLESIŃSKA Beata , KWINKOWSKI Marek , ARABSKI Michał , KONIECZNA Iwona
发明人： KACA Wiesław , KAMIŃSKI Zbigniew , KOLESIŃSKA Beata , KWINKOWSKI Marek , ARABSKI Michał , KONIECZNA Iwona
IPC分类号： B01J20/289 , C08B11/14 , C12N1/06 , C12N9/80 , C12Q1/58
CPC分类号： C12N9/80 , C08B11/14 , G01N33/573
摘要： Synthetic peptides mimicking urease, said peptides comprising structures with general formula: when: R 1 means BK-61A: H 2 N-Ser-Ile-Lys-Glu-Asp-Val-Gln-Phe-; R 2 means BK-61B: H2N-Cys-His-His-Leu-Asp-Lys-Ser-Ile-Lys-Glu-Asp-Val-Gln- Phe-Ala-Asp-Ser-Arg-Ile-; R 3 means BK-65A: H 2 N-GIy- Arg-Arg-Asn-Leu-Lys-Trp-Met-Leu-Arg-; R 4 means BK-65B: H 2 N-Met-Leu-Met-Val-Cys-His-His-Leu-Asp-Pro-Ser-Ile-Pro- Glu-Asp-Val-Ala-; R 5 means BK-65C: H 2 N-Met-Val-Met-Ile-Thr-His-His-Leu-Asn-Ala-Ser-Ile-Pro-Glu- Asp-Ile-Ala-; R 6 means BK-65D: H 2 N-Met-Leu-Met-Val-Cys-His-His-Leu-Asn-Arg-Glu-Ile-Pro- Glu-Asp-Ile-Ala-. Methods for manufacturing of peptides mimicking urease said methods consisting im- mobilization of 2,4-dichloro-6-methoxy-l,3,5-triazine (DCMT) on cellulose plate fol- lowed by treatment with solution of N-methylmorpholine in THF and removing of excess of NMM by washing with THF, treatment with solution of protected amino acid from the group R 1 -R 6 , followed by treated with acetic anhydride solution, washed with DMF, washed with dichloromethane and ethanol (EtOH) and dried. The synthetic peptides mimicking urease recognized anti-urease antibodies as markers of autoimmune and atherosclerotic diseases. Method for performance the test for identification of antibodies againt ureases said meth- od comprising insertion of celullose pieces with immobilized synthetic peptides mimick- ing urease into microtiter plate wells, blocking of nonspecific binding sites with blocking buffer, than - after removal of blocking buffer - incubation with human serum, followed by washing with buffer, addition of anti-human antibodies conjugated with enzyme, se- quential washing and developing of colour.
摘要翻译：模拟脲酶的合成肽，所述肽包含具有以下通式的结构：当：R 1表示BK-61A：H 2 N-Ser-Ile-Lys-Glu-Asp-Val-Gln-Phe-; R2表示BK-61B：H2N-Cys-His-His-Leu-Asp-Lys-Ser-Ile-Lys-Glu-Asp-Val-Gln-Phe-Ala-Asp-Ser-Arg-Ile-; R3表示BK-65A：H2N-Gly-Arg-Arg-Asn-Leu-Lys-Trp-Met-Leu-Arg-; R4表示BK-65B：H2N-Met-Leu-Met-Val-Cys-His-His-Leu-Asp-Pro-Ser-Ile-Pro-Glu-Asp-Val-Ala- R5表示BK-65C：H 2 N-Met-Val-Met-Ile-Thr-His-His-Leu-Asn-Ala-Ser-Ile-Pro-Glu-Asp-Ile-Ala- R6表示BK-65D：H2N-Met-Leu-Met-Val-Cys-His-His-Leu-Asn-Arg-Glu-Ile-Pro-Glu-Asp-Ile-Ala-。模拟尿素酶的肽的制备方法说，所述方法包括在纤维素板上进行2,4-二氯-6-甲氧基-1,3,5-三嗪（DCMT）的活化，然后用N-甲基吗啉在THF中的溶液用THF洗涤去除过量的NMM，用R1-R6的保护氨基酸溶液处理，然后用乙酸酐溶液处理，用DMF洗涤，用二氯甲烷和乙醇（EtOH）洗涤，干燥。模仿脲酶的合成肽识别抗脲酶抗体作为自身免疫和动脉粥样硬化疾病的标志物。用于进行抗体鉴定的试验的方法再次测定脲酶，所述方法包括将固定的合成肽插入纤维素片段，将模拟脲酶插入微量滴定板孔中，用封闭缓冲液阻断非特异性结合位点，与人血清孵育，然后用缓冲液洗涤，加入与酶缀合的抗人抗体，顺序洗涤和显色。

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