CN1173224A - 多层分析元件 - Google Patents
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Abstract
本发明涉及含可与待测样相互作用的试剂、可定量检测待测样某一组分的多层分析元件,该元件包括一种固体载体和一层附着其上的亲水层,后者由明胶、聚乙烯醇或上列的混合物组成且含用量占总重5%~30%的一种显影剂及生色剂,显影剂和生色剂重量比为1∶0.3~1∶3.0,显影剂在所述试剂中的氧化剂作用下发生氧化反应,由此被氧化后的显影剂与生色剂反应生成有色产物。本发明多层分析元件可用于医学和药学,以及刑事领域。它可用于胆甾醇、氯离子和葡萄糖检测,故便于囊纤维病、糖尿病以及高血脂症的诊断。
Description
发明背景
本发明涉及材料分析领域的改进。具体而言,本发明涉及用于皮肤及其它固体材料中胆甾醇含量检测的多层分析元件。
苏联(USSR)作者证书第1054784号介绍了一种类脂物薄层色谱法分析用分析元件,该元件含一种载体(如玻璃板),板上附一层吸收材料,如硅胶,石膏,硫酸铵及乙酸铜的混合物。该类分析元件不适用人皮肤中胆甾醇检测。
苏联(USSR)作者证书第1495378号介绍了一种双层分析元件,该元件含一种载体(如聚乙烯膜)及含固定化氧化酶的明胶层。元件在有胺或水溶液与其中的明胶层相接触时变色。该类分析元件也不适用人皮肤中胆甾醇检测。
加拿大专利第1335968号介绍一种检测胆甾醇的方法为,使皮肤或其它含胆甾醇表面与通式A-C-B的酶亲和性化合物相接触,其中A为对胆甾醇具亲和性的检测剂,B为可与生色剂反应生成有色产物的酶性显色剂,且C为将显色剂和检测剂连结在一起的胶粘剂。胆甾醇通过生色剂变色检出。该法之不足在于,溶液及显色后的反应产物不够稳定,胆甾醇含量低时敏感度及可重复性极差。
鉴于上述已有分析元件及检测方法存在不足,有必要结合现有胆甾醇液相检测方法的优点和带固体载体多层分析元件在实用中的便利,开发新的分析元件。
美国专利第4895704号已提出一种多层分析元件,它含隔水、透光固体载体及一个亲水层,后者至少含一种试剂可与待检测样某一组分反应生成可经光辐射检出的物质。该分析元件包括含光散射颗粒的附加层,含量足以保证光透射率介于10%~2.5%。这种已知的多层分析元件不宜用于皮肤中胆甾醇检测,显色反应强度低,且待检测物含量低时分析衬度高。
发明概要
本发明旨在提高分析元件的敏感度、可重复性及显色反应强度,增加显色时酶反应的持续时间,强化皮肤中胆甾醇含量分析手段,并且拓展其在胆甾醇即时诊断中的应用范围。
广义而言,本发明提供含可与待测样相互作用试剂、可定量检测待测样某一组分的多层分析元件。本发明多层分析元件包括一种载体和一层附着其上的亲水层,后者由明胶,聚乙烯醇或上列的混合物组成且含用量占总重5%~30%的一种显影剂及生色剂,显影剂和生色剂重量比为1∶0.3~1∶3.0。显影剂在所述试剂中的氧化剂作用下发生氧化反应,故此被氧化后的显影剂与生色剂反应生成有色产物。
本发明多层分析元件用于胆甾醇检测时,所用试剂为如前通式A-C-B定义的酶亲和性化合物,其中酶显色剂B为过氧化酶。此时,显影剂在过氧化氢中的过氧化酶作用下发生氧化反应,故此显影剂被氧化后与生色剂反应生成标示皮肤中胆甾醇含量的有色产物。优选实施方案的说明
所用显影剂优选下式的化合物其中:R,R′及R″可相同也可不同,且各自分别为氢原子,烷基或链烷醇基。这类化合物的例子包括N,N-二乙基对苯二胺,对乙基氨基-邻甲苯胺,对-[N-乙基-N-(β-羟乙烯基)氨基]-苯胺,对-[N-乙基-N-(β-甲磺酰氨基乙基)氨基]-邻甲苯胺,对-[N-乙基-N-(β-羟乙基)氨基]-邻甲苯胺,以及对-[N-丁基-N-(γ-磺酰丙基)氨基]-苯胺。
二乙基对甲苯二胺,乙氧基乙基对苯二胺,乙基甲磺酰氨基乙基对甲苯二胺,乙氧基乙基对甲苯二胺,丁磺酰丙基对苯二胺,或二乙基对苯二胺也可用作显影剂。
生色剂可采用吡唑酮-5,苯甲酰乙酸或萘酸的衍生物。举例而言,适选生色剂包括1-(2,4,6-三氯苯基)-3-十五烷基-氨基苯基-氨基吡唑酮,3,5-二羧苯基十八烷基氨基-1-氧-2-萘酸,2-氯-5-[γ-(2,4-二-四胺苯氧基)-丁酰氨基)-苯胺-α-N-苯甲酰海因酰新戊酰乙酸,间-(N-乙基-N-十八烷基)-氨基-对苯甲酰乙酰胺苯磺酸,间羧基-间-{N-(对氯-间-(N-邻十八烷基-羟基苯甲酰基乙酰基)氨基)苯酰基氨基)-羧酸,邻苯氧基-间-(5-氧-1,2-吡唑啉-3-基)苯磺酸,2-甲基十八烷基氨基-5′-磺酰苯胺苯甲酰乙酸,2′-氯-5′-(3″,5″-二羧苯基磺氨基)-苯胺基-2-十八烷氧基苯甲酰乙酸,十八烷基氨基-1-氧-4-硫-2-萘酸,2′-甲基十八烷基氨基-5′-磺酰苯胺基-1,2-羟萘酸,以及1-(4′-苯氧基-3′-磺苯基)-3-甾基-氨基吡唑酮。
生色剂可以微囊、微粒或分散的分子均匀分布在亲水层中,这取决于生色剂的性质。如欲使生色剂在亲水层以微粒或分子态分布,则采用具亲水性(极性)支链的化合物。如欲使生色剂以微囊态分布,则采用具憎水性(非极性)支链的化合物。
载体优选由聚对苯二甲酸乙二醇酯,聚乙烯或三硝基纤维素亲水膜制得。无色聚合物,纸或聚乙烯包覆纸也可采用。
亲水层也可含增湿和鞣制剂。增湿剂可保证载体上亲水层以不变的均一厚度,优选2-100微米涂敷。鞣制剂可保证多层分析元件所需的物理-力学性能。乙酸铬及2,4-二氯-6-羟三唑-1,3,5的一钠盐可用作鞣制剂。
本发明多层分析元件可用于医学和药学,以及刑事领域。它不仅可用于胆甾醇检测,也可用于检测氯离子和葡萄糖,故便于囊纤维病、糖尿病以及高血脂症的诊断。
下列非限制实施例进一步说明。实施例1
制得一个用于皮肤中胆甾醇检测的多层分析元件,方法是,在厚50微米的对苯二甲酸膜载体上涂敷一层光成象明胶,胶中含75%(重量)亲水性紫色生成剂,后者由1-(2,4,6-三氯苯基)-3-十五烷基-氨基苯基-氨基吡唑酮组成。该生色剂平均粒径小于0.1微米。明胶层含7.5%(重量)的N,N-二乙基对苯二胺,后者系以分子水平分散的显影剂。该层中也含乙酸铬鞣制剂
该多层分析元件分别用于患者手外皮及健康人皮肤外皮中胆甾醇的检测。取如前式A-C-B(其中酶显色剂B为过氧化酶)的酶亲和化合物水溶液各一滴,涂于每一人的掌皮。经1分钟培养后,干燥或以棉球除去液体。在同一处,滴涂一滴过氧化氢,随即即刻使多层分析元件紧贴并压实于皮表面1分钟。其后,在流水中冲洗分析元件15~20秒。将所得色迹与带刻度的色标对比,或以分光光度方法如LCD(液晶显示)分光光度计测定色点的强度,则得检测结果。
分别以现有液相法和本发明多层分析元件测定血液及皮肤中的胆甾醇含量,结果如表1所示。
表1
测试组 已有方法测定 本发明测定
血胆甾醇含量 皮胆甾醇含量 皮胆甾醇含量健康人(24人) 185±25 1.9±0.3 0.13±0.04前巩膜患者(29人) 228±32 2.8±0.5 0.32±0.07
对同一组患者依次进行10次检测,考察显色反应的可重复性,标准偏差不大于8%。
多层分析元件分别隔一周和六个月,得同样重复结果。
检测时间需2~3分钟,准备时间约3分钟。使用该多层分析元件,可制成配套仪器,供患者自检。实施例2
制得一个用于皮肤中胆甾醇检测的多层分析元件,方法是,在白色聚乙烯包覆光成象纸载体上涂敷一层含蓝色生色剂的聚乙烯醇,生色剂由3,5-二羧苯基十八烷基氨基-1-氧-萘酸组成。聚乙烯醇尚含对乙基氨基-邻甲苯胺显影剂。
该多层分析元件用于外皮中胆甾醇检测,步骤同实施例1所述,得基本相同结果。实施例3
制得一个柔性多层分析元件,方法是,在厚100微米、经电晕放电等离子体亲水处理的聚乙烯膜载体上涂敷一层明胶与聚乙烯醇的混合物,层中尚含亲水性黄色生色剂分散物,也即2-氯-5-[γ-(2,4-二-四胺苯氧基)-丁酰氨基)-苯胺-α-N-苯甲酰基海因酰基新戊酰乙酸。明胶及聚乙烯醇层尚含对-[N-乙基-N-(β-羟乙烯基)氨基]-苯胺显影剂。
该多层分析元件用于皮中胆甾醇检测,步骤同实施例1所述,得基本相同结果。
Claims (22)
1含可与待测样相互作用的试剂、可定量检测待测样某一组分的多层分析元件,该元件包括一种固体载体和一层附着其上的亲水层,后者由明胶,聚乙烯醇或上列的混合物组成且含用量占总重5%~30%的一种显影剂及生色剂,显影剂和生色剂重量比为1∶0.3~1∶3.0,显影剂在所述试剂中的氧化剂作用下发生氧化反应,由此被氧化后的显影剂与生色剂反应生成有色产物。
3权利要求2的多层分析元件,其中,所述显影剂为选自下列一组的一种化合物:N,N-二乙基对苯二胺,对乙基氨基-邻甲苯胺,对-[N-乙基-N-(β-羟乙烯基)氨基]-苯胺,对-[N-乙基-N-(β-甲磺酰氨基乙基)氨基]-邻甲苯胺,对-[N-乙基-N-(β-羟乙基)氨基]-邻甲苯胺,以及对-[N-丁基-N-(γ-磺酰丙基)氨基]-苯胺。
4权利要求1的多层分析元件,其中,所述显影剂为选自下列一组的一种化合物:二乙基对甲苯二胺,乙氧基乙基对苯二胺,乙基甲磺酰氨基乙基对甲苯二胺,乙氧基乙基对甲苯二胺,丁磺酰丙基对苯二胺,以及二乙基对苯二胺。
5权利要求1的多层分析元件,其中,所述生色剂为选自一组由吡唑酮-5、苯甲酰乙酸或萘酸的衍生物组成的化合物中的一种。
6权利要求5的多层分析元件,其中,所述生色剂为选自下列一组的一种化合物:1-(2,4,6-三氯苯基)-3-十五烷基-氨基苯基-氨基吡唑酮,3,5-二羧苯基十八烷基氨基-1-氧-2-萘酸,2-氯-5-[γ-(2,4-二-四胺苯氧基)-丁酰氨基)-苯胺-α-N-苯甲酰海因酰新戊酰乙酸,间-(N-乙基-N-十八烷基)-氨基-对苯甲酰乙酰胺苯磺酸,间羧基-间-{N-(对氯-间-(N-邻十八烷基-羟基苯甲酰基乙酰基)氨基)苯酰基氨基)-羧酸,邻苯氧基-间-(5-氧-1,2-吡唑啉-3-基)苯磺酸,2-甲基十八烷基氨基-5′-磺酰苯胺苯甲酰乙酸,2′-氯-5′-(3″,5″-二羧苯基磺氨基)-苯胺基-2-十八烷氧基苯甲酰乙酸,十八烷基氨基-1-氧-4-硫-2-萘酸,2′-甲基十八烷基氨基-5′-磺酰苯胺基-1,2-羟萘酸,以及1-(4′-苯氧基-3′-磺苯基)-3-甾基-氨基吡唑酮。
7权利要求5的多层分析元件,其中,所述生色剂为粒径小于0.1微米的憎水性化合物。
8权利要求7的多层分析元件,其中,所述憎水性化合物为1-(2,4,6-三氯苯基)-3-十五烷基氨基苯基-氨基吡唑酮,或2-氯-5-[γ-(2,4-二-四胺苯氧基)-丁酰氨基)-苯胺-α-N-苯甲酰海因酰新戊酰乙酸。
9权利要求1的多层分析元件,其中,所述亲水层厚介于2~100微米。
10权利要求1的多层分析元件,其中,所述亲水层尚含至少一种增湿剂。
11权利要求1的多层分析元件,其中,所述亲水层尚含至少一种鞣制剂。
12在通式A-C-B的酶亲和性化合物存在下检测胆甾醇的多层分析元件,其中:A为对胆甾醇具亲和性的检测剂,B为含过氧化酶的酶性显色剂,且C为将显色剂和检测剂连结在一起的胶粘剂;所述元件包括一种固体载体和一层附着其上的亲水层,后者由明胶,聚乙烯醇或上列的混合物组成且含用量占总重5%~30%的一种显影剂及生色剂,显影剂和生色剂重量比为1∶0.3~1∶3.0,显影剂在过氧化氢中的过氧化酶作用下发生氧化反应,由此被氧化后的显影剂与生色剂反应生成有色产物。
14权利要求13的多层分析元件,其中,所述显影剂为选自下列一组的一种化合物:N,N-二乙基对苯二胺,对乙基氨基-邻甲苯胺,对-[N-乙基-N-(β-羟乙烯基)氨基]-苯胺,对-[N-乙基-N-(β-甲磺酰氨基乙基)氨基]-邻甲苯胺,对-[N-乙基-N-(β-羟乙基)氨基]-邻甲苯胺,以及对-[N-丁基-N-(γ-磺酰丙基)氨基]-苯胺。
15权利要求12的多层分析元件,其中,所述显影剂为选自下列一组的一种化合物:二乙基对甲苯二胺,乙氧基乙基对苯二胺,乙基甲磺酰氨基乙基对甲苯二胺,乙氧基乙基对甲苯二胺,丁磺酰丙基对苯二胺,以及二乙基对苯二胺。
16权利要求12的多层分析元件,其中,所述生色剂为选自一组由吡唑酮-5、苯甲酰乙酸或萘酸的衍生物组成的化合物中的一种。
17权利要求16的多层分析元件,其中,所述生色剂为选自下列一组的一种化合物:1-(2,4,6-三氯苯基)-3-十五烷基-氨基苯基-氨基吡唑酮,3,5-二羧苯基十八烷基氨基-1-氧-2-萘酸,2-氯-5-[γ-(2,4-二-四胺苯氧基)-丁酰氨基)-苯胺-α-N-苯甲酰海因酰新戊酰乙酸,间-(N-乙基-N-十八烷基)-氨基-对苯甲酰乙酰胺苯磺酸,间羧基-间-{N-(对氯-间-(N-邻十八烷基-羟基苯甲酰基乙酰基)氨基)苯酰基氨基)-羧酸,邻苯氧基-间-(5-氧-1,2-吡唑啉-3-基)苯磺酸,2-甲基十八烷基氨基-5′-磺酰苯胺苯甲酰乙酸,2′-氯-5′-(3″,5″-二羧苯基磺氨基)-苯胺基-2-十八烷氧基苯甲酰乙酸,十八烷基氨基-1-氧-4-硫-2-萘酸,2′-甲基十八烷基氨基-5′-磺酰苯胺基-1,2-羟萘酸,以及1-(4′-苯氧基-3′-磺苯基)-3-甾基-氨基吡唑酮。
18权利要求16的多层分析元件,其中,所述生色剂为粒径小于0.1微米的憎水性化合物。
19权利要求18的多层分析元件,其中,所述憎水性化合物为1-(2,4,6-三氯苯基)-3-十五烷基-氨基苯基-氨基吡唑酮,或2-氯-5-[γ-(2,4-二四胺苯氧基)-丁酰氨基)-苯胺-α-N-苯甲酰海因酰新戊酰乙酸。
20权利要求12的多层分析元件,其中,所述亲水层厚介于2~100微米。
21权利要求12的多层分析元件,其中,所述亲水层尚含至少一种增湿剂。
22权利要求12的多层分析元件,其中,所述亲水层尚含至于少一种鞣制剂。
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RU94044169/14A RU94044169A (ru) | 1994-12-16 | 1994-12-16 | Многослойный аналитический элемент |
RU94044169 | 1994-12-16 |
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CN1173224A true CN1173224A (zh) | 1998-02-11 |
CN1154844C CN1154844C (zh) | 2004-06-23 |
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US (1) | US6605440B2 (zh) |
EP (1) | EP0797774B1 (zh) |
JP (1) | JP3755007B2 (zh) |
KR (1) | KR100235211B1 (zh) |
CN (1) | CN1154844C (zh) |
AT (1) | ATE282204T1 (zh) |
AU (1) | AU702663B2 (zh) |
BR (1) | BR9510038A (zh) |
CA (1) | CA2207555C (zh) |
DE (1) | DE69533756T2 (zh) |
ES (1) | ES2227564T3 (zh) |
MX (1) | MX9704469A (zh) |
PT (1) | PT797774E (zh) |
RU (1) | RU94044169A (zh) |
WO (1) | WO1996018898A1 (zh) |
Cited By (2)
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CN101052874B (zh) * | 2004-09-17 | 2010-09-08 | 富士胶片株式会社 | 用于分析液体样品的多层分析元件 |
CN101530327B (zh) * | 2009-04-20 | 2012-09-05 | 湖州美奇医疗器械有限公司 | 皮下组织实时监测用针状电流测定式葡萄糖传感器及其制作方法 |
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WO2001011359A2 (en) | 1999-08-06 | 2001-02-15 | Imi International Medical Innovations Inc. | Color space analysis in biochemical and immunological assays |
US7250304B2 (en) | 2000-03-31 | 2007-07-31 | The Regents Of The University Of California | Functional assay of high-density lipoprotein |
US6596544B1 (en) | 2000-03-31 | 2003-07-22 | The Regents Of The University Of California | Functional assay of high-density lipoprotein |
DE60113976T2 (de) * | 2000-03-31 | 2006-07-27 | The Regents Of The University Of California, Oakland | Funktionaler nachweis von lipoprotein mit hoher dichte |
US7347879B2 (en) | 2004-02-27 | 2008-03-25 | L'Oreál, S.A. | Sulfur-containing secondary para-phenylenediamines dye compositions comprising such para-phenylenediamines, processes, and uses thereof |
FR2866882B1 (fr) * | 2004-02-27 | 2017-04-14 | Oreal | Para-phenylenediamines secondaires soufrees, composition tinctoriale comprenant de telles para-phenylenediamines, procede mettant en oeuvre cette composition et utilisation |
US20050272112A1 (en) * | 2004-04-28 | 2005-12-08 | Imi International Medical Innovations, Inc. | Direct assay of cholesterol in skin removed by tape stripping |
US7238494B2 (en) * | 2004-04-30 | 2007-07-03 | Imi International Medical Innovations, Inc. | Direct assay of cholesterol in skin samples removed by tape stripping |
US20070287991A1 (en) * | 2006-06-08 | 2007-12-13 | Mckay William F | Devices and methods for detection of markers of axial pain with or without radiculopathy |
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FR2499444A1 (fr) | 1981-02-11 | 1982-08-13 | Termet Pierre | Nouvel appareil pour un enfoncement de chevilles ou d'objets analogues |
JPS57150399A (en) * | 1981-03-13 | 1982-09-17 | Fuji Photo Film Co Ltd | Reagent for determining hydrogen peroxide and its determination method |
JPS5946854A (ja) * | 1982-09-10 | 1984-03-16 | Fuji Photo Film Co Ltd | 多層分析材料 |
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JPS6211167A (ja) | 1985-07-09 | 1987-01-20 | Fuji Photo Film Co Ltd | コレステロ−ル分析用多層分析要素 |
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JPS63158000A (ja) | 1986-12-18 | 1988-06-30 | Fuji Photo Film Co Ltd | 総コレステロ−ル定量用一体型多層分析要素 |
JPH07102156B2 (ja) | 1987-04-02 | 1995-11-08 | 東洋紡績株式会社 | 脱水素酵素または基質の測定法 |
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- 1994-12-16 RU RU94044169/14A patent/RU94044169A/ru unknown
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1995
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- 1995-12-14 CN CNB951973673A patent/CN1154844C/zh not_active Expired - Fee Related
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- 1995-12-14 AU AU41694/96A patent/AU702663B2/en not_active Ceased
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Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN101052874B (zh) * | 2004-09-17 | 2010-09-08 | 富士胶片株式会社 | 用于分析液体样品的多层分析元件 |
CN101530327B (zh) * | 2009-04-20 | 2012-09-05 | 湖州美奇医疗器械有限公司 | 皮下组织实时监测用针状电流测定式葡萄糖传感器及其制作方法 |
Also Published As
Publication number | Publication date |
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JPH10510159A (ja) | 1998-10-06 |
US20010007773A1 (en) | 2001-07-12 |
ATE282204T1 (de) | 2004-11-15 |
DE69533756D1 (de) | 2004-12-16 |
ES2227564T3 (es) | 2005-04-01 |
EP0797774A1 (en) | 1997-10-01 |
MX9704469A (es) | 1998-05-31 |
KR100235211B1 (ko) | 1999-12-15 |
AU4169496A (en) | 1996-07-03 |
US6605440B2 (en) | 2003-08-12 |
CA2207555A1 (en) | 1996-06-20 |
CA2207555C (en) | 2004-02-24 |
WO1996018898A1 (en) | 1996-06-20 |
JP3755007B2 (ja) | 2006-03-15 |
AU702663B2 (en) | 1999-02-25 |
DE69533756T2 (de) | 2005-11-10 |
CN1154844C (zh) | 2004-06-23 |
EP0797774B1 (en) | 2004-11-10 |
BR9510038A (pt) | 1998-11-03 |
PT797774E (pt) | 2005-03-31 |
RU94044169A (ru) | 1996-10-20 |
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