CN112708657A - 一种快速消除羟苯磺酸钙、酚磺乙胺药物干扰的肌酐试剂盒及其制备方法 - Google Patents
一种快速消除羟苯磺酸钙、酚磺乙胺药物干扰的肌酐试剂盒及其制备方法 Download PDFInfo
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- CN112708657A CN112708657A CN202011578371.XA CN202011578371A CN112708657A CN 112708657 A CN112708657 A CN 112708657A CN 202011578371 A CN202011578371 A CN 202011578371A CN 112708657 A CN112708657 A CN 112708657A
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- reagent
- creatinine
- etamsylate
- trinder
- calcium dobesilate
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- HBGOLJKPSFNJSD-UHFFFAOYSA-N Etamsylate Chemical compound CC[NH2+]CC.OC1=CC=C(O)C(S([O-])(=O)=O)=C1 HBGOLJKPSFNJSD-UHFFFAOYSA-N 0.000 title claims abstract description 54
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- C12Q1/26—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving oxidoreductase
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Abstract
本发明的一种快速消除羟苯磺酸钙、酚磺乙胺药物干扰的肌酐试剂盒及其制备方法,其技术方案要点是:包括试剂R1和试剂R2,所述试剂R1中包含缓冲液、肌酸酶、肌氨酸氧化酶、抗坏血酸氧化酶、过氧化物酶或过氧化氢酶、血清白蛋白、非离子表面活性剂、防腐剂、Trinder’s反应剂A、漆酶或金属酸盐;所述试剂R2包含缓冲液、血清白蛋白、肌酐酶、防腐剂、及Trinder’s反应剂B。本发明克服现在各医院临床检验测定肌酐时,由于血清标本中含羟苯磺酸钙、酚磺乙胺造成的负偏差,使肌酐测定时不准确的缺陷,适用于全自动生化分析仪血清肌酐的检测。
Description
本案为分案申请,具体为申请号:2020108215100,名称为:一种可消除羟苯磺酸钙,酚磺乙胺的肌酐试剂盒及其制备方法,申请日为:2020.08.15的分案申请。
【技术领域】
本发明属于医学检验技术领域,本发明涉及一种快速消除羟苯磺酸钙、酚磺乙胺药物干扰的肌酐试剂盒及其制备方法。
【背景技术】
肌酐为临床检验上肾功能的主要指标及检测项目,测定血清肌酐的方法主要有两种:一种是碱性苦味酸速率法(Jaffe法),一种是肌氨酸氧化酶法;Jaffe法由于线性范围窄,特异性差,易受样本中其他物质(如酮体、抗坏血酸、胆红素、头孢、多巴胺等)干扰,出现检测偏差,已逐渐退出市场。目前市场使用较多的为肌氨酸氧化酶法,此法已被各医院逐渐淘汰弃用改由肌氨酸氧化酶法检测,其检测原理如下:
现行市售检测试剂盒均为双试剂,试剂R1内含肌酸酶,肌氨酸氧化酶先将血清中之肌酸分解产生H2O2,再以过氧化物酶及Trinder’s试剂与形成的H2O2反应生产无色化合物,或以过氧化氢酶用来消除血清肌酸产生的H2O2,将生成的H2O2分解成水和氧,然后加入R2试剂,R2试剂里含肌酐酶及另一种Trinder’s试剂,及叠氮钠,肌酐酶将肌酐水解成肌酸再由R1里的肌酸酶,肌氨酸氧化酶,过氧化物酶及Trinder’s试剂之4-AAP及另一种Trinder’s试剂苯酚类化合物共同存在下呈色进行测定,试剂R1里如含过氧化氢酶则被试剂R2内之叠氮钠反应掉,此反应过程均利用了酶的专一性,但生成的H2O2为强氧化剂,极易被血清中的还原性药物消耗而产生负干扰。
肌酐作为肾功能最重要的指标,在肾病造成的微血管病变治疗及保护上常用的羟苯磺酸钙常用药具极强的还原性可与H2O2直接反应,消耗掉H2O2;另在常用于预防及治疗外科手术出血过多的常用药酚磺乙胺,亦具有强的还原性,此两种药物均会消耗现今医院常用的肌酐酶法测定时产生的H2O2,造成肌酐值会有严重的负偏差,造成医师诊断治疗时的误判。
羟苯磺酸钙和酚磺乙胺对肌氨酸氧化酶法检测肌酐的负干扰对被施用这两种药物的患者造成很大的不便。例如,对于糖尿病患者而言,常伴发肾功能不全,而所施用的羟苯磺酸钙对肌氨酸氧化酶法检测肌酐的负干扰可能造成糖尿病患者肾功能不正确评估,掩盖并延误病情。而对于手术患者,所施用的酚磺乙胺对肌氨酸氧化酶法检测肌酐的负干扰可能使得不能正确评估其肾功能,造成术后并发症,影响患者的术后恢复。
【发明内容】
本发明公开一种快速消除羟苯磺酸钙、酚磺乙胺药物干扰的肌酐试剂盒,克服现在各医院临床检验测定肌酐时,由于血清标本中含羟苯磺酸钙、酚磺乙胺造成的负偏差,使肌酐测定时不准确,导致医师治疗过程中的误判的缺陷,试剂盒稳定性好,操作简单迅速,使测定准确,适用于全自动生化分析仪血清肌酐的检测。
本发明还公开一种快速消除羟苯磺酸钙、酚磺乙胺药物干扰的肌酐试剂盒的制备方法。
本发明的可消除血清中羟苯磺酸钙、酚磺乙胺药物引起肌酐测定负偏差干扰的肌酐试剂盒,包括独立的试剂R1和试剂R2,所述试剂 R1中包含缓冲液、肌酸酶、肌氨酸氧化酶、抗坏血酸氧化酶、过氧化物酶或过氧化氢酶、血清白蛋白、非离子表面活性剂、防腐剂、Trinder’s反应剂A、漆酶或金属酸盐;所述试剂R2包含缓冲液、血清白蛋白、肌酐酶、防腐剂、可含亦可不含非离子表面活性剂,及Trinder’s反应剂B。漆酶可利用氧(O2)将羟苯磺酸钙及酚磺乙胺氧化成苯醌类化合物;上述金属酸盐或五氧化二钒或三氯氧钒作为氧化剂可将羟苯磺酸钙,酚磺乙胺氧化,避免消耗掉在肌酐测定时产生的H2O2。上述R1试剂中,漆酶或氧化态金属酸盐或五氧化二钒或三氯氧钒组分亦可由试剂R1中剥离出来,单独一并使用前再加入成为试剂R1。
优选的,试剂R1中有时也会包含抗坏血酸氧化酶,抗坏血酸氧化酶的量在5-10KU/L,优选5.5-10KU/L的范围内,抗坏血酸氧化酶的量在上述范围内时可以有效地降低抗坏血酸对肌酐测定所造成的干扰,在一些具体实施方式中,所述抗坏血酸氧化酶的量可以为5.5KU/L、 6.4KU/L、7.5KU/L、8.8KU/L、8.6KU/L、6KU/L、7.2KU/L、9.2KU/L、 9.5KU/L或10KU/L。
优选的,所述漆酶是Sigma-Alderich公司产品号SAE0050之漆酶。所述漆酶是一种含铜的多酚氧化还原酶,可利用水中的氧分子氧化多种苯酚类化合物生成醌类化合物,氧分子本身被还原成水,为污水处理中常用于环境友好酶用来消除污水中的酚类化合物。羟苯磺酸钙及酚磺乙胺均为含苯酚环结构之还原剂,漆酶可以将其氧化消除;但漆酶一般在PH2~4时活力最高,稳定性较好,而肌酐酶法试剂之PH值须在7~9,本专利试验不同来源之漆酶后,最终筛选出 Sigma-Alderich公司产品号SAE0050之漆酶,其在PH7.0~9.0时有很好之活性及稳定性,可加入肌酐酶R1试剂中用以消除羟苯磺酸钙及酚磺乙胺之干扰。
优选的,所述金属酸盐包括氧化态镍酸盐、铁酸盐、钴酸盐、铬酸盐、锰酸盐、钒酸盐,所述钒酸盐包括多聚钒酸盐、偏钒酸盐、五氧化二钒或三氯氧钒一种或一种以上混合。本发明采用元素周期表中23、24、25、26、27、28位各金属元素酸盐,具体选择氧化还原电位高的氧化态镍酸盐、钴酸盐、铬酸盐、锰酸盐、钒酸盐,所述钒酸盐包括偏钒酸盐、多聚钒酸盐、五氧化二钒或三氯氧钒均可做为氧化剂来消除肌酐测定时血清中羟苯磺酸钙及酚磺乙胺的干扰造成的负偏差,同时不影响试剂R1及试剂R2中各组分的稳定性。
优选的,所述R1中缓冲液浓度为5~200mM,PH7.0~9.0;肌酸酶含量5~50ku/升,肌氨酸氧化酶含量5~50ku/升,抗坏血酸氧化酶1~10ku/升,过氧化物酶含量1~10ku/升,或含过氧化氢酶100~500ku/升,血清蛋白1~2克/升,Trinder’s反应剂A 0.5~5mM/升[所述的Trinder’s反应剂A为4-氨基安替比林 (4-Aminoantipyrine,简称4-AAP)];防腐剂重量比含量 0.005%-0.2%,如采用漆酶消除羟苯磺酸钙及酚磺乙胺之负干扰时,漆酶含量0.1~10ku/升;如采用上述金属酸盐氧化剂或五氧化二钒或三氯氧钒氧化剂时则单一种或混合二种及以上时其含量在0.01~10mM;所述的高氧化还原电位金属酸盐、包含所述含金属酸的钠、钾、铵盐。
优选的,所述R1的缓冲液为BES、BICINE、DIPSO、EPPS、PIPES、 HEPES、MOPS、TAPS、TAPSO、TES、Tricine、Tris、双甘肽、磷酸盐中的一种或一种以上混合;试剂R2所述的缓冲液为BES、BICINE、 DIPSO、EPPS、PIPES、HEPES、MOPS、TAPS、TAPSO、TES、TRICINE、 TRTS、双甘肽、磷酸盐中的一种或一种以上混合。
优选的,所述试剂R1及试剂R2中还含有表面活性剂,其重量比含量为0.01%~0.3%,所述表面活性剂为非离子型表面活性剂,具体为TRITON、TWEEN、BRIJ、EMULGEN中的一种或两种。
优选的,所述防腐剂为Proclin、硫酸庆大霉素、PARABEN、氯霉素、叠氮钠之一种或两种。
优选的,所述R2中,缓冲液浓度为10~200mM、PH6.5~9.0;Trinder’s反应剂B含量为0.1~20mM;其肌酐酶之含量50~10 00ku /升。
优选的,所述试剂R2内之Trinder’s反应剂B为TOOS、DHBS、 DAOS、HDAOS、TBHBA、TOPS、TODB、4-氯酚的一种或两种以上混合。这些组分与Trinder’s反应试剂各种苯酚或苯胺类衍生物混合,生成色原组分,Trinder’s反应剂A必须在试剂R1内,Trinder’s反应剂B必须在试剂R2内。
试剂R1及试剂R2,其中试剂R1中包括消除羟苯磺酸钙及酚磺乙胺干扰的漆酶,可以在加入肌酐酶(试剂R2)之前先将血清中的羟苯磺酸钙及酚磺乙胺氧化避免其消耗在肌酐测定时产生的H2O2,并在R2试剂中加入叠氮钠,于试剂R1与血清中成份反应完成后加入试剂R2,将漆酶杀灭,亦将过氧化氢酶杀灭,避免了肌酐酶加入后对肌酐测定的影响。
一种可消除血清中羟苯磺酸钙、酚磺乙胺药物引起肌酐测定负偏差干扰的肌酐试剂盒的制备方法,包括如下步骤:
制备R1试剂:
称取缓冲液→加入防腐剂→加入漆酶或金属酸盐,搅拌1小时以IN盐酸或氢氧化钠(或钾)调至PH8.0→加入Trinder’s反应剂A→非离子表面活性剂→加入BSA重量比0.1%搅拌1小时→放入2~8℃隔夜→再加入肌酸酶,肌氨酸氧化酶,过氧化物酶或过氧化氢酶,抗坏血酸氧化酶→搅拌1小时→检测后获得;所述金属酸盐为氧化态镍酸盐、铁酸盐、钴酸盐、铬酸盐、锰酸盐、钒酸盐中的一种或一种以上混合,所述的Trinder’s反应剂A为4 -氨基安替比林;
制备R2试剂:
称取缓冲液→以IN盐酸或氢氧化钠(或钾)调至PH8.0→加入Trinder’s反应剂B→加入防腐剂→加入非离子表面活性剂→血清白蛋白→搅拌1小时→放冷,2~8℃隔夜→再加入肌酐酶后,检测后获得。
由试剂R1及试剂R2组成,以试剂R1先与标本混合,除了先将标本中的肌酸消除,如标本中含治疗药物羟苯磺酸钙或酚磺乙胺时,试剂 R1中的漆酚或上述氧化态金属酸盐或五氧化二钒或三氯氧钒可先将其氧化,避免了加入试剂R2中含有的肌酐酶经系列反应产生的H2O2被上述药物消耗掉,同时R2试剂中含有Trinder’s反应剂B,即苯酚类化合物与试剂R1中4-AAP与肌酐酶经系列反应产生的H2O2呈色,测定其吸光度,计算其含量。
本发明的有益效果是:
本申请的可消除血清中羟苯磺酸钙、酚磺乙胺药物引起肌酐测定负偏差干扰的肌酐试剂盒,目前市面上所有肌氨酸氧化酶法测定肌酐时均采用双试剂及Trinder’s反应来测定,均在试剂R1中含Trinder’s反应剂常用之苯酚或苯胺类化合物,如4-氯酚, TOOS,DHBS,DAOS,HDAOS,TBHBA,TOPS,TODB在R2试剂中含4-AAP,本发明在一系列试验中发现必须将4-AAP加在R1试剂中才可与漆酶或上述金属酸盐或五氧化二钒及三氯氧钒及R1试剂中其他组分共同存在,Trinder’s反应另一试剂苯酚类化合物必须与肌酐酶共同存在R2中,才可使本肌酐试剂盒除了可消除羟苯磺酸钙及酚磺乙胺的干扰外,才可使本试剂盒在保持2~8℃一年以上的稳定。
本申请的可消除血清中羟苯磺酸钙、酚磺乙胺药物引起肌酐测定负偏差干扰的肌酐试剂盒,试剂R1中含过氧化氢酶或过氧化物酶与 4-AAP共同存在下,可将血清标本中的肌酸先由试剂R1中肌酸酶及肌氨酶氧化酶产生的H2O2先行分解消除血清中肌酸的干扰,R1试剂中亦含抗坏血酸氧化酶,先将血清中抗坏血酸消除,避免抗坏血酸造成的干扰,所得试剂盒稳定性好,操作简单迅速,消除了羟苯磺酸钙及酚磺乙胺的负干扰,准确度高,避免医师在临床诊断时的误判,可以精准用药及治疗,适用于全自动生化分析仪,以双试剂上机,操作简单,安全可靠。
本申请的可消除血清中羟苯磺酸钙、酚磺乙胺药物引起肌酐测定负偏差干扰的肌酐试剂盒的制备方法,方法独特,工艺简便,适合批量制备作业。
【附图说明】
附图1:为吸光度测试过程示意。
【具体实施方式】
下面结合具体实施例对本发明的可消除血清中羟苯磺酸钙、酚磺乙胺药物引起肌酐测定负偏差干扰的肌酐试剂盒做进一步描述:
以目前市售具中国国家药监局批准之肌酐酶法试剂盒测定羟苯磺酸钙及酚磺乙胺之干扰,结果如表一:
其试剂组成依说明书所述为:
[主要组成成份]由试剂R1、试剂R2和校准品组成。
试剂R1:三羟甲基氨基甲烷(Tris)缓冲液、肌酸脒基水解酶、肌氨酸氧化酶、抗坏血酸氧化酶、2-羟基-3-间甲苯胺丙磺酸钠(TOOS)、过氧化氢酶、表面活性剂;
试剂R2:三羟甲基氨基甲烷(Tris)缓冲液、肌酐氨基水解酶、 4-氨基安替比林、过氧化物酶。
校准品:含肌酐水溶液,浓度见标签,可溯源至Randox CAL3校准品。
[测定方法]
(1)双试剂无需配制,直接使用;
(2)试验条件:样本(S):10μl,试剂1(R1):210μl,试剂2(R2): 70μl,温度:37℃;
测定类型:终点法,主波长:546nm,副波长:660nm,反应方向:上升;
方法:先将样本与R1混合,37℃5分钟后加入R2试剂,然后测定加入R2后5分钟的反应吸光度。
(3)校准程序:使用校准品进行校准,每次更换试剂批次时都应进行校准。校准后,各实验室要用质控品验证。如果质控结果不在可接受范围值内,则需要进行重新校准。
测定仪器及参数:
单位:μmol/L,正常值≤70μmol/L
测定仪器:日立7180
测定参数:R1:R2:S(标本量)=210:70:10μl
主/副波长:540/660nm
2POINT END,INC.
表1现有双试剂肌酐酶法试剂盒测定干扰结果:
结论:测定结果显示羟苯磺酸钙及酚磺乙胺均对肌酐测定结果造成严重的负偏差。
具体结合实施例1-4说明本申请的可消除血清中羟苯磺酸钙、酚磺乙胺药物引起肌酐测定负偏差干扰的肌酐试剂盒的作用:
表2实施例1-4的试剂组分:
依照本发明的肌酐试剂具体组份描述如下(所述实施例中百分比为重量百分比):
实施例1:
试剂R1组份:
试剂R2组份:
实施例2:
试剂R1组份:
试剂R2组份:
实施例3:
试剂R1组份:
试剂R2组份:
实施例4:
试剂R1组份:
试剂R2组份:
实施例1-4的具体制备方法入下:
制备R1试剂:
称取缓冲液→加入防腐剂→加入漆酶或金属酸盐,搅拌1小时→以IN盐酸或氢氧化钠(或钾)调至PH8.0→加入Trinder’s反应剂A→非离子表面活性剂→加入BSA 重量比0.1%→搅拌1小时→放入2~8℃隔夜→再加入肌酸酶,肌氨酸氧化酶,过氧化物酶或过氧化氢酶,抗坏血酸氧化酶搅拌1小时→检测后获得;
制备R2试剂:
称取缓冲液→以IN盐酸或氢氧化钠(或钾)调至PH8.0 加入Trinder’s反应剂B→加入防腐剂→加入非离子表面活性剂→血清白蛋白→搅拌1小时→放冷,2~8℃隔夜再加入肌酐酶后,检测后获得。
测定方法:
将实施例1-4所得试剂R1及试剂R2放置于全自动凝血分析仪 (日立7180)相应试剂位置,吸取试剂R1 210μl,试剂R2 70μl,定标液或标本10μl,定标液浓度300μM,主/副波长540/660nm,二点终点法,反应方向:INC,以定标液定标测定含不同浓度羟苯磺酸钙或酚磺乙胺各标本肌酐值。
表3:实施例1测定结果:(单位:μmol/L)
测定 | 1 | 2 | 3 | 平均值 | 相对偏差 |
母液 | 139 | 140 | 139 | 139.30 | |
羟苯磺酸钙8ml/L | 141 | 137 | 138 | 138.70 | ﹣0.95% |
羟苯磺酸钙16ml/L | 141 | 141 | 140 | 140.7 | 1.0% |
羟苯磺酸钙32ml/L | 141 | 141 | 139 | 140.30 | 0.13% |
羟苯磺酸钙64ml/L | 139 | 138 | 138 | 138.3 | ﹣0.017% |
酚磺乙胺12.5mg/L | 140 | 137 | 139 | 138.70 | ﹣0.15% |
酚磺乙胺25mg/L | 141 | 140 | 139 | 140.00 | 0.15% |
酚磺乙胺50mg/L | 139 | 140 | 138 | 144.00 | ﹣0.2% |
酚磺乙胺100mg/L | 139 | 138 | 137 | 138 | ﹣1.0% |
表4:实施例2测定结果:(单位:μmol/L)
测定 | 1 | 2 | 3 | 平均值 | 相对偏差 |
母液 | 148 | 146 | 147 | 147.01 | |
羟苯磺酸钙8ml/L | 149 | 148 | 147 | 148.01 | 0.7% |
羟苯磺酸钙16ml/L | 149 | 147 | 148 | 148.01 | 0.7% |
羟苯磺酸钙32ml/L | 148 | 148 | 149 | 151.01 | 1.0% |
羟苯磺酸钙64ml/L | 147 | 148 | 148 | 152.01 | 0.5% |
酚磺乙胺12.5mg/L | 147 | 146 | 147 | 146.70 | ﹣0.2% |
酚磺乙胺25mg/L | 148 | 147 | 147 | 147.30 | 0.2% |
酚磺乙胺50mg/L | 147 | 148 | 148 | 147.70 | 0.5% |
酚磺乙胺100mg/L | 147 | 148 | 146 | 147.0 | 0.0% |
表5:实施例3测定结果:(单位:μmol/L)
测定 | 1 | 2 | 3 | 平均值 | 相对偏差 |
母液 | 152 | 152 | 152 | 152.01 | |
羟苯磺酸钙8ml/L | 154 | 153 | 152 | 153.01 | 0.7% |
羟苯磺酸钙16ml/L | 152 | 152 | 152 | 152.01 | 0.0% |
羟苯磺酸钙32ml/L | 152 | 152 | 151 | 151.7 | -0.2% |
羟苯磺酸钙64ml/L | 152 | 151 | 151 | 151.3 | -0.5% |
酚磺乙胺12.5mg/L | 153 | 153 | 153 | 153.01 | 0.7% |
酚磺乙胺25mg/L | 155 | 153 | 153 | 153.7 | 1.1% |
酚磺乙胺50mg/L | 155 | 153 | 153 | 153.7 | 1.1% |
酚磺乙胺100mg/L | 153 | 154 | 153 | 153.3 | 0.9% |
表6:实施例4测定结果:(单位:μmol/L)
上面各实施例1-4测定结果表明,本申请的可消除血清中羟苯磺酸钙、酚磺乙胺药物引起肌酐测定负偏差干扰的肌酐试剂盒,由试剂 R1及试剂R2组成,以试剂R1先与标本混合,除了先将标本中的肌酸消除,如标本中含治疗药物羟苯磺酸钙或酚磺乙胺时,试剂R1中的漆酚或上述氧化态金属酸盐或五氧化二钒或三氯氧钒可先将其氧化,采用漆酶或氧化还原电位高的氧化态的高铁酸盐、多聚偏钒酸盐及偏钒酸盐,可有效的消除肌酐测定时羟苯磺酸钙及酚磺乙胺的干扰,得到准确的肌酐测定值,避免了加入试剂R2中含有的肌酐酶经系列反应产生的H2O2被上述药物消耗掉,同时R2试剂中含有Trinder’s 反应剂B,即苯酚类化合物与试剂R1中4-AAP与肌酐酶经系列反应产生的H2O2呈色,测定其吸光度,计算其含量;
试剂R1中含过氧化氢酶或过氧化物酶与4-AAP共同存在下,可将血清标本中的肌酸先由试剂R1中肌酸酶及肌氨酶氧化酶产生的H2O2先行分解消除血清中肌酸的干扰,所得试剂盒稳定性好,操作简单迅速,消除了羟苯磺酸钙及酚磺乙胺的负干扰,准确度高,避免医师在临床诊断时的误判,可以精准用药及治疗,适用于全自动生化分析仪,以双试剂上机,操作简单,安全可靠。采用漆酶或氧化还原电位高的氧化态的高铁酸盐、多聚偏钒酸盐及偏钒酸盐,均可有效的消除肌酐测定时羟苯磺酸钙及酚磺乙胺的干扰,得到准确的肌酐测定值。
Claims (7)
1.一种快速消除羟苯磺酸钙、酚磺乙胺药物干扰的肌酐试剂盒,其特征在于:包括试剂R1和试剂R2,所述试剂R1中包含缓冲液、肌酸酶、肌氨酸氧化酶、抗坏血酸氧化酶、过氧化物酶、血清白蛋白、非离子表面活性剂、防腐剂、Trinder’s反应剂A、漆酶或铬酸盐或铁酸盐或五氧化二钒;所述试剂R2包含缓冲液、血清白蛋白、肌酐酶、防腐剂、Trinder’s反应剂B;所述的Trinder’s反应剂A为4-氨基安替比林,所述Trinder’s反应剂B为TOOS、DHBS、DAOS、HDAOS、TBHBA、TOPS、TODB、4-氯酚的一种或两种以上混合;所述试剂R1及试剂R2中还含有表面活性剂,其重量比含量为0.01%~0.3%,所述表面活性剂为非离子型表面活性剂,具体为TRITON、TWEEN、BRIJ、EMULGEN中的一种或两种。
2.根据权利要求1所述的一种快速消除羟苯磺酸钙、酚磺乙胺药物干扰的肌酐试剂盒,其特征在于:所述漆酶是Sigma-Alderich公司产品号SAE0050之漆酶。
3.根据权利要求1所述的一种快速消除羟苯磺酸钙、酚磺乙胺药物干扰的肌酐试剂盒,其特征在于:
所述R1中,缓冲液浓度为5~200mM,PH7.0~9.0;
肌酸酶含量5~50ku/升;
肌氨酸氧化酶含量5~50ku/升;
过氧化物酶含量1~10ku/升;
抗坏血酸氧化酶1~10ku/升;
血清蛋白1~2克/升;
Trinder’s反应剂A 0.5~5mM/升;
防腐剂重量比为0.005%-0.2%时,漆酶含量0.1~10ku/升,或采用含量在0.01~10mM的铬酸盐或铁酸盐或五氧化二钒。
4.根据权利要求1所述的一种快速消除羟苯磺酸钙、酚磺乙胺药物干扰的肌酐试剂盒,其特征在于:所述R1的缓冲液为BES、BICINE、DIPSO、EPPS、PIPES、HEPES、MOPS、TAPS、TAPSO、TES、Tricine、Tris、双甘肽、磷酸盐中的一种或一种以上混合;试剂R2所述的缓冲液为BES、BICINE、DIPSO、EPPS、PIPES、HEPES、MOPS、TAPS、TAPSO、TES、TRICINE、TRTS、双甘肽、磷酸盐中的一种或一种以上混合。
5.根据权利要求1所述的一种快速消除羟苯磺酸钙、酚磺乙胺药物干扰的肌酐试剂盒,其特征在于:所述防腐剂为Proclin、硫酸庆大霉素、PARABEN、氯霉素、叠氮钠之一种或两种。
6.根据权利要求1所述的一种快速消除羟苯磺酸钙、酚磺乙胺药物干扰的肌酐试剂盒,其特征在于:所述R2中,缓冲液浓度为10~200mM、PH6.5~9.0;Trinder’s反应剂B含量为0.1~20mM;其肌酐酶之含量50~10 00ku/升。
7.一种快速消除羟苯磺酸钙、酚磺乙胺药物干扰的肌酐试剂盒的制备方法,其特征在于包括如下步骤:
制备R1试剂:
称取缓冲液→加入防腐剂→加入漆酶或铬酸盐或铁酸盐或五氧化二钒,搅拌1小时→以IN盐酸或氢氧化钠(或氢氧化钾)调至PH8.0→加入Trinder’s反应剂A→非离子表面活性剂→加入BSA重量比0.1%→搅拌1小时→放入2~8℃隔夜→再加入肌酸酶,肌氨酸氧化酶,过氧化物酶,抗坏血酸氧化酶搅拌1小时→检测后获得;所述的Trinder’s反应剂A为4-氨基安替比林;
制备R2试剂:
称取缓冲液→以IN盐酸或氢氧化钠(或氢氧化钾)调至PH8.0加入Trinder’s反应剂B→加入防腐剂→加入非离子表面活性剂→血清白蛋白→搅拌1小时→放冷,2~8℃隔夜再加入肌酐酶后,所述Trinder’s反应剂B为TOOS、DHBS、DAOS、HDAOS、TBHBA、TOPS、TODB、4-氯酚的一种或两种以上混合,检测后获得试剂。
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