大鼠抗心肌抗体(AMA)ELISA试剂盒产品规格：96T/48T
产品库存：现货
产品价格：询价
使用范围：仅供科研检测，不得用于临床。
保存条件：2-8℃
发货方式：专门快递免费送货上门
产品价格：*，洽谈！
大鼠抗心肌抗体(AMA)ELISA试剂盒说明书:说明书随货发送，您也可以直接我司在线销售人员索取。全国：
代理多种品牌进口原装、分装“ELISA试剂盒”。齐一生物科技有限公司作为酶联免疫供应商，凭借*的技术，*的售前，售中，售后免费代测服务，共同铸就高品质的产品。多年专业酶免服务，质量保证，可免费提供代测服务，一周内出结果。咨询订购。
ELISA试剂盒操作步骤
ELISA试剂盒使用前，将所有试剂充分混匀。不要使液体产生大量的泡沫，以免加样时加入大量的气泡，产生加样上的误差。
根据待测样品数量加上标准品的数量决定所需的板条数。每个标准品和空白孔建议做复孔。每个样品根据自己的数量来定，能使用复孔的尽量做复孔。标本用标本稀释液1：1稀释后加入50ul于反应孔内。
加入稀释好后的标准品50ul于反应孔、加入待测样品50ul于反应孔内。立即加入50ul的生物素标记的抗体。盖上膜板，轻轻振荡混匀，37℃温育1小时。
甩去孔内液体，每孔加满洗涤液，振荡30秒，甩去洗涤液，用吸水纸拍干。重复此操作3次。如果用洗板机洗涤，洗涤次数增加一次。
每孔加入80ul的亲和链酶素-HRP，轻轻振荡混匀，37℃温育30分钟。
甩去孔内液体，每孔加满洗涤液，振荡30秒，甩去洗涤液，用吸水纸拍干。重复此操作3次。如果用洗板机洗涤，洗涤次数增加一次。
每孔加入底物A、B各50ul，轻轻振荡混匀，37℃温育10分钟。避免光照。
取出酶标板，迅速加入50ul终止液，加入终止液后应立即测定结果。
在450nm波长处测定各孔的OD值。
ELISA操作步骤
酶联免疫吸附实验材料，试剂，溶液   
1．酶标板，100μltip头，1ml Ep管，湿盒
2．包被稀释液（0.05mol/L碳酸钠-碳酸氢钠buffer,pH 9.6）
  碳酸钠0.15g,碳酸氢钠0.29g,叠氮钠0.02g,加双蒸水至100ml,调至pH     9.6
3.  封闭液(5%小牛血清/PBS溶液)
    小牛血清5ml, 1*PBS(pH 7.4)95ml
4.  洗涤液(PBST, pH 7.4)
    NaCl 0.8g, KH2PO40.02g,Na2HPO4.12H2O 0.29g, KCl 0.02g,Tween20 0.05ml,叠氮钠0.01g,加双蒸水至100ml,调至pH 7.4
5.  样本稀释液(PBS, pH 7.4)
     NaCl 0.8g, KH2PO40.02g,Na2HPO4.12H2O 0.29g, KCl 0.02g,叠氮钠0.01g,加双蒸水至100ml,调至pH 7.4
6.  酶标第2抗体(羊抗兔)稀释范围1:5,000-1:100,000
7.  底物液(TMB-过氧化氢尿素溶液)
① 底物液A:TMB20mg,无水乙醇10ml,加双蒸水至100ml.
②底物液B(0.1mol/L柠檬酸-0.2mol/L磷酸二氢钠缓冲液, pH 5.0-5.4)
Na2HPO41.46g, 柠檬酸0.933g, 0.75%过氧化氢尿素0.64ml, 加三蒸水至100ml,调至pH 5.0-5.4
③底物A和B按1:1混合即成TMB-过氧化氢尿素溶液.
8. 终止液(2mol/LH2SO4溶液)双蒸水200ml,浓硫酸34ml,(缓慢滴加并不断搅拌),加双蒸水至300ml
9. 0.9%生理盐水

大鼠抗心肌抗体(AMA)ELISA试剂盒操作步骤: 
1.     包被过程(注意设置空白对照,阴性对照):
将所用抗原用包被稀释液稀释到适当浓度(一般所需抗原包被量为每孔20-200μg),每孔抗原加入100μl, 置37℃,4h,或4℃,24h;弃去孔中液体.(为避免蒸发,板上应加盖或将板平放在底部有湿纱布的金属湿盒中)
2.     封闭酶标反应孔:
5%小牛血清置37℃封闭40min.封闭时将封闭液加满各反应孔,并去除各孔中的气泡,封闭结束后用洗涤液满孔洗涤3遍,每遍3min.
洗涤方法:吸干孔内反应液,将洗涤液注满板孔,放置2min略作摇动,吸干孔内液,倾去液体后在吸水纸上拍干         
        洗涤次数3次
3.     加入待检测样品(建立合适的浓度梯度):
检测时一般采用1:50-1:400的稀释度, 应采用较大稀释体积进行,一般保证样品吸取量>20μl.
将稀释好的样品加入酶标反应孔中,每样品至少加双孔, 每孔100μl,置于37℃,40-60min.用洗涤液满孔洗涤3遍,每遍3min.
4.     加入酶标抗体:
酶标抗体: 根据酶结合物提供商提供的参考工作稀释度进行. 37℃,30-60min之间.短于30min往往结果不稳定.每孔加100μl.洗涤同前。
5.     加入底物液(现用现配):
*TMB-过氧化氢尿素溶液,OPD-过氧化氢底物液系统次之.
底物加入量:每孔100μl,置37℃避光放置3-5分钟,加入终止液显色.
6.     终止反应:
每孔加入终止液50μl终止反应,于20min内测定实验结果.
7.     结果判断:
OPD显色后采用492nm波长,TMB反应产物检测需要450nm波长.检测时一定要首*行空白孔系统调零.用测定标本孔的吸收值与一组阴性标本测定孔平均值的比值(P/N)表示,当P/N大于2时作为抗体的效价.(数值的大小依具体检测要求而定.)

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