Mouse TNF-α(Tumor Necrosis Factor Alpha) ELISA Kit

产品货号:E-EL-M0049c

规格: / 96T / 48T
价格: / ¥2400 / ¥1650

货号:E-EL-M0049c

货期:3个工作日+顺丰周期

反应性: 小鼠

检测范围: 31.25~2000 pg/mL

灵敏度: 18.75 pg/mL

  • 详情
  • 文献(37)
  • 用途

    该试剂盒用于体外定量检测小鼠Serum, plasma and other biological fluids中天然及部分重组TNF-α浓度

    检测原理

    本试剂盒采用双抗体夹心ELISA法。用抗小鼠TNF-α抗体包被于酶标板上,实验时样品或标准品中的小鼠TNF-α会与包被抗体结合,游离的成分被洗去。依次加入生物素化的抗小鼠TNF-α抗体和辣根过氧化物酶标记的亲和素。抗小鼠TNF-α抗体与结合在包被抗体上的小鼠TNF-α结合、生物素与亲和素特异性结合而形成免疫复合物,游离的成分被洗去。加入显色底物(TMB),TMB在辣根过氧化物酶的催化下呈现蓝色,加终止液后变成黄色。用酶标仪在450nm波长处测OD值,TNF-α浓度与OD450值之间呈正比,通过绘制标准曲线计算出样品中TNF-α的浓度。

    反应类型 夹心法
    规格 96T
    反应时间 3.5h
    反应性 小鼠
    检测方法 Colormetric
    检测范围 31.25—2000 pg/mL
    灵敏度 18.75 pg/mL
    样本体积 100μL
    样本类型 Serum, plasma and other biological fluids

    特异性

    可检测样本中的小鼠TNF-α,且与其它相关蛋白无明显交叉反应。

    重复性

    板内,板间变异系数均<10%。

    典型数据

    由于实验操作条件的不同(如操作者、移液技术、洗板技术和稳定条件等),标准曲线的OD值会有所差异。以下数据和曲线仅供参考,实验者需根据自己的实验建立标准曲线。

    (pg/mL) O.D Average Corrected
    2000 2.449
    2.491
    2.47 2.39
    1000 1.726
    1.752
    1.739 1.659
    500 0.993
    0.991
    0.992 0.912
    250 0.483
    0.509
    0.496 0.416
    125 0.3
    0.282
    0.291 0.211
    62.5 0.196
    0.168
    0.182 0.102
    31.25 0.128
    0.136
    0.132 0.052
    0 0.079
    0.081
    0.08 --

    精密度

    板内精密度:低浓度样本,中浓度样本和高浓度样本分别在1块板子上检测20次。
    板间精密度:低浓度样本,中浓度样本和高浓度样本分别在3块板子上检测20次。

    Intra-assay Precision Inter-assay Precision
    Sample 1 2 3 1 2 3
    n 20 20 20 20 20 20
    Mean
    (pg/mL)
    92.50 285.40 722.10 85.50 297.30 763.30
    Standard
    deviation
    6.00 16.60 33.90 4.50 11.90 41.20
    C V (%) 6.49 5.82 4.69 5.26 4.00 5.40

    回收率

    分别往5个不同样本中添加已知浓度的目标蛋白,做回收实验,得出回收率范围和平均回收率。

    Sample Type Range (%) Average Recovery (%)
    Serum (n=5) 94-111 101
    EDTA plasma (n=5) 96-110 102
    Cell culture media (n=5) 88-104 96

    线性

    分别往5个样本中添加已知浓度的目标蛋白,做回收实验,得出回收率范围及平均回收率。将5个样本分别稀释2倍,4倍,8倍,16倍做回收实验,得出回收率范围及平均回收率。

    Serum (n=5) EDTA plasma (n=5) Cell culture media (n=5)
    1:2 Range (%) 95-106 89-101 88-102
    Average (%) 101 94 94
    1:4 Range (%) 95-111 81-91 91-105
    Average (%) 103 86 98
    1:8 Range (%) 96-112 84-97 91-106
    Average (%) 103 91 98
    1:16 Range (%) 95-112 85-96 95-106
    Average (%) 103 91 101

    试剂盒组成及保存

    未拆封的试剂盒可在4℃保存一周;如果一周以后才使用试剂盒,请拆开试剂盒按照下表中的条件分别保存各组分。

    中文名称 英文名称 规格 保存条件
    ELISA酶标板(可拆卸) Micro ELISA Plate(Dismountable) 8 孔×12 条 -20℃,可存放6个月
    冻干标准品 Reference Standard 2 支
    浓缩生物素化抗体(100×) Concentrated Biotinylated Detection Ab 1支 120 μL
    浓缩HRP酶结合物(100×) Concentrated HRP Conjugate 1支 120 μL -20℃(避光),可存放6个月
    标准品&样品稀释液 Reference Standard & Sample Diluent 1瓶 20 mL 4℃.可存放6个月
    生物素化抗体稀释液 Biotinylated Detection Ab Diluent 1瓶 14 mL
    酶结合物稀释液 HRP Conjugate Diluent 1瓶 14 mL
    浓缩洗涤液(25×) Concentrated Wash Buffer (25×) 1瓶 30 mL
    底物溶液(TMB) Substrate Reagent 1瓶 10 mL 4℃(避光)
    反应终止液 Stop Solution 1瓶 10 mL 4℃
    封板覆膜 Plate Sealer 5 张
    产品说明书 Product Description 1 份
    质检报告 Certificate of Analysis 1 份

    说明:所有试剂瓶盖须旋紧以防止蒸发和微生物的污染。
    试剂体积以实际发货版说明书为准。相关试剂在分装时会比标签上标明的体积稍多一些,请在使用时量取而非直接倒出。

    试验所需自备物品

    • 1. 酶标仪(450nm波长滤光片)
    • 2. 高精度移液器,EP管及一次性吸头:0.5-10 μL, 2-20 μL, 20-200 μL, 200-1000 μL
    • 3. 37℃恒温箱,双蒸水或去离子水
    • 4. 吸水纸

    操作步骤

    1.对应板孔中加入100μL标准品工作液或样本,37℃孵育90分钟
    2.弃掉板内液体后,立即加入100μL生物素化抗体工作液,37℃孵育60分钟
    3.弃掉板内液体,洗板3次
    4.每孔加入100μL HRP酶结合物工作液,37℃孵育30分钟,弃掉板内液体,洗板5次
    5.每孔加入90μL底物溶液,37℃孵育15分钟
    6.每孔加入50μL终止液
    7.立即在450nm波长下读数,处理数据

    客户发表文献

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