HIS(Histamine) ELISA Kit
组胺(HIS)酶联免疫吸附测定试剂盒
规格: | / 96T / 48T / 96T*5 |
价格: | / ¥2400 / ¥1650 / ¥10200 |
货号:E-EL-0032
反应性: 通用
检测范围: 1.56~100 ng/mL
灵敏度: 0.94 ng/mL
用途
该试剂盒用于体外定量检测血清、血浆或其他相关生物液体中HIS浓度。
检测原理
本试剂盒采用竞争ELISA法。用HIS抗原包被于酶标板上,实验时样品或标准品中的HIS与包被的HIS竞争生物素标记的抗HIS单抗上的结合位点,游离的成分被洗去。加入辣根过氧化物酶标记的亲和素,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,HIS浓度与OD450值之间呈反比,通过绘制标准曲线计算出样品中HIS的浓度。
反应类型 | 竞争法 |
规格 | 96T |
反应时间 | 2.0h |
反应性 | 通用 |
检测方法 | Colormetric |
检测范围 | 1.56-100 ng/mL |
灵敏度 | 0.94 ng/mL |
样本体积 | 50μL |
样本类型 | 血清、血浆或其他相关生物液体 |
特异性
可检测样本中的HIS,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。
(ng/mL) | O.D | Average |
---|---|---|
100 | 0.396 0.44 |
0.418 |
50 | 0.523 0.529 |
0.526 |
25 | 0.727 0.705 |
0.716 |
12.5 | 0.998 1.016 |
1.007 |
6.25 | 1.373 1.363 |
1.368 |
3.13 | 1.724 1.7 |
1.712 |
1.56 | 1.965 1.977 |
1.971 |
0 | 2.299 2.305 |
2.302 |
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 (ng/mL) |
5.17 | 13.32 | 45.10 | 5.30 | 13.54 | 42.52 |
Standard
deviation |
0.27 | 0.72 | 1.51 | 0.28 | 0.61 | 1.68 |
C V (%) | 5.22 | 5.41 | 3.35 | 5.28 | 4.51 | 3.95 |
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) | 90-100 | 95 |
EDTA plasma (n=5) | 87-100 | 93 |
Cell culture media (n=5) | 89-105 | 97 |
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 (%) | 88-103 | 88-100 | |
Average (%) | 95 | 95 | 94 | |
1:4 | Range (%) | 85-98 | 86-101 | 87-99 |
Average (%) | 91 | 92 | 94 | |
1:8 | Range (%) | 87-98 | 89-100 | 87-99 |
Average (%) | 93 | 95 | 92 | |
1:16 | Range (%) | 86-99 | 90-104 | 89-102 |
Average (%) | 92 | 97 | 96 |
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50μL标准品工作液或样本后,立即每孔加入50μL生物素化抗体工作液,37℃孵育90分钟 |
2. 弃掉板内液体,洗板3次 | |
3. 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 | |
4. 每孔加入90μL底物溶液,37℃孵育15分钟 | |
5. 每孔加入50μL终止液 | |
6. 立即在450nm波长下读数,处理数据 |
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