人抗利尿激素/血管加压素/精氨酸加压素（ADH/VP/AVP）ELIS..
人抗利尿激素/血管加压素/精氨酸加压素（ADH/VP/AVP）ELISA试剂盒说明
检测范围： 48T 25 ng/L -800 ng/L
使用目的：
本试剂盒用于测定人血清、血浆及相关液体样本抗利尿激素/血管加压素/精氨酸加压素（ADH/VP/AVP）含量。
实验原理
本试剂盒应用双抗体夹心法测定标本中人抗利尿激素/血管加压素/精氨酸加压素（ADH/VP/AVP）水平。用纯化的人抗利尿激素/血管加压素/精氨酸加压素（ADH/VP/AVP）抗体包被微孔板，制成固相抗体，往包被单抗的微孔中依次加入抗利尿激素/血管加压素/精氨酸加压素（ADH/VP/AVP），再与HRP标记的抗利尿激素/血管加压素/精氨酸加压素（ADH/VP/AVP）抗体结合，形成抗体-抗原-酶标抗体复合物，经过彻底洗涤后加底物TMB显色。TMB在HRP酶的催化下转化成蓝色，并在酸的作用下转化成最终的黄色。颜色的深浅和样品中的抗利尿激素/血管加压素/精氨酸加压素（ADH/VP/AVP）呈正相关。用酶标仪在450nm波长下测定吸光度（OD值），通过标准曲线计算样品中人抗利尿激素/血管加压素/精氨酸加压素（ADH/VP/AVP）浓度。
试剂盒组成
1 | 20倍浓缩洗涤液 | 20ml×1瓶 | 7 | 终止液 | 3ml×1瓶 |
2 | 酶标试剂 | 3ml×1瓶 | 8 | 标准品（1600ng/L） | 0.5ml×1瓶 |
3 | 酶标包被板 | 12孔×4条 | 9 | 标准品稀释液 | 1.5ml×1瓶 |
4 | 样品稀释液 | 3ml×1瓶 | 10 | 说明书 | 1份 |
5 | 显色剂A液 | 3ml×1瓶 | 11 | 封板膜 | 2张 |
6 | 显色剂B液 | 3ml×1/瓶 | 12 | 密封袋 | 1个 |
标本要求
1．标本采集后尽早进行提取，提取按相关文献进行，提取后应尽快进行实验。若不能马上进行试验，可将标本放于-20℃保存，但应避免反复冻融
2．不能检测含NaN3的样品，因NaN3抑制辣根过氧化物酶的（HRP）活性。
操作步骤
1. 标准品的稀释：本试剂盒提供原倍标准品一支，用户可按照下列图表在小试管中进行稀释。
800 ng/L | 5号标准品 | 150μl的原倍标准品加入150μl标准品稀释液 |
400 ng/L | 4号标准品 | 150μl的5号标准品加入150μl标准品稀释液 |
200 ng/L l | 3号标准品 | 150μl的4号标准品加入150μl标准品稀释液 |
100 ng/L | 2号标准品 | 150μl的3号标准品加入150μl标准品稀释液 |
50 ng/L | 1号标准品 | 150μl的2号标准品加入150μl标准品稀释液 |
2. 加 样：分别设空白孔（空白对照孔不加样品及酶标试剂，其余各步操作相同）、标准孔、待测样品孔。在酶标包被板上标准品准确加样50μl，待测样品孔中先加样 品稀释液40μl，然后再加待测样品10μl（样品最终稀释度为5倍）。加样将样品加于酶标板孔底部，尽量不触及孔壁，轻轻晃动混匀。
3. 温育：用封板膜封板后置37℃温育30分钟。
4. 配液：将20倍浓缩洗涤液用蒸馏水20倍稀释后备用
5. 洗涤：小心揭掉封板膜，弃去液体，甩干，每孔加满洗涤液，静置30秒后弃去，如此重复5次，拍干。
6. 加酶：每孔加入酶标试剂50μl，空白孔除外。
7. 温育：操作同3。
8.洗涤：操作同5。
9. 显色：每孔先加入显色剂A50μl，再加入显色剂B50μl，轻轻震荡混匀，37℃避光显色15分钟.
10.终止：每孔加终止液50μl，终止反应（此时蓝色立转黄色）。
11.测定：以空白空调零，450nm波长依序测量各孔的吸光度（OD值）。 测定应在加终止液后15分钟以内进行。
注意事项：
1．试剂盒从冷藏环境中取出应在室温平衡15-30分钟后方可使用，酶标包被板开封后如未用完，板条应装入密封袋中保存。
2．浓洗涤液可能会有结晶析出，稀释时可在水浴中加温助溶，洗涤时不影响结果。
3．各步加样均应使用加样器，并经常校对其准确性，以避免试验误差。一次加样时间控制在5分钟内，如标本数量多，推荐使用排枪加样。
4． 请每次测定的同时做标准曲线，做复孔。如标本中待测物质含量过高（样本OD值大于标准品孔孔的OD值），请先用样品稀释液稀释一定倍数（n倍）后再测定，计算时请最后乘以总稀释倍数（×n×5）。
5． 封板膜只限一次性使用，以避免交叉污染。
6．底物请避光保存。
7．严格按照说明书的操作进行，试验结果判定必须以酶标仪读数为准.
8．所有样品，洗涤液和各种废弃物都应按传染物处理。
9．本试剂不同批号组分不得混用。
10. 如与英文说明书有异，以英文说明书为准。
保存条件及有效期
1．试剂盒保存：；2-8℃。
2．有效期：6个月
The performance of kit:
1 sensitivity: minimum detection concentration is less than 1 standard. Linearity of dilution. Sample linear regression and the expected concentration correlation coefficient R value is 0.990.
2: no specific reaction with other cytokines.
3 repeatability: plate, plate between the coefficients of variation were less than 10%.
Human type III procollagen amino terminal peptide ELISA kit steps:
1 before use, all reagents and mixing. Do not allow liquid to produce a large number of bubbles, so as to avoid adding a large number of bubbles, resulting in the addition of the error.
2 according to determine the number of sample number plus standard strip number. Each standard and blank hole is recommended to do the hole. Each sample can be made according to its own quantity, and can be used as a hole in the hole.
3 diluted after standard 50ul in reaction hole, added to the sample 50 UL in reaction hole to be measured. Immediately joined the 50 UL antibody biotin. Cover the membrane plate, gently oscillating mixing, 37 degrees Celsius for 45 minutes.
4 left hole liquid, each hole filled with washing liquid, oscillating 30 seconds off the washing liquid, pat dry with absorbent paper. Repeat this operation 4 times. If the washing machine with washing, washing times increased once.
5 per hole adding chain affinity enzyme -HRP 100ul, gently oscillating mixing, 37 degrees 30 minutes incubation.
6 left hole liquid, each hole filled with washing liquid, oscillating 30 seconds off the washing liquid, pat dry with absorbent paper. Repeat this operation 4 times. If the washing machine with washing, washing times increased once.
7 per hole adding substrate A, B 50ul, gently oscillating mixing, 37 degrees 5 minutes incubation. Avoid light.
8 remove ELISA plate, quickly add 50ul terminated liquid, adding the stop solution immediately after the determination results.
9 od determination of each hole at the wavelength of 450nm.
The result of judgment and analysis:
1, instrument value: Yu Bo 450nm ELISA od read the hole on the instrument
2, to the OD value as a vertical coordinate (y), corresponding ot standard concentrations as a horizontal coordinate (x), do have corresponding curve, sample ot content can be according to the OD value by standard curve conversion out corresponding concentration, multiplied by the dilution multiple; or with the standard concentration and the OD value calculated the regression equation of the standard curve, the sample OD value in the equation to calculate the sample concentration, multiplied by the dilution factor is the actual concentration of the sample.
3, detection range: 0-100ng/ml
4, sensitivity: 0.39ng/ml
