For Research Use Only

仅供研究用

货号：QY-M30038

小鼠胰岛素(INS)酶联免疫分析(ELISA)

试剂盒使用说明书

 

本试剂仅供研究使用 目的：本试剂盒用于测定小鼠血清，细胞上清及相关液体样本中胰岛素(INS)的含量。

实验原理：

本试剂盒应用双抗体夹心法测定标本中小鼠胰岛素(INS)水平。用纯化的小鼠抗-胰岛素(INS)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胰岛素(INS),再与 HRP 标记的胰岛素（INS）抗体结合，形成抗体-抗原-酶标抗体复合物，经过*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zui终的黄色。颜色的深浅和样品中的胰岛素(INS)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小鼠胰岛素(INS)浓度。

Specimen requirements

N

Assay procedure

 

1.Dilute and add sample to Standard: set 10 Standard wells on the ELISA plates coated, add Standard 100μl to the first and the second well, then add Standard dilution 50μl to the first and the second well, mix; take out 100μl form the first and the second well then add it to the third and the forth well separay. then add Standard dilution 50μl to the third and the forth well ,mix ; then take out 50μl from the third and the forth well discard, add 50μl to the fifth and the sixth well ,then add Standard dilution 50μl to the fifth and the sixth well, mix ; take out 50μl from the fifth and the sixth well and add to the seventh and the eighth well, then add Standard dilution 50μl to the seventh and the eighth well ,mix ; take out 50μl from the seventh and the eighth well and add to the ninth and the tenth well, add Standard dilution 50μl to the ninth and the tenth well, mix , take out 50μl from the ninth and the tenth well discard(add Sample 50μl to each well after Diluting ,(density: 12 mU/L,8mU/L ,4 mU/L,2 mU/L, 1 mU/L)

2.add sample ： Set blank wells separay (blank comparison wells don’t add sample and HRP-Conjugate reagent, other each step operation is same). testing sample well. add Sample dilution 40μl to testing sample well, then add testing sample 10μl (sample final dilution is 5-fold), add sample to wells , don’t touch the well wall as far as possible, and Gently mix.

3.add enzyme：Add HRP-Conjugate reagent 50μl to each well, except blank well.

4.Incubate: After closing plate with Closure plate membrane ,incubate for 30 min at 37℃. 5.Configurate liquid: 30-fold (or 20-fold)wash solution diluted 30-fold (or 20-fold) with distilled water and reserve.

6.washing：Uncover Closure plate membrane, discard Liquid, dry by swing, add washing buffer to every well, still for 30s then drain, repeat 5 times, dry by pat.

7.color：Add Chromogen Solution A 50ul and Chromogen Solution B to each well, evade the light preservation for 10 min at 37℃

8.Stop the reaction ： Add Stop Solution50μl to each well, Stop the reaction(the blue color change to yellow color).

9.assay：take blank well as zero , Read absorbance at 450nm after Adding Stop Solution and within 15min.

Important notes

• The kit takes out from the refrigeration environment should be balanced 15-30 minutes in the room temperature, ELISA plates coated if has not use up after opened, the plate should be stored in Sealed bag.
• washing buffer will Crystallization separation, it can be heated the water helps dissolve when dilute . Washing does not affect the result.
• add Sample with sampler Each step, And proofread its accuracy frequently, avoids the experimental error. add sample within 5 min, if the number of sample is much , recommend to use Volley .
• if the testing material content is excessively higher (The sample OD is bigger than the first standard well ),please dilute Sample (n-fold), Please diluente and multiplied by the dilution factor.（×n×5）.
• Closure plate membrane only limits the disposable use, to avoid cross-contamination.
• The substrate evade the light preservation.
• Please according to use instruction strictly, The test result determination must take the microtiter plate reader as a standard.
• All samples, washing buffer and each kind of reject should according to infective material process.
• Do not mix reagents with those from other lots.

Assay range

0.3 mU/L -15 mU/L

 

Storage and validity

1．Storage： 2-8℃.

2．validity： six months.

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