AE-200160-2 猪流行性乙型脑炎ELISA试剂盒Porcine Epidemic Encephalitis B Ab ELISA kit, 192 tests Control Disease (Serum)

Bring test kit to the room temperature (20-25 ℃) for at least 30 min, note that each reagent must be shaken evenly before use; put the required micro-well strips into plate frames. Resealed the unused microplate, stored at 2-8 ℃, not frozen.

1. Add 100 μL of the positive control serum, negative control serum and sample in duplicate wells. (Note: dilute the sample 40 times with the sample diluent (add 6 μL sample into 234 μL sample diluent).

2. Mix gently manually for 5-10 seconds. Cover the microplate with adhesive foil and incubate at 37°C for 30 min.

3. Aspirate the plates and tap dry on absorbent paper. Wash the plate 3 times with 300 ul wash buffer (1X) (note: (dilute the 20×concentrated washing buffer at 20 times to get 1X buffer with deionized water).

4. Add 100 μL enzyme conjugate to each well. Mix gently manually for 5-10seconds, cover the plate and incubate at 37 °C for 30 min. Aspirate the plates and tap dry on absorbent paper. Wash 3 times same as in step 3.

5. Add 50 μL of the substrate A solution followed by 50 μL of the B solution into each well. (Alternatively it is possible to mix Soln A and B in 1:1 ratio in a clean tube, mix, and add 100 ul of the mix in a single step). Mix gently manually for 5-10 seconds. Cover the plate and incubate at 25 °C for 15-20min in the dark (blue color develops in standards and samples).

6. Add 50 μL of the stop solution into each well. Mix gently manually for 5-10seconds . (blue color does not change after addition of stop solution). Read the plate at 630 nm to determine the OD value of every well within 5-10 min.

操作步骤:

将检测试剂盒置于室温(20-25℃)中至少30分钟，注意每种试剂在使用前必须均匀摇动，并放置所需的微孔分成板框。储存在2-8℃。

1.加入100μL的阳性对照血清、阴性对照血清，复孔取样。(注：用样品稀释剂稀释样品40次(将6μL样品加到2 34μL样品稀释剂中)。

2.用手轻轻搅拌5-10秒。用胶纸覆盖微板，在37°C下孵育30分钟。

3.抽吸轻拍吸水纸。用300μl的洗涤缓冲液(1x)洗3次(注：(将20倍的浓缩洗涤缓冲液稀释20倍，用去离子水得到1倍的缓冲液)。

4.每孔加入100μL酶结合物。轻轻搅拌5-10秒，盖上盖板，在37°C下孵育30分钟。抽吸微孔板，然后轻敲吸水纸上的干纸片。清洗3次，与步骤3相同。

5.在每孔中加入50μL的底物A溶液，然后再加入50μL的B溶液。(或者也可以将溶剂A和B按1：1的比例在干净的试管中混合，然后在一步中加入100μl的混合物)。轻轻搅拌5-10秒。盖上盘子，在25°C的黑暗中孵育15-20分钟(标准和样品中呈现蓝色)。

6.在每孔中加入50μL的终止液。用手轻轻搅拌5-10秒。(添加终止液后，蓝色不会改变)。在630 nm处读板，在5-10分钟内测定每口井的OD值。