Oriscience Supersensitive ECL Kit 是一種基于辣根過氧化物酶(horseradish peroxidase, HRP)的極敏型ECL化學發光試劑盒。本試劑盒對化學發光液成分進行了優化，應用了不同于常見的魯米諾-H2O2體系的新型的增強劑和氧化劑，具有靈敏度極高，發光迅速，亮度強，穩定性好等特點，可使用較高的抗體稀釋倍數（1:5000~1:10000），因此在很大程度上節約抗體，且試劑中不含有毒成分，實驗安全性更高。
靈敏度更高
發光迅速，曝光時間更短
穩定性更好，發光信號持久
節約抗體，背景更低
安全性高
4℃避光保存
Oriscience Supersensitive ECL與某進口品牌超敏ECL對比圖
不同量的Hela細胞裂解液的Anti-actin Western Blotting結果（一抗稀釋倍數 1：10000，二抗稀釋倍數：1：10000）。除使用的ECL底物不同而外，其余實驗條件完全一致。
1. 在配置ECL工作液過程中吸取A液和B液時應避免交叉污染。
2. ECL工作液應在每次使用前新鮮配制，避免室溫放置時間過長。
3. 曝光時間過長或蛋白過量，將會導致過曝，使條帶強弱變化失去線性關系；曝光不足則條帶模糊。
4. ECL發光液是HRP的顯色底物，因此檢測對象必須由HRP標記。
5. 本產品僅供科研使用。請勿用于醫藥、臨床診斷或治療，食品及化妝品等用途。
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Q1：背景臟，信噪比低，出現非特異性條帶
a. 抗體特異性低：選用特異性更高的抗體。
b. 顯影時間過長：在確保獲得足夠強度的目標蛋白信號的前提下，縮短顯影時間。
c. 抗體孵育和洗膜過程中的污染：確保操作過程中所有試劑和用具的潔凈度。
d. 未進行非特異性封閉或封閉不充分：延長封閉時間，考慮更換合適的封閉劑。
e. 一抗濃度過高導致非特異性結合：稀釋抗體至合適濃度，以更高稀釋度抗體孵育更長時間（耗時長但特異性結合最好）。
f. 二抗與封閉劑非特異性結合：設置二抗對照(不加一抗)以明確，如確實存在此種情況，可在孵育和洗滌液中加入溫和去污劑，如吐溫20。
OriNote：脫脂奶粉含有酪蛋白，對于酪蛋白激酶，該蛋白本身就是一種磷酸化蛋白底物，有可能會結合磷酸化特異性抗體而易產生高背景。使用 BSA 代替奶粉作為封閉劑。
g. 膜干燥，導致高背景： 在孵育過程中防止膜變干，在任何步驟都保證膜有充分的反應液。
h. 未結合蛋白質洗滌不充分：增加洗滌次數。
Q2：目的條帶過曝
a. 減少蛋白上樣量。
b. 縮短曝光時間。
c. 用緩沖液適當洗滌后重新顯影。
d. 滴加ECL后放置一段時間再顯影。
Q3：目的條帶弱
a. 轉膜不充分：優化轉膜條件，提高轉膜效率。
b. 增加蛋白上樣量。
c. 抗體稀釋倍數過大：降低抗體稀釋倍數以提高其濃度。
d. 抗體使用時間過長導致抗體失效：重新配制抗體。
Q4：背景有不均勻的白色斑點
a. 轉膜時膜上有氣泡或抗體在膜上分布不均：轉膜過程中盡量去除氣泡，抗體孵育時保持搖動。
Q5：背景有黑色斑點
a. 抗體結合了封閉劑：過濾封閉劑。
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