人轉鐵蛋白(TRF)ELISA試劑盒

英文名：Human transferrin (TRF) ELISA kit

規    格  ：96T/48T

保存條件：2-8℃   

產  地  ：國產/進口

elisa試劑盒操作步驟

　　1. 標準品的稀釋與加樣：在酶標包被板上設標準品孔10孔，在*、第二孔中分別加標準品100μl，然后在*、第二孔中加標準品稀釋液50μl，混勻;然后從*孔、第二孔中各取100μl分別加到第三孔和第四孔，再在第三、第四孔分別加標準品稀釋液50μl，混勻;然后在第三孔和第四孔中先各取50μl棄掉，再各取50μl分別加到第五、第六孔中，再在第五、第六孔中分別加標準品稀釋液50ul，混勻;混勻后從第五、第六孔中各取50μl分別加到第七、第八孔中，再在第七、第八孔中分別加標準品稀釋液50μl，混勻后從第七、第八孔中分別取50μl加到第九、第十孔中，再在第九第十孔分別加標準品稀釋液50μl，混勻后從第九第十孔中各取50μl棄掉。(稀釋后各孔加樣量都為50μl，濃度分別為1200pg/ml，800pg/ml ，400pg/ml，200pg/ml，100pg/ml)。

　2.加樣：分別設空白孔(空白比照孔不加樣品及酶標試劑，其他各步操作相同)、待測樣品孔。在酶標包被板上待測樣品孔中先加樣品稀釋液40μl，然后再加待測樣品10μl(樣品zui終稀釋度為5倍)。加樣將樣品加于酶標板孔底部，盡量不涉及孔壁，輕輕晃動混勻。

　3.溫育：用封板膜封板后置37℃溫育30分鐘。

　4.配液：將30(48T的20倍)倍濃縮洗滌液用蒸餾水30(48T的20倍)倍稀釋后備用。

　5.洗滌：小心揭掉封板膜，棄去液體，甩干，每孔加滿洗滌液，靜置30秒后棄去，如此重復5次，拍干。

　6.加酶：每孔加入酶標試劑50μl，空白孔除外。

　7.溫育：操作同3。

　8.洗滌：操作同5。

　9.顯色：每孔先加入顯色劑A50μl，再加入顯色劑B50μl，輕輕震蕩混勻，37℃避光顯色15分鐘.

　10.終止：每孔加終止液50μl，終止反應(此時藍色立轉黃色)。

　11.測定：以空白空調零，450nm波長依序丈量各孔的吸光度(OD值)。 測定應在加終止液后15分鐘以內進行。Procedure elisa kit
A standard sample dilution and added : In the ELISA plate coated with standard holes 10 holes , add Standard 100μl in the first and second holes , then add the standard in the first and second holes dilution 50μl, mix ; then from the first hole , the second hole from each 100μl were added to the third hole and the fourth hole , and then in the third and fourth holes add Standard dilution 50μl, mix ; then on the third hole in the first and fourth holes from each 50μl discard, depicting 50μl were added to the fifth and sixth holes , then add Standard dilution 50ul in the fifth and sixth holes , mix ; mix, take from the fifth and sixth holes were added to 50μl seventh, eighth hole, then add Standard dilution 50μl in the seventh , eighth hole , mix from seventh after the first eight holes were taken 50μl added to the ninth , tenth hole, then add Standard dilution 50μl in the ninth and the tenth hole , mix, take 50μl discard from the ninth and tenth holes. (Diluted sample volume of each hole are 50μl, concentrations were 1200pg/ml, 800pg/ml, 400pg/ml, 200pg/ml, 100pg/ml).
2 plus sample: Set blank wells ( blank cf hole not add sample and enzyme reagent, other each step operation ) , the sample holes . In the sample well microtiter plates coated before adding the sample dilution 40μl, then add the sample 10μl ( sample final dilution is 5 -fold ) . Add sample to the bottom of the wells, and try not to involve the cell wall , and Gently mix .
3 incubation : After Closure plate membrane seal plates were incubated at 37 ℃ for 30 minutes.
4 with liquid: 30 (48T 20- fold) concentrate was washed with distilled water and 30 (48T 20- fold) diluted spare .
5 Washing : Beware Uncover Closure plate membrane , discard liquid, drying , washing liquid fill every hole , after standing for 30 seconds, drain , repeat 5 times , and pat dry.
6 enzyme : enzyme reagent added to each well , except 50μl, empty holes.
7 incubation : Operation with 3.
8 Washing : Operation with 5.
9 color : each hole to join the chromogenic agent A50μl, then add color reagent B50μl, gently mixing shock , 37 ℃ dark color 15 minutes .
10 Termination: each well stop solution 50μl, termination of the reaction (the blue color change to yellow ) .
11 Determination : take blank zero , 450nm wavelength sequentially measuring the absorbance of each well (OD) . Measurement should be carried out after adding the stop solution within 15 minutes.