操作步驟：
1. 標(biao)準(zhun)(zhun)品的稀(xi)(xi)釋與加(jia)(jia)樣：在(zai)酶(mei)標(biao)包被(bei)板上(shang)設標(biao)準(zhun)(zhun)品孔(kong)(kong)(kong)(kong)(kong)(kong)10孔(kong)(kong)(kong)(kong)(kong)(kong)，在(zai)*、第(di)(di)(di)(di)(di)(di)二孔(kong)(kong)(kong)(kong)(kong)(kong)中(zhong)(zhong)分(fen)(fen)別(bie)(bie)加(jia)(jia)標(biao)準(zhun)(zhun)品100μl，然后(hou)在(zai)*、第(di)(di)(di)(di)(di)(di)二孔(kong)(kong)(kong)(kong)(kong)(kong)中(zhong)(zhong)加(jia)(jia)標(biao)準(zhun)(zhun)品稀(xi)(xi)釋液50μl，混(hun)勻(yun)；然后(hou)從(cong)(cong)*孔(kong)(kong)(kong)(kong)(kong)(kong)、第(di)(di)(di)(di)(di)(di)二孔(kong)(kong)(kong)(kong)(kong)(kong)中(zhong)(zhong)各取(qu)(qu)100μl分(fen)(fen)別(bie)(bie)加(jia)(jia)到(dao)(dao)第(di)(di)(di)(di)(di)(di)三(san)孔(kong)(kong)(kong)(kong)(kong)(kong)和第(di)(di)(di)(di)(di)(di)四孔(kong)(kong)(kong)(kong)(kong)(kong)，再(zai)在(zai)第(di)(di)(di)(di)(di)(di)三(san)、第(di)(di)(di)(di)(di)(di)四孔(kong)(kong)(kong)(kong)(kong)(kong)分(fen)(fen)別(bie)(bie)加(jia)(jia)標(biao)準(zhun)(zhun)品稀(xi)(xi)釋液50μl，混(hun)勻(yun)；然后(hou)在(zai)第(di)(di)(di)(di)(di)(di)三(san)孔(kong)(kong)(kong)(kong)(kong)(kong)和第(di)(di)(di)(di)(di)(di)四孔(kong)(kong)(kong)(kong)(kong)(kong)中(zhong)(zhong)先各取(qu)(qu)50μl棄掉(diao)，再(zai)各取(qu)(qu)50μl分(fen)(fen)別(bie)(bie)加(jia)(jia)到(dao)(dao)第(di)(di)(di)(di)(di)(di)五(wu)、第(di)(di)(di)(di)(di)(di)六孔(kong)(kong)(kong)(kong)(kong)(kong)中(zhong)(zhong)，再(zai)在(zai)第(di)(di)(di)(di)(di)(di)五(wu)、第(di)(di)(di)(di)(di)(di)六孔(kong)(kong)(kong)(kong)(kong)(kong)中(zhong)(zhong)分(fen)(fen)別(bie)(bie)加(jia)(jia)標(biao)準(zhun)(zhun)品稀(xi)(xi)釋液50ul，混(hun)勻(yun)；混(hun)勻(yun)后(hou)從(cong)(cong)第(di)(di)(di)(di)(di)(di)五(wu)、第(di)(di)(di)(di)(di)(di)六孔(kong)(kong)(kong)(kong)(kong)(kong)中(zhong)(zhong)各取(qu)(qu)50μl分(fen)(fen)別(bie)(bie)加(jia)(jia)到(dao)(dao)第(di)(di)(di)(di)(di)(di)七、第(di)(di)(di)(di)(di)(di)八孔(kong)(kong)(kong)(kong)(kong)(kong)中(zhong)(zhong)，再(zai)在(zai)第(di)(di)(di)(di)(di)(di)七、第(di)(di)(di)(di)(di)(di)八孔(kong)(kong)(kong)(kong)(kong)(kong)中(zhong)(zhong)分(fen)(fen)別(bie)(bie)加(jia)(jia)標(biao)準(zhun)(zhun)品稀(xi)(xi)釋液50μl，混(hun)勻(yun)后(hou)從(cong)(cong)第(di)(di)(di)(di)(di)(di)七、第(di)(di)(di)(di)(di)(di)八孔(kong)(kong)(kong)(kong)(kong)(kong)中(zhong)(zhong)分(fen)(fen)別(bie)(bie)取(qu)(qu)50μl加(jia)(jia)到(dao)(dao)第(di)(di)(di)(di)(di)(di)九、第(di)(di)(di)(di)(di)(di)十(shi)孔(kong)(kong)(kong)(kong)(kong)(kong)中(zhong)(zhong)，再(zai)在(zai)第(di)(di)(di)(di)(di)(di)九第(di)(di)(di)(di)(di)(di)十(shi)孔(kong)(kong)(kong)(kong)(kong)(kong)分(fen)(fen)別(bie)(bie)加(jia)(jia)標(biao)準(zhun)(zhun)品稀(xi)(xi)釋液50μl，混(hun)勻(yun)后(hou)從(cong)(cong)第(di)(di)(di)(di)(di)(di)九第(di)(di)(di)(di)(di)(di)十(shi)孔(kong)(kong)(kong)(kong)(kong)(kong)中(zhong)(zhong)各取(qu)(qu)50μl棄掉(diao)。
2. 加(jia)(jia)樣(yang)(yang)：分別設空白(bai)孔(kong)（空白(bai)對照孔(kong)不(bu)加(jia)(jia)樣(yang)(yang)品(pin)(pin)(pin)及(ji)酶(mei)標(biao)(biao)試劑，其余各步操作(zuo)相(xiang)同）、待測(ce)樣(yang)(yang)品(pin)(pin)(pin)孔(kong)。在(zai)酶(mei)標(biao)(biao)包被板(ban)上待測(ce)樣(yang)(yang)品(pin)(pin)(pin)孔(kong)中先(xian)加(jia)(jia)樣(yang)(yang)品(pin)(pin)(pin)稀釋液40μl，然(ran)后再加(jia)(jia)待測(ce)樣(yang)(yang)品(pin)(pin)(pin)10μl（樣(yang)(yang)品(pin)(pin)(pin)zui終稀釋度為5倍）。加(jia)(jia)樣(yang)(yang)將樣(yang)(yang)品(pin)(pin)(pin)加(jia)(jia)于酶(mei)標(biao)(biao)板(ban)孔(kong)底部，盡量不(bu)觸及(ji)孔(kong)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(de)(de)20倍(bei)）倍(bei)濃縮洗滌液用蒸餾水30（48T的(de)(de)20倍(bei)）倍(bei)稀釋(shi)后備用。
5. 洗滌(di)：小心揭掉封板膜，棄去(qu)液(ye)體，甩干，每孔(kong)加滿(man)洗滌(di)液(ye)，靜置30秒后棄去(qu)，如此(ci)重復5次，拍干。 6. 加酶(mei)：每孔(kong)加入酶(mei)標試(shi)劑50μl，空白(bai)孔(kong)除外。
7. 溫育：操(cao)作同3。
8. 洗滌(di)：操作(zuo)同5。
9. 顯(xian)色(se)(se)：每孔先(xian)加入顯(xian)色(se)(se)劑(ji)A50μl，再加入顯(xian)色(se)(se)劑(ji)B50μl，輕輕震蕩混勻，37℃避光(guang)顯(xian)色(se)(se)15分鐘.
10. 終(zhong)止：每孔加(jia)終(zhong)止液(ye)50μl，終(zhong)止反應（此時藍色(se)立轉(zhuan)黃色(se)）。
11. 測(ce)定：以(yi)(yi)空白空調(diao)零，450nm波長依序(xu)測(ce)量各孔的吸光度（OD值）。 測(ce)定應在加終止液(ye)后15分(fen)鐘以(yi)(yi)內進行。
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