大鼠抗心肌抗體(AMA)ELISA試劑盒產(chǎn)品規(guī)格：96T/48T
產(chǎn)品庫存：現(xiàn)貨
產(chǎn)品價(jià)格：詢價(jià)
使用范圍：僅供科研檢測(cè)，不得用于臨床。
保存條件：2-8℃
發(fā)貨方式：專門快遞免費(fèi)送貨上門
產(chǎn)品價(jià)格：*，洽談！
大鼠抗心肌抗體(AMA)ELISA試劑盒說明書:說明書隨貨發(fā)送，您也可以直接我司在線銷售人員索取。全國(guó)：
代理多種品牌進(jìn)口原裝、分裝“ELISA試劑盒”。齊一生物科技有限公司作為酶聯(lián)免疫供應(yīng)商，憑借*的技術(shù)，*的售前，售中，售后免費(fèi)代測(cè)服務(wù)，共同鑄就高品質(zhì)的產(chǎn)品。多年專業(yè)酶免服務(wù)，質(zhì)量保證，可免費(fèi)提供代測(cè)服務(wù)，一周內(nèi)出結(jié)果。咨詢訂購。
ELISA試劑盒操作步驟
ELISA試劑盒使用前，將所有試劑充分混勻。不要使液體產(chǎn)生大量的泡沫，以免加樣時(shí)加入大量的氣泡，產(chǎn)生加樣上的誤差。
根據(jù)待測(cè)樣品數(shù)量加上標(biāo)準(zhǔn)品的數(shù)量決定所需的板條數(shù)。每個(gè)標(biāo)準(zhǔn)品和空白孔建議做復(fù)孔。每個(gè)樣品根據(jù)自己的數(shù)量來定，能使用復(fù)孔的盡量做復(fù)孔。標(biāo)本用標(biāo)本稀釋液1：1稀釋后加入50ul于反應(yīng)孔內(nèi)。
加入稀釋好后的標(biāo)準(zhǔn)品50ul于反應(yīng)孔、加入待測(cè)樣品50ul于反應(yīng)孔內(nèi)。立即加入50ul的生物素標(biāo)記的抗體。蓋上膜板，輕輕振蕩混勻，37℃溫育1小時(shí)。
甩去孔內(nèi)液體，每孔加滿洗滌液，振蕩30秒，甩去洗滌液，用吸水紙拍干。重復(fù)此操作3次。如果用洗板機(jī)洗滌，洗滌次數(shù)增加一次。
每孔加入80ul的親和鏈酶素-HRP，輕輕振蕩混勻，37℃溫育30分鐘。
甩去孔內(nèi)液體，每孔加滿洗滌液，振蕩30秒，甩去洗滌液，用吸水紙拍干。重復(fù)此操作3次。如果用洗板機(jī)洗滌，洗滌次數(shù)增加一次。
每孔加入底物A、B各50ul，輕輕振蕩混勻，37℃溫育10分鐘。避免光照。
取出酶標(biāo)板，迅速加入50ul終止液，加入終止液后應(yīng)立即測(cè)定結(jié)果。
在450nm波長(zhǎng)處測(cè)定各孔的OD值。
ELISA操作步驟
酶聯(lián)免疫吸附實(shí)驗(yàn)材料，試劑，溶液   
1．酶標(biāo)板，100μltip頭，1ml Ep管，濕盒
2．包被稀釋液（0.05mol/L碳酸鈉-碳酸氫鈉buffer,pH 9.6）
  碳酸鈉0.15g,碳酸氫鈉0.29g,疊氮鈉0.02g,加雙蒸水至100ml,調(diào)至pH     9.6
3.  封閉液(5%小牛血清/PBS溶液)
    小牛血清5ml, 1*PBS(pH 7.4)95ml
4.  洗滌液(PBST, pH 7.4)
    NaCl 0.8g, KH2PO40.02g,Na2HPO4.12H2O 0.29g, KCl 0.02g,Tween20 0.05ml,疊氮鈉0.01g,加雙蒸水至100ml,調(diào)至pH 7.4
5.  樣本稀釋液(PBS, pH 7.4)
     NaCl 0.8g, KH2PO40.02g,Na2HPO4.12H2O 0.29g, KCl 0.02g,疊氮鈉0.01g,加雙蒸水至100ml,調(diào)至pH 7.4
6.  酶標(biāo)第2抗體(羊抗兔)稀釋范圍1:5,000-1:100,000
7.  底物液(TMB-過氧化氫尿素溶液)
① 底物液A:TMB20mg,無水乙醇10ml,加雙蒸水至100ml.
②底物液B(0.1mol/L檸檬酸-0.2mol/L磷酸二氫鈉緩沖液, pH 5.0-5.4)
Na2HPO41.46g, 檸檬酸0.933g, 0.75%過氧化氫尿素0.64ml, 加三蒸水至100ml,調(diào)至pH 5.0-5.4
③底物A和B按1:1混合即成TMB-過氧化氫尿素溶液.
8. 終止液(2mol/LH2SO4溶液)雙蒸水200ml,濃硫酸34ml,(緩慢滴加并不斷攪拌),加雙蒸水至300ml
9. 0.9%生理鹽水

大鼠抗心肌抗體(AMA)ELISA試劑盒操作步驟: 
1.     包被過程(注意設(shè)置空白對(duì)照,陰性對(duì)照):
將所用抗原用包被稀釋液稀釋到適當(dāng)濃度(一般所需抗原包被量為每孔20-200μg),每孔抗原加入100μl, 置37℃,4h,或4℃,24h;棄去孔中液體.(為避免蒸發(fā),板上應(yīng)加蓋或?qū)迤椒旁诘撞坑袧窦啿嫉慕饘贊窈兄?
2.     封閉酶標(biāo)反應(yīng)孔:
5%小牛血清置37℃封閉40min.封閉時(shí)將封閉液加滿各反應(yīng)孔,并去除各孔中的氣泡,封閉結(jié)束后用洗滌液滿孔洗滌3遍,每遍3min.
洗滌方法:吸干孔內(nèi)反應(yīng)液,將洗滌液注滿板孔,放置2min略作搖動(dòng),吸干孔內(nèi)液,傾去液體后在吸水紙上拍干         
        洗滌次數(shù)3次
3.     加入待檢測(cè)樣品(建立合適的濃度梯度):
檢測(cè)時(shí)一般采用1:50-1:400的稀釋度, 應(yīng)采用較大稀釋體積進(jìn)行,一般保證樣品吸取量>20μl.
將稀釋好的樣品加入酶標(biāo)反應(yīng)孔中,每樣品至少加雙孔, 每孔100μl,置于37℃,40-60min.用洗滌液滿孔洗滌3遍,每遍3min.
4.     加入酶標(biāo)抗體:
酶標(biāo)抗體: 根據(jù)酶結(jié)合物提供商提供的參考工作稀釋度進(jìn)行. 37℃,30-60min之間.短于30min往往結(jié)果不穩(wěn)定.每孔加100μl.洗滌同前。
5.     加入底物液(現(xiàn)用現(xiàn)配):
*TMB-過氧化氫尿素溶液,OPD-過氧化氫底物液系統(tǒng)次之.
底物加入量:每孔100μl,置37℃避光放置3-5分鐘,加入終止液顯色.
6.     終止反應(yīng):
每孔加入終止液50μl終止反應(yīng),于20min內(nèi)測(cè)定實(shí)驗(yàn)結(jié)果.
7.     結(jié)果判斷:
OPD顯色后采用492nm波長(zhǎng),TMB反應(yīng)產(chǎn)物檢測(cè)需要450nm波長(zhǎng).檢測(cè)時(shí)一定要首*行空白孔系統(tǒng)調(diào)零.用測(cè)定標(biāo)本孔的吸收值與一組陰性標(biāo)本測(cè)定孔平均值的比值(P/N)表示,當(dāng)P/N大于2時(shí)作為抗體的效價(jià).(數(shù)值的大小依具體檢測(cè)要求而定.)

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