抗副流感病毒IgG抗體(anti-PIVIgG)ELISA檢測試劑盒本試劑盒僅供研究使用檢測范圍:歡迎或或索取原版說明書zui低檢測限:歡迎或或索取原版說明書,全國:歡迎選購!
抗副流感病毒IgG抗體(anti-PIVIgG)ELISA檢測試劑盒類別:酶聯(lián)免疫分析檢測試劑盒
規(guī)格:96T/48T
包裝:盒
檢測原理:采用雙抗夾心法
保質(zhì)期:六個月
產(chǎn)品保存條件:2-8°C
應(yīng)用范圍:僅供科研研究實驗使用
樣品形式:血清/血漿/組織勻漿/細胞培養(yǎng)上清//其它生物液體(具體情況請咨詢)
標本的采集及保存:
1.血清:室溫血液自然凝固10-20分鐘后,離心20分鐘左右(2000-3000轉(zhuǎn)/分)。仔細收集上清。保存 過程 中 如有沉淀形成,應(yīng)再次離心。
2.血漿:應(yīng)根據(jù)標本的要求選擇EDTA、檸檬酸鈉或肝素作為抗凝劑,混合10-20分鐘后,離心20分鐘左右(2000-3000轉(zhuǎn)/分)。仔細收集上清。保存過程中如有沉淀形成,應(yīng)再次離心。
3.尿液:用無菌管收集。離心20分鐘左右(2000-3000轉(zhuǎn)/分)。仔細收集上清。保存過程中如有沉淀形成,應(yīng)再次離心。胸腹水、腦脊液參照此實行。
4.細胞培養(yǎng)上清:檢測分泌性的成份時,用無菌管收集。離心20分鐘左右(2000-3000轉(zhuǎn)/ 分)。仔細收集上清。 檢測細胞內(nèi)的成份時,用PBS(PH7.2-7.4)稀釋細胞懸液,細胞濃度達到100萬/ml左右。通過反復(fù)凍融,以 使細胞破壞并 放出細胞內(nèi)成份。離心20分鐘左右(2000-3000轉(zhuǎn)/分)。仔細收集上清。保存過程中如有沉淀形成,應(yīng)再次離心。
5.組織標本:切割標本后,稱取重量。加入一定量的PBS,PH7.4。用液氮迅速冷凍保存?zhèn)溆?。標本融化后仍然保?-8℃的溫度。加入一定量的PBS(PH7.4),用手工或勻漿器將標本勻漿充分。離心20分鐘左右(2000-3000轉(zhuǎn)/分)。仔細收集上清。分裝后一份待檢測,其余冷凍備用。
標本要求:
1.標本采集后盡早進行提取,提取按相關(guān)文獻進行,提取后應(yīng)盡快進行實驗。若不能馬上進行試驗,可將標本放于-20℃保存,但應(yīng)避免反復(fù)凍融
2.不能檢測含NaN3的樣品,因NaN3抑制辣根過氧化物酶的(HRP)活性。
試劑盒組成:
48孔配制:
1. 說明書: 1份
2. 封板膜: 2片(48)
3. 密封袋: 1個
4 酶標包被板: 1×48 (2-8℃保存)
5. 標準品: 0.5ml×1瓶(2-8℃保存)
6. 標準品稀釋液: 1.5ml×1瓶(2-8℃保存)
7. 酶標試劑: 3 ml×1瓶(2-8℃保存)
8. 樣品稀釋液: 3 ml×1瓶(2-8℃保存)
9. 顯色劑A液: 3 ml×1瓶(2-8℃保存)
10 .顯色劑B液: 3 ml×1瓶(2-8℃保存)
11 .終止液:3ml×1瓶(2-8℃保存)
12. 濃縮洗滌液:(20ml×20倍)×1瓶(2-8℃保存)
96孔配置:
1. 說明書:1份
2. 封板膜:2片(96)
3. 密封袋: 1個
4. 酶標包被板:1×96 (2-8℃保存)
5. 標準品:0.5ml×1瓶(2-8℃保存)
6. 標準品稀釋液:1.5ml×1瓶(2-8℃保存)
7. 酶標試劑: 3 ml×1瓶(2-8℃保存)
8. 樣品稀釋液: 3 ml×1瓶(2-8℃保存)
9. 顯色劑A液:3 ml×1瓶(2-8℃保存)
10. 顯色劑B液:3 ml×1瓶(2-8℃保存)
11. 終止液:3ml×1瓶(2-8℃保存)
12. 濃縮洗滌液:(20ml×20倍)×1瓶(2-8℃保存)
需要自備的試劑和器材:
1.37℃恒溫箱。
2.標準規(guī)格酶標儀。
3.精密移液器及一次性吸頭
4.蒸餾水,
5.一次性試管
6.吸水紙
操作步驟:
1、標準品的稀釋:在酶標包被板上設(shè)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9mmol/L,6 mmol/L,3 mmol/L,1.5mmol/L, 0.75 mmol/L)。
2、加樣:分別設(shè)空白孔(空白對照孔不加樣品及酶標試劑,其余各步操作相同)、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,盡量不觸及孔壁,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(fù)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(yīng)(此時藍色立轉(zhuǎn)黃色)。
11、測定:以空白空調(diào)零,450nm波長依序測量各孔的吸光度(OD值)。 測定應(yīng)在加終止液后15分鐘以內(nèi)進行。
洗板方法:
手工洗板方法:甩掉酶標板內(nèi)的液體;在實驗臺上鋪墊幾層吸水紙,酶標板朝下用力拍幾次;將稀釋后的洗滌液少0.35ml注入孔內(nèi),浸泡1-2分鐘。根據(jù)需要,重復(fù)此過程數(shù)次。
自動洗板:如果有自動洗板機,應(yīng)在熟練使用后再用到正式實驗過程中。
注意事項:
試劑盒從冷藏環(huán)境中取出應(yīng)在室溫平衡15-30分鐘后方可使用,酶標包被板開封后如未用完,板條應(yīng)裝入密封袋中保存。
濃洗滌液可能會有結(jié)晶析出,稀釋時可在水浴中加溫助溶,洗滌時不影響結(jié)果。
各步加樣均應(yīng)使用加樣器,并經(jīng)常校對其準確性,以避免試驗誤差。一次加樣時間控制在5分鐘內(nèi),如標本數(shù)量多,推薦使用排槍加樣。
請每次測定的同時做標準曲線,做復(fù)孔。如標本中待測物質(zhì)含量過高(樣本OD值大于標準品孔*孔的OD值),請先用樣品稀釋液稀釋一定倍數(shù)(n倍)后再測定,計算時請zui后乘以總稀釋倍數(shù)(×n×5)。
封板膜只限一次性使用,以避免交叉污染。
底物請避光保存。
嚴格按照說明書的操作進行,試驗結(jié)果判定必須以酶標儀讀數(shù)為準.
所有樣品,洗滌液和各種廢棄物都應(yīng)按傳染物處理。
本試劑不同批號組分不得混用。
如與英文說明書有異,以英文說明書為準。
計算:
以標準物的濃度為橫坐標,OD值為縱坐標,
在坐標紙上繪出標準曲線,根據(jù)樣品的OD
值由標準曲線查出相應(yīng)的濃度;再乘以稀釋
倍數(shù);或用標準物的濃度與OD值計算出標
準曲線的直線回歸方程式,將樣品的OD值
代入方程式,計算出樣品濃度,再乘以稀釋
倍數(shù),即為樣品的實際濃度。
檢測范圍、 OD值:
具體請咨詢在線客服,或者來電索要詳細說明書!
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