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人纖溶酶原激活物抑制因子(PAI)ELISA檢測試劑盒實(shí)驗(yàn)所涉及到的儀器如下:酶標(biāo)儀、洗板機(jī)、離心機(jī)、培養(yǎng)箱
酶標(biāo)比色儀介紹:
酶標(biāo)比色儀簡稱酶標(biāo)儀,通常指于測讀ELISA結(jié)果吸光度的光度計(jì)。針對固相載體形式的不同,各有特制的適用于板、珠和小試管的設(shè)計(jì)。許多試劑公司配套供應(yīng)酶標(biāo)儀。酶標(biāo)儀的主要性能指標(biāo)有:測讀速度、讀數(shù)的準(zhǔn)確性、重復(fù)性、度和可測范圍、線性等等。優(yōu)良的酶標(biāo)儀的讀數(shù)一般可到0.001,準(zhǔn)確性為lusm1%,重復(fù)性達(dá)0.5%。舉例說,若某孔測得的A值為1.083,則該孔相對于空氣的真實(shí)A值應(yīng)為1.083lusm0.01(1.073~1.093),復(fù)測定數(shù)次,其A值均應(yīng)1.083lusm0.05(1.078~1.088)在之間。酶標(biāo)儀的可測范圍視各酶標(biāo)儀的性能而不同。普通的酶標(biāo)儀在0.000~2.000,新型號的酶標(biāo)儀上限拓寬達(dá)2.900,甚至更高。超出可測上限的A值常以"*"或"over"或其它符號表示。應(yīng)注意可測范圍與線性范圍的不同,線性范圍常小于可測范圍,比如某一酶標(biāo)儀的可測范圍為0.000~2.900,而其線性范圍僅0.000~2.000,這在定量ELISA中制作標(biāo)準(zhǔn)曲線時(shí)應(yīng)予注意。
酶標(biāo)儀不應(yīng)安置在陽光或強(qiáng)光照射下,操作時(shí)室溫宜在15~30℃,使用前先預(yù)熱儀器15-30分鐘,測讀結(jié)果更穩(wěn)定。
測讀A值時(shí),要選用產(chǎn)物的敏感吸收峰,如OPD用492nm波長。有的酶標(biāo)儀可用雙波長式測讀,即每孔先后測讀兩次,*次在zui適波長(W1),第二次在不敏感波長(W2),兩次測定間不移動ELISA板的位置。例如OPD用492nm為W1,630nm為W2,zui終測得的A值為兩者之差(W1-W2)。雙波長式測讀可減少由容器上的劃痕或指印等造成的光干擾。各種酶標(biāo)儀性能有所不同,使用中應(yīng)參考對應(yīng)儀器說明書。
人纖溶酶原激活物抑制因子(PAI)ELISA檢測試劑盒兩種設(shè)定空白對照的方案:
(1)將 TMB 空白顯色孔設(shè)為對照。所有的標(biāo)準(zhǔn)品和樣品的吸光值減去 TMB 空白顯色孔的 吸 光 值 后 , 在 坐 標(biāo) 紙 上 畫 出 曲 線 , 以 吸 光 值 作 為 縱 坐 標(biāo) , 以 濃 度 作 為 橫 坐 標(biāo) 。
(2)將零孔設(shè)為對照。所有的標(biāo)準(zhǔn)品和樣品的吸光值減去零孔的吸光值后,得到的數(shù)據(jù)可以 直接在坐標(biāo)紙上畫出曲線。
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