小鼠內脂素/內髒脂肪素（visfatin）ELISA試劑盒

 小鼠內脂素/內髒脂肪素（visfatin）ELISA試劑盒含税含运费　, 
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    産品規格：96T/48T 
    96T指可以做94個標本，2個標准對照 
    48T指可以做47個標本，1個標准對照 
    96T-84個樣本 
    48T-42個樣本 
    操作步驟 
    1.標准品的稀釋：本試劑盒提供原倍標准品一支，用戶可按照下列圖表在小試管中進行稀釋。 
    3600 pg/ml5号標准品150μl的原倍標准品加入150μl標准品稀释液 
    1800pg/ml4號標准品150μl的5號標准品加入150μl標准品稀釋液 
    900pg/ml3號標准品150μl的4號標准品加入150μl標准品稀釋液 
    450 pg/ml2号標准品150μl的3号標准品加入150μl標准品稀释液 
    225pg/ml1號標准品150μl的2號標准品加入150μl標准品稀釋液 
     
     
    2.加樣：分別設空白孔（空白對照孔不加樣品及酶標試劑，其余各步操作相同）、標准孔、待測樣品孔。在酶標包被板上標准品准確加樣50μl，待測樣品孔中先加樣品稀釋液40μl，然後再加待測樣品10μl（樣品zui終稀釋度爲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：用封板膜封板後置37℃溫育30分鍾。 
    4.配液：將20倍濃縮洗滌液用蒸餾水20倍稀釋後備用 
    5.洗滌：小心揭掉封板膜，棄去液體，甩幹，每孔加滿洗滌液，靜置30秒後棄去，如此重複5次，拍幹。 
    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分鍾. 
    10.終止：每孔加終止液50μl，終止反應（此時藍色立轉黃色）。 
    11.测定：以空白空调零，450nm波长依序测量各孔的吸光度（OD值）。 测定应在加终止液后15分钟以内进行。 
    Assay procedure 
    1.Dilute and add sample:Dilute Original density Standard as follow table: 
    800 nmol/L5 Standard150μl Original density Standard+150μl Standard diluent 
    400 nmol/L4 Standard150μl 5 Standard+150μl Standard diluent 
    200 nmol/L3 Standard150μl 4 Standard+150μl Standard diluent 
    100 nmol/L2 Standard150μl 3 Standard +150μl Standard diluent 
    50 nmol/L1 Standard150μl 2 Standard +150μl Standard diluent 
    2.add sample：Set blank wells separay （blank comparison wells don’t add sample and HRP-Conjugate reagent, other each step operation is same）. testing sample well. add Sample dilution 40μl to testing sample well, then add testing sample 10μl （sample final dilution is 5-fold）, add sample to wells , don’t touch the well wall as far as possible, and Gently mix. 
    3.Incubate: After closing plate with Closure plate membrane ,incubate for 30 min at 37℃. 
    4.Configurate liquid: 30-fold wash solution diluted 30-fold （or 20-fold） with distilled water and reserve. 
    5.washing：Uncover Closure plate membrane, discard Liquid, dry by swing, add washing buffer to every well, still for 30s then drain, repeat 5 times, dry by pat. 
    6.add enzyme：Add HRP-Conjugate reagent 50μl to each well, except blank well. 
    7.incubate：Operation with 3. 
    8.washing：Operation with 5. 
    9.color：Add Chromogen Solution A 50ul and Chromogen Solution B to each well, evade the light preservation for 15 min at 37℃ 
    10.Stop the reaction：Add Stop Solution50μl to each well, Stop the reaction（the blue color change to yellow color）. 
    11.assay：take blank well as zero , Read absorbance at 450nm after Adding Stop Solution and within 15min. 
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