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 大鼠细胞间粘附分子3(ICAM-3/CD50)ELISA Kit
  • 大鼠细胞间粘附分子3(ICAM-3/CD50)ELISA Kit
  • 大鼠细胞间粘附分子3(ICAM-3/CD50)ELISA Kit

    参考价格: ¥3000.00
    品  牌:USA
    产品型号:RC01612B (大鼠细胞间粘附分子3(ICAM-3/CD50)ELISA Kit)
    所在地区:上海
    上架时间:2012年08月01日
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    RC01612B 大鼠细胞间粘附分子3(ICAM-3/CD50)ELISA 试剂盒 大鼠细胞间粘附分子3(ICAM-3/CD50)ELISA Kit
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    RC01626B 大鼠白介素13(IL-13)ELISA 试剂盒  大鼠白介素13(IL-13)ELISA Kit
    RC01627B 大鼠白介素15(IL-15)ELISA 试剂盒 大鼠白介素15(IL-15)ELISA Kit
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    PREPARATION OF REAGENTS
     Standards: Standard have to be reconstituled with the volume of standard buffer diluent indicated on the vial. This reconstitution produces a stock solution of 40ng/ml AFP-L3. Allow standard to stand for 5
     minutes with gentle swirling prior to making dilutions. Serial dilutions of standard must be made before each assys and cannot be stored.

    40 ng/ml (6  Standard) Original density 50ul。
    20 ng/ml (5  Standard) 100ul  6 Standard  +100ul diludent
    10 ng/ml (4  Standard) 100ul  5 Standard  +100ul diludent
    5.0 ng/ml (3  Standard) 100ul  4 Standard  +100ul diludent
    2.5 ng/ml (2  Standard) 100ul  3 Standard  +100ul diludent
    1.25 ng/ml (1  Standard) 100ul  2 Standard  +100ul diludent
    0 ng/ml Blank Control 50ul。

     Washing buffer 50×concentrate:  Dilute 50 times in distilled water.

    ASSAY METHOD
     Before use, mix all reagents thoroughly without making foam.
     Determine the number of microwell strips required to test the desired number of samples,plus appropriate number of wells needed for running blanks standards. Each sample, standard and blank should be assayed in duplicate. Remove sufficient microwell strips from the pouch.
     Add 50ul of standard diluent to standard wells B1,B2,  C1,C2,  D1,D2,  E1, E2,  F1,F2. Reconstitute standard vial with the appropriate volume as described in the chapter reagents preparation. Preparation. Pipet 100ul of standard into wells A1 and A2 (see plate scheme below). Transfer 50ul from A1 and A2 to B1 and B2 wells. Mix the contents by repeated aspirations and ejections. Take care not to scratch the inner surface of microwells. Repat this procedure from the wells B1,B2 to wells C1,C2 and from wells C1,C2 to D1,D2 and so on creating two parallel rows of AFP-L3 standard dilutions ranging,Add 50ul of standard diluent to the bland wells.
     Dilute samples 1:1 distribing 50ul of sample into 50ul of dilluent ,Add 50ul of diluted sample to wells..
     Add 50ul of diluted biotinylated anti-AFP-L3 to all wells.
     Cover with a plate vover and incubate for 1 hour at 37℃.
     Remove the cover and wash the plate as follows: ⑴ aspirate the liquid from each well, ⑵ dispensse 0.3ml of washing solution into each well. ⑶ Aspirate again the contet of each well after 0.5 minute. ⑷ Repeat steps ⑵ and ⑶ three times.
     Distribute 80ul of streptavidin-HRP solution to all wells, including blank wells.
     Cover and incubate 30 min at 37℃.
     Remove the cover and empty wells, Wash microwell strips according to step, Proceed immediately to the next step.
     Add 50ul Substrate A and Substrate B to each well。Incubate for 10 min at 37℃。
     The enzyme-substrate reaction is stopped by quickly pipetting 50ul of H2SO4. stop reagent into each well, including the blank wells, to completely and uniformly inactivate the enzyme. Results must be red immediately after the addition of H2SO4.
     Read absorbance of each well on a spectrophotometer using 450nm as the primary wavelength and optionally 620nm (610nm to 650nm is acceptable ) as the reference wavelength.

     

    SUGGESTED PLATE SCHEME
     Standard
    concentrations(ng/ml) 
    A 40 40 sample sample sample sample sample sample sample sample sample sample
    B 20 20 sample sample sample sample sample sample sample sample sample sample
    C 10 10 sample sample sample sample sample sample sample sample sample sample
    D 5.0 5.0 sample sample sample sample sample sample sample sample sample sample
    E 2.5 2.5 sample sample sample sample sample sample sample sample sample sample
    F 1.25 1.25 sample sample sample sample sample sample sample sample sample sample
    G 0 0 sample sample sample sample sample sample sample sample sample sample
    H sample sample sample sample sample sample sample sample sample sample sample sample

    LIMITATIONS OF THE PROCEDURE
    Do not extrapolate the standard curve beyond the max  standard curve point. The dose-response is non-linear in this region and good accruacy is difficult to obtain.

    CALCULATION OF RESULTS
    The minimum detectable concentration in this assay is estimated to be 0.1ng/ml