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Please according to use instruction strictly, The test result determination must take the microtiter plate reader as a standard. Use new disposal plastic pipette tips and Closure plate membrane for each standard, in order to avoid cross contamination. Do not mix reagents with those from other lots. The substrate evade the light preservation. Specimen requirements extract as soon as possible after Specimen collection,and according to the relevant literature, and should be experiment as soon as possible after the extraction. If it can t, specimen can be kept in -20 ! to preserve, Avoid repeated freeze-thaw cycles. Can t detect the sample which contain NaN3, because NaN3 inhibits HRP active. Assay procedure Dilute and add sample:Dilute Original density Standard as follow table: 800U/L5 Standard120l Original density Standard+120l Standard diluent400U/L4 Standard120l 5 Standard+120l Standard diluent200U/L3 Standard120l 4 Standard+120l Standard diluent100U/L2 Standard120l 3 Standard +120l Standard diluent50U/L1 Standard120l 2 Standard +120l Standard diluent2. according testing Sample numbers to define how many wells nedd, Standard and blank suggested Do holes. 3.add sample1) blank wells: (blank comparison wells don t add sample , Biotinylated anti  TM -antibody and Streptavidin-HRP ,other each step operation is same); 2) Standard wells: add Standard 50l and Streptavidin-HRP 50l; 3) testing Sample wells: add sample 40l,then add anti  TM -antibody 10l , Streptavidin-HRP 50l. closing plate with Closure plate membrane ,incubate for 60 min at 37!. 4.Configurate liquid: 30-fold(or 20-fold) wash solution diluted 30-fold (or 20-fold) with distilled water and reserve. 5.washingUncover 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.colorAdd Chromogen Solution A 50ul and Chromogen Solution B to each well, evade the light preservation for 10 min at 37! 7.Stop the reactionAdd Stop Solution50l to each well, Stop the reaction(the blue color change to yellow color). 8.assaytake blank well as zero , Read absorbance at 450nm after Adding Stop Solution and within 10min. 9. Calculate of result Steps description Standard, Sample diluent Add Standard, Sample diluent, Biotinylated and HRP ,incubate for 60 min at 37!. Wash 5 times,Add Chromogen Solution A and B, incubate for 15 min at 37!. Add Stopp Solution Read absorbance at 450nm within 15 min calculate Assay range 30U/L!800U/L Storage and validity 1Storage 2-8!. 2validity six months.     PAGE  PAGE 1 AMEKO TEL:+86-021-55785280 557585281 FAX:+86-021-55785279 E-mail:biolzy@yahoo.cn QYՋBR 7hTThQT ReQQY}Yv7hTThQT uir }hvNbTvhՋBR 37!S^60R mg5!k ReQ>frmA0B 37!>fr15R ReQ~bkm 15RQOD

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