The kit assay Human TNF-α level in the sample,use Purified Human TNF-α antibody tocoat microtiter plate wells, make solid-phase antibody, then add TNF-α to wells, Combined TNF-α antibody which With HRP labeled, become antibody - antigen - enzyme-antibody complex, after washing Completely, Add TMB substrate solution,TMB substrate becomes blue color At HRP enzyme-catalyzed, reaction is terminated by the addition of a sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450 nm. The concentration of TNF-α in the samples is then determined by comparing the O.D. of the samples to the standard curve. 人肿瘤坏死因子α(TNF-α)ELISA酶法试剂盒 样本提取方法Specimen requirements
1. serum- coagulation at room temperature 10-20 mins,centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant, If precipitation appeared, Centrifugal again.
2. plasma-use suited EDTA or citrate plasma as an anticoagulant,mix 10-20 mins ,centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant, If precipitation appeared, Centrifugal again.
Urine-collect sue a sterile container, centrifugation 20-min at the speed of 2000-3000 r.p.m.remove supernatant, If precipitation appeared, Centrifugal again. The Operation of Hydrothorax and cerebrospinal fluid Reference to it.
cell culture supernatant-detect secretory components, collect sue a sterile container,centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant,detect the composition of cells, Dilut cell suspension with PBS ( PH7.2-7.4 ) , Cell concentration reached 1 million / ml, repeated freeze-thaw cycles, damage cells and release of intracellular components, centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant, If precipitation appeared, Centrifugal again.
Tissue samples- After cutting samples, check the weight,add PBS(PH7.2-7.4), Rapidlyfrozen with liquid nitrogen, maintain samples at 2-8℃ after melting,add PBS (PH7.4 ),Homogenized by hand or Grinders, centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant.
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.