Selected article for: "room temperature and TMB substrate"
Author: Lee, Yu-Ching; Tsai, Keng-Chang; Leu, Sy-Jye; Wang, Tuan-Jen; Liu, Chia-Yu; Yang, Yi-Yuan
Title: Isolation, Characterization, and Molecular Modeling of a Rheumatoid Factor from a Hepatitis C Virus Infected Patient with Sjögren's Syndrome
  • Document date: 2013_12_30
    • ID: 0zsn4lu3_16
    Snippet: Competitive inhibition assays were carried out to determine the binding affinities of Fc-binding Fab molecules. In brief, microtitre plates were coated with human Fc fragment (0.5 g/well) and blocked with 5% skim milk. After one hour incubation at room temperature, plates were washed with PBST twice. Each purified Fab was incubated first with various concentrations of soluble Fc fragment diluted in PBS (ranging from 2.5 to 80 g/mL) at room temper.....
    Document: Competitive inhibition assays were carried out to determine the binding affinities of Fc-binding Fab molecules. In brief, microtitre plates were coated with human Fc fragment (0.5 g/well) and blocked with 5% skim milk. After one hour incubation at room temperature, plates were washed with PBST twice. Each purified Fab was incubated first with various concentrations of soluble Fc fragment diluted in PBS (ranging from 2.5 to 80 g/mL) at room temperature for 1 hr. Then, the mixtures were added to the wells to react with coated human Fc fragment. After incubating at room temperature for 1 hr, plates were washed with PBST six times. Thereafter, HRP-conjugated goat anti-human or light chain antibodies (Jackson ImmunoResearch Laboratories) were incubated at room temperature for 1 hr to detect bound Fab protein. After washing with PBST six times, TMB substrate (Sigma) was added to each well for development. Reaction was stopped with 1 N HCl and signal intensity was measured at OD 450 nm.

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