Browsing by keyword "Enzyme-linked immunosorbent assay"

Now showing items 1-3 of 3

    • Quantification of Murine AAT by Direct ELISA
      Cox, Andrew; Mueller, Christian (2017-07-28)
      This methods chapter elaborates on how a direct enzyme-linked immunosorbent assay (ELISA) is used to specifically detect and quantify murine alpha-1 antitrypsin (AAT). As a direct ELISA, it lacks some sensitivity as compared to the "sandwich" ELISA method; however, it does reliably differentiate between samples with varying amounts of the mouse AAT protein. This protocol relies on the principle of adsorption to coat each well with sera proteins, whereas detection occurs specifically using a two-step antibody combination. This procedure effectively identifies and quantifies murine AAT from a wide variety of samples including mouse serum, cell culture medium, and cell or tissue lysate.

    • Quantification of Total Human Alpha-1 Antitrypsin by Sandwich ELISA
      Tang, Qiushi; Gruntman, Alisha; Flotte, Terence R. (2017-07-28)
      In this chapter we describe an enzyme-linked immunosorbent assay (ELISA) to quantitatively measure human alpha-1 antitrypsin (AAT) protein levels in serum, other body fluids or liquid media. This assay can be used to measure the expression of the human AAT (hAAT) gene in a variety of gene transfer or gene downregulation experiments.A hAAT-specific capture antibody and a HRP-conjugated anti-AAT detection antibody are used in this assay. The conjugated anti-AAT used in this protocol, instead of the typical sandwich which employs an unconjugated antibody followed by a specifically conjugated IgG, makes the assay simpler and decreases variability. This provides a useful tool to evaluate the AAT levels in clinical and research samples and can allow fairly rapid testing of a large number of samples.

    • Quantification of Z-AAT by a Z-Specific "Sandwich" ELISA
      Borel, Florie; Tang, Qiushi; Mueller, Christian (2017-07-28)
      This protocol describes an enzyme-linked immunosorbent assay (ELISA) to specifically detect Z-alpha-1 antitrypsin (AAT), the most common protein variant associated with alpha-1 antitrypsin deficiency. This "sandwich" ELISA relies on an anti-Z-AAT specific capture antibody and a HRP-conjugated anti-AAT detection antibody. This method would be of interest to identify and quantify Z-AAT in a variety of samples such as cell culture medium, cell or tissue lysate, animal or patient serum. Because this method is specific and sensitive, it would be particularly valuable for detection of Z-AAT in the presence of background M-AAT, for instance when quantifying silencing of Z-AAT in patients undergoing M-AAT augmentation therapy.