The identification of antibody pair for using in sandwich immunoassays is a tiresome and time-consuming process. As an expert in developing and validating novel immunoassays, Scientists at Novatein Biosciences use the bead-based alphascreen method to screen and select the best complementary pair for sandwich immunoassays.
The whole process of antibody pair screening is divided in to three stages:
Selection of antibodies
Sensitivity, selectivity and overall performance of an immunoassay is largely dependent on the antibody selection and hence, this is the critical step in immunoassay development. We use our in-house developed antibodies or source them from a trusted commercial manufacturer. The client also has the flexibility of using their own hybridoma supernatants/ purified antibodies.
Conjugation of antibodies to the beads
The selected antibodies are conjugated to the acceptor and donor beads at saturating concentrations. Using a matrix approach, the selected antibodies will be tested against each other. Each antibody is captured on both donor and acceptor beads and complemented with its pairing antibody in the opposite bead.
Data collection and analysis
Assay will be performed by one of our expert scientists and read on BMG Pherastar. A color-coded matrix will be generated including the alphascreen counts (see below). The client will receive a detailed technical report along with our recommendation regarding the best antibody pair.
Antibody Pair screening to detect STING/TMEM173
A total of 3 antibodies were shortlisted for the screening process. After conjugating the antibodies to the acceptor and donor beads, alphalisa assay was performed and quantified using BMG Pherastar reader. A color-coded matrix was generated as shown below:
In Antibody 1A and Antibody 1D, 1 refers to the antibody# and A or D refer to conjugation to acceptor or donor beads respectively.
Using the above recommended antibody pair (Antibody 2 on acceptor beads and antibody 1 on donor beads) the assay range is determined to be 62.5 pg to 2000 pg/mL.
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