Sandwich ELISA

Affimer® reagents to complement an existing antibody

Dependent upon assay requirements specific Affimer® reagents can be generated to complement an existing antibody within an assay, or as Affimer® pairs for the detection and capture of antigens of interest within a sandwich ELISA format.

Affimer® proteins have been engineered to be robust to the assay environment, demonstrating sustained performance within a wide range of experimental conditions or following long term storage. The highly specific nature of these reagents ensures accurate results with no matrix effects. Using site-specific chemistries, the Affimer binders are easy to orientate on a variety of surfaces to provide high capacity surfaces for sensitive protein capture and detection.

Binders can be developed to a wide range of targets from monomeric proteins to protein-protein complexes, including small molecules and peptides, that often prove intractable with classical antibodies. Moreover, Affimer reagents can be generated to form a capture and detection pair with an existing antibody or as an Affimer-Affimer pair to meet all assay requirements.

Broad dynamic assay range meeting regulatory compliance

ELISA plates coated with anti-trastuzumab Affimer® reagent (1µg/mL) using standard Fc detection in human serum samples showed a broader dynamic range and increased consistency in performance compared to standard antibody-based methods, whilst meeting FDA and EMA criteria for critical reagents in bioanalytical assays.

Simple Affimer® formatting yields flexibility in assay design

Affimer® reagents to human C-reactive protein were formatted as a dimer and used as a capture reagent to coat ELISA plates (2µg/mL) with monomeric Affimer detection reagents (1µg/mL in a sandwich ELISA format. Simple formatting with a cysteine residue allowed dimerization to increase assay sensitivity with increased surface capture. This assay offered a clinically relevant dynamic assay range with improved assay parameters compared to antibody-based assays, that were within the regulatory guidelines.

Achieving specificity and sensitivity with Affimer® immunoassay reagents

Introduction of dimeric Affimer® binders specific for C. difficile toxin B as capture reagents within a diagnostic sandwich ELISA increased the specificity of the assay, towards the more virulent and disease-causing form of this bacterial toxin, Toxin B, offering greater accuracy in assay results and improved diagnosis.