Antibody-Enzyme Conjugates & Other Enzyme-Labelling Applications

With the use of suitable substrates, enzymes such as horseradish peroxidase (HRP) and alkaline phosphatase (AP) can be exploited in colorimetric, fluorescent and luminescent signal generation approaches, capable of very high sensitivity. These approaches are employed in a very wide range of applications, including commercial immunoassays, cytochemical and histochemical procedures, Western blots, etc.

Antibody-HRP and antibody-AP conjugates form some of Fleet Bioprocessing Ltd.’s most popular product categories, and our expertise in optimising these conjugation procedures is unparalleled. We use “clean” heterobifunctional coupling techniques which give a much more homogeneous product than older chemistries. We typically use high-resolution size exclusion chromatography (HR-SEC) to purify our enzyme conjugates, the exact choice of purification conditions often being a key factor in obtaining optimal conjugate performance. Fleet’s knowhow will always make sure that you get the best out of your enzyme-labelled antibodies!

Of course we regularly couple HRP and other enzymes to targets other than antibodies, and have made countless conjugates between enzymes and small molecules, protein antigens, streptavidin, oligonucleotides, etc.

Example Projects

From R&D to Production

Fleet have worked with a world renowned diagnostics company based in the Midlands, to develop their Ab-HRP conjugate taking it through process optimisation, robustness and validation all the way to manufacture within Fleet's labs.

Development of mAb-HRP conjugates for immunoassay

Fleet carried out a program of work to develop monoclonal-HRP conjugates giving excellent performance in the customer's assay (high signal, minimum non-specific binding). The conjugation has been validated for multiple batches of key raw materials and for a range of production scales.

Antibody-alkaline phosphatase conjugate for TB diagnostic

Fleet developed a panel of monoclonal antibody-alkaline phosphatase conjugates for use in a novel TB test. Full robustness studies were executed by Fleet to confirm that all key variables were under control.

Redevelopment of an established Ab-AP conjugate to improve process robustness

Working with a company based in Europe, Fleet helped to redevelop some reaction parameters and the linker strategy for their Ab-AP reagent, which has increased reliability of the AP functionalisation and robustness of the conjugation.

Poly-HRP

Building on their expertise, Fleet have developed strategies for the polymerisation of HRP, which has already shown promise compared against the same traditional Ab-HRP conjugates, for ever greater ELISA sensitivity.