Faster and Cheaper Techniques for the Characterisation of Immunoassay Conjugates

Fleet Bioprocessing Ltd

National Physical Laboratory
National Measurement Laboratory at LGC
Round 3

Fleet Bioprocessing Ltd.

are expert in the development of immunoassays, and fluorescent-labelled antibodies which enable exquisitely sensitive detection _via_ fluorometric quantification.

The performance of any given immunoassay is largely defined by the behaviour of these conjugates, which ideally display 100, whilst incorporating an optimally high level of label to facilitate immobilisation or detection.

A trade-off between these competing aims is required for optimal performance.

The chemistry required to _create_ such conjugates is well-established; with Fleet’s expertise, existing bioconjugation techniques are routinely exploited to yield conjugates with world-class immunoassay performance.

By contrast, the availability of rapid and inexpensive procedures for effective analysis and characterisation of these conjugates is extremely limited.

Simple cheap techniques can be used to determine basic conjugate characteristics such as antibody concentration or mean biotin or enzyme incorporation, but these provide no information on whether the antibody or antigen has retained immunological functionality; without this information it is impossible to know whether the conjugate will perform effectively in the immunoassay.

This means that the only available additional approach is to “”try it in the assay””, which (see _Need or Challenge_ section) has serious weaknesses.

The knowledge gap that this application aims to address, therefore, is to identify inexpensive and relatively fast techniques capable of confirming that antibody or antigen has retained adequate functionality during the conjugation process.

A technique capable of showing quickly and cheaply that the tertiary structure of the antigen or antibody remained intact after conjugation would be a massive step forward of great significance to our company and to the industry in general.

Initial discussions with LGC and NPL have indicated that both parties have access to analytical techniques which may directly or indirectly help to address this knowledge gap, including (from LGC) hydrogen/deuterium exchange mass spectrometry (HDX-MS) and ion mobility spectrometry mass spectrometry (IMS-MS) and (from NPL) circular dichroism spectroscopy (CD), Fourier transform infrared spectroscopy (FT-IR) and isothermal titration calorimetry (ITC).

This project comprises the preparation of suitable conjugate panels and determination of their immunoassay performance (Fleet), coupled with comparative evaluation of these conjugates using the techniques outlined above (LGC, NPL).

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