New methods for hapten bioanalysis
Anti-hapten antibodies are difficult and slow to generate limiting cheap and efficient bioanalysis in this field. Our proprietary methods offer standard assay formats, such as ELISA and lateral flow tests, for simple hapten analysis with anti-hapten Optimers.
We previously discussed the issues with anti-hapten antibody production and examples of diagnostic assays that lack the required sensitivity and selectivity due to poor anti-hapten antibodies. Here we show how we have developed solutions to working with this target class. Using Optimer technology we have developed proprietary methods to enable increased selectivity and sensitivity in standard assay formats when working with haptens, to overcome the limitations of anti-hapten antibodies.
Optimer reagents to small molecule targets offer a simple and sensitive solution for the study of traditional haptens.
What is a hapten?
A hapten is an antigen that is too small to raise an immune response when injected into an animal on its own. To generate antibodies to a hapten, the hapten molecule must be conjugated to a carrier protein, such as BSA or KLH, to increase its size and allow it to be recognized by the immune system. Hapten-carrier complexes must exceed 3000 MW to be recognized by the immune system. Following recognition antibodies to the hapten-carrier complex, some of which will target the small-molecule hapten.
Small but still mighty
The pharmaceutical industry has long been dominated by small molecule drugs for the treatment of a wide variety of diseases and conditions. Diversity in their mechanism of action is provided by the small size and physicochemical profiles, offering use as enzyme inhibitors, allosteric modifiers and to interact with both intra- and extracellular targets. Despite the rise of antibody and large molecule therapeutics, small molecule drugs remain the major component of the pharmaceutical pipeline, with over three-quarters of the new molecular entities approved by the FDA between 2010 and 2017 being small molecules.
As small molecules are the main constituent of the pharmaceutical pipeline, sensitive analysis methods are essential in drug development and monitoring.
How do we analyze haptens with antibodies?
Currently, the main method used to analyze and monitor small-molecule drugs and their metabolites is mass spectrometry. This often requires coupling to liquid chromatography to remove contaminants that can affect results. Obvious limitations are the high cost, labor-intensive nature, the need for skilled staff to operate these systems and large footprints of this machinery.
The ability to use antibody-based assays, such as ELISA and biosensors, in place of MS analysis could:
- Reduce cost
- Increase speed
- Allow point-of-care diagnosis
- Improve patient outcomes
Immunoassays of hapten targets are typically based on competitive assays. This is because the small size of the hapten target is limited in the number of binding epitopes for anti-hapten antibodies to bind, making the development of matched pairs very challenging. Yet, competitive assays offer a negative readout and suffer sensitivity issues.
Optimer technology delivers an anti-hapten antibody alternative
Many researchers are investigating Optimers as anti-hapten antibody alternatives for targets such as bacterial quorum sensing molecules, antibiotics and chemotherapeutics. We have a success rate of over 80% in selecting Optimers to traditional haptens using our selection platform specifically to select Optimers to small molecules.
We understand the challenges of working with haptens and have developed a proprietary assay format for researchers to enable them to use ELISA, lateral flow and biosensor assays for the bioanalytical analysis of hapten targets with Optimer reagents.
Aptamer Group’s proprietary displacement method allows the development of sensitive, gain-of-signal assays using a single Optimer reagent to hapten targets.
ELISA or lateral flow assay
For traditional ELISA and lateral flow assays, Aptamer Group’s proprietary assay can be used to generate a single reagent assay that yields high sensitivity and selectivity to hapten targets with a positive readout.
For an ELISA the Optimer is bound to a compatible oligonucleotide on the bottom of the ELISA well. If the hapten target is present in the sample, the Optimer will preferentially bind the hapten over the oligonucleotide tag. The sample containing the hapten-Optimer complex can be collected and concentration determined based on the Optimer eg via a fluorescent tag or RT-PCR for increased sensitivity.
Researchers are also choosing to use these Optimers in bioanalytical assays to develop biosensors as point-of-care assays. Studies have shown sensitivities in the clinical concentration range for antibiotics and chemotherapeutics and assay criteria that meet the regulatory requirements, using anti-hapten Optimers on these platforms. Some researchers are working with Optimers and our proprietary assay format in this way to develop multiplex diagnostics.
Our Optimers to traditional hapten targets and proprietary single reagent assay format for small molecule monitoring are simplifying and accelerating the development of many research and diagnostic assays. The previous reliance on anti-hapten antibodies for the development and monitoring of small molecule drugs has hindered bioanalytical development in this field. The turn towards anti-hapten antibody alternatives should drive progression to increase speed and reduce costs, with the use of single-reagents assays leading to a much-needed step-change in the efficiency of the hapten analysis.