Non-Optical Sensor Technologies

Ruizhi Wang at Abselion talks to IPT about how non-optical sensor technologies are unlocking critical process data at the bench and bioreactor

IPT: Why is access to real-time protein quantification data so important for biologic and viral vector development?

Ruizhi Wang (RW): The development and production of biotherapeutics, such as antibody-based biologics, requires complex multi-step processes to reliably express and purify the therapeutic protein from cell culture at optimal yield and quality. Producing a therapeutic product from a biological system brings inherent variation. While bioprocessing strategies for antibody-based therapies have advanced over the years, multiple rounds of experimentation are required to define the optimal scalable production processes for commercialisation of each asset.1

More recently, advanced therapy medicinal products (ATMPs), such as cell and gene therapies (CGTs) that utilise viral vectors to deliver the therapeutic payload to the patient, have required the development of new production processes with their own challenges.2 As with biologics, cell culture techniques play a crucial role in viral vector production, impacting yield, purity and quality.3

Optimising bioprocess performance across multiple parameters – such as culture media components, environmental conditions, harvest timing or column loading for purification – is time-consuming, resource-intensive and costly. Frequent, accurate titre measurement of the relevant protein or viral vector capsid is needed at multiple points throughout cell line and process development to check the impact of process steps or parameter changes on yield and quality. Successful growth and expression in cell culture can be influenced by small adjustments in parameters requiring assessment across bioreactors and experimental conditions. Upstream and downstream bioprocessing teams need rapid access to titre data to make informed, on-the-spot decisions, whether that is to select the best clones or to optimise process conditions efficiently.

IPT: What are some of the technological and operational challenges that limit access to titre data for scientific decision-makers?

RW: While real-time titre data is extremely valuable to support informed process optimisation, in practice protein analysis systems can be complex to operate and are not always available in individual cell line development laboratories or process development environments.

High-performance liquid chromatography (HPLC) is a well-established technique in protein analysis. Target proteins are separated from impurities based on molecular interactions, polarity or size as they pass through the HPLC column. Protein identification and quantification is based on the specific peak area in a chromatogram, often analysed by spectroscopy. This powerful but complex instrumentation requires highly trained personnel to operate and calibrate. While a key tool for protein purification, HPLC was not devised for agile at-line titre monitoring of crude cell cultures and lysates, requiring frequent maintenance.4

Bio-layer interferometry (BLI) and surface plasmon resonance (SPR) are widely applied optical biosensor technologies for protein analysis. 5,6 First described for antibody-based detection of proteins in 1983, SPR uses microfluidics to flow samples across immobilised ligands on a biosensor surface.7 In BLI, biosensor tips are immersed into samples directly. Both methods measure changes in optical properties on the sensor surface as molecular complexes form.

Due to the significant investment, training and maintenance required, optical instruments are often housed in specialised analytical laboratories and managed by experienced operators. Titre analysis therefore occurs offline, away from the point of cell culture, with some delay as samples are batched, transferred and processed from multiple labs.

Another quantification method more applicable to at-line titre measurement is the enzyme-linked immunosorbent assay (ELISA). ELISA is simple, specific and has a relatively low equipment cost. However, it requires labourious assay procedures and lengthy incubations, which can lead to samples being frozen to manage throughput and delay data availability.8 As with any extensively manual process, it is also open to errors and variability.8

IPT: What is non-optical biosensor technology and how has it advanced?

RW: A non-optical biosensor converts a biological response into an electrical signal that is detected without reliance on light. A typical biosensor comprises of an analyte, bioreceptor, transducer, electronics and display. The non-optical transducer (device that transforms the biological interaction into a quantifiable signal) may be gravimetric, electronic, thermal or electrochemical.9

Technology for electrochemical sensing of biomolecules was initially developed in the late 1960s for measuring glucose, aimed at the application of diabetic care. 10 Electrochemical sensing of proteins followed shortly after with academic publications mentioning related methods in 1975. 11 Today, probably the most well-established use of electrochemical detection of proteins is the use in handheld point-of-care instrumentation for the detection of biomarkers.9

The main benefits of electrochemical detection are specificity, robustness and speed. Signal detection and processing are carried out using integrated circuits, resulting in compact and affordable equipment that is extremely robust and portable, in contrast to optical systems whose calibration can be easily disturbed. Measurements can be carried out in seconds and are highly specific as they are associated with changes to electrochemical properties uniquely linked to the analyte of interest.

IPT: Why is electrochemical sensor technology well placed to address at-line titre measurement?

RW: As bioprocessing technology has advanced, increases in yield have been driven by process efficiencies.1 Higher upstream cell densities require techniques that can rapidly measure titre within a dynamic range that does not require extensive manual dilution of samples or additional purification steps.

Electrochemical detection measures electron flow at the sensor surface associated with an enzymatic reaction, rather than changes within the whole sample. This means that the signal only occurs if the specific protein of interest is present, and few molecules are required for measurement to occur. Measurement is not impacted by other protein contaminants in the sample, or changes in buffer viscosity or temperature, for example, enabling direct analysis of crude and complex samples without extensive dilution.

Sensor technology combined with automation ensures consistent incubation times, removing the need for extended incubation to achieve binding equilibrium across all wells and reducing analysis times. The sensitivity and specificity of the electrochemical biosensor technology and its robustness, which is a consequence of the lack of optical components, means that sensors can be incorporated into compact instrumentation that can be positioned directly at the point of need – on a laboratory bench or beside a bioreactor.

Removing the reliance on optical measurement also simplifies the analysis, calibration and maintenance process, making non-optical sensor technology accessible to operate without extensive training.

IPT: How do you see the field of protein quantification developing in the next five years?

RW: Cost efficiency in R&D is a constant pressure, and utilising lab resources effectively is a key optimisation strategy. Providing agile access to protein quantification data as close to the biopharmaceutical development and production process as possible will allow upstream and downstream teams to optimise and monitor processes efficiently by adapting in real time. Electrochemical biosensor technologies can be deployed at-line to provide consistent rapid measurement across multiple unit operations as they are not affected by the sample matrix. Selecting and utilising protein quantification technologies for their ability to deliver the required results, depending on circumstance, will support the optimal use of resources. For instance, reserving complex equipment for detailed protein analysis under the auspices of experienced operators, while empowering individual laboratories with tools for robust, protein titre measurement at-line for rapid decisions.

References:

1. Mirasol F A (2023), ‘Look into Biologic Scale-Up Strategies’, BioPharm International, 36:(8), 15-18

2. Visit: ema.europa.eu/en/human-regulatory-overview/advanced-therapy-medicinal-products-overview

3. Visit: pharmafocuseurope.com/articles/optimizingpharmaceutical-viral-vector-production-techniquesand-challenges

4. Jadaun G (2017), ‘HPLC for Peptides and Proteins: Principles, Methods and Applications’, Pharm Methods, 8(1), 1-6

5. Jug A et al (2024), ‘Biolayer interferometry and its applications in drug discovery and development’, TrAC Trends in Analytical Chemistry, 176, 117741

6. Schasfoort R (2017), ‘Handbook of Surface Plasmon Resonance’, Royal Society of Chemistry

7. Liedberg B et al (1983), ‘Surface plasmon resonance for gas detection and biosensing’, Sensors and actuators, 4, 299-304

8. Song J G (2023), ‘Quantitative analysis of therapeutic proteins in biological fluids: recent advancement in analytical techniques’, Drug Delivery, 30(1)

9. Naresh V (2021), ‘A Review on Biosensors and Recent Development of Nanostructured Materials-Enabled Biosensors’, Sensors (Basel), 5;21(4), 1109

10. Updike S J (1967), ‘The enzyme electrode’, Nature, 214(5092), 986-988

11. Janata J (1975), ‘Immunoelectrode,’ Journal of the American Chemical Society, 97(10), 2914-2916

Ruizhi Wang is CEO of Abselion. Ruizhi received his PhD on semiconductor technologies from the University of Cambridge, UK, and MSc/BSc from ETH Zurich, Switzerland. He has co-authored 20 publications, with a total of more than 900 citations. Ruizhi has received awards, personally and for his involvement at Abselion. These include the Emerging Technologies Award of the Royal Society of Chemistry, the Displaying Future Award of Merck Group and the Grand Prize of the Cambridge University Entrepreneurs. He is an enterprise fellow of the Royal Academy of Engineering, UK, and a recipient of the Scholarship of the German Academic Merit Foundation.