Lateral flow assays are fast to deploy but slow to develop. SPR accelerates the critical early steps — antibody selection, affinity ranking, and surface optimization.
Introduction
From virus detection to pregnancy testing, lateral flow assays (LFA) have revolutionized biological testing. It requires only a few drops of biological fluids like saliva, blood, or urine to achieve the detection of biomarkers within 30 minutes. Disposable and with a long shelf-life, LFA are the perfect point-of-care test. During the COVID-19 pandemic, LFA proved to be an effective tool against the spread of the disease. However, despite the rapid commercialization of LFA, numerous ones proved to be inaccurate. To meet the rising demand, LFA developers need better analytical tools to accelerate a development process that currently takes more than a year and costs hundreds of thousands of dollars.
Most LFA developers rely on Enzyme-Linked Immunosorbent Assay (ELISA) throughout feasibility and development processes. Slow, labour-intensive, and expensive, ELISA provides limited insights as this complex method translates poorly to the more simplistic LFA format. Surface plasmon resonance (SPR) is a type of biosensor format much closer to LFA. Label-free SPR provides direct binding results for any biomolecular interactions in real-time.
1. Immobilization of Antibodies
Proteins can be immobilized using biotin-streptavidin linkages or through NHS-EDC chemistry. Affinité also offers a surface coating called Afficoat™ that serves as a linker to enable carbodiimide conjugation to form covalent amide bonds between proteins and the surface linker's carboxyl groups. Immobilizing capture probes via high affinity or covalent linkages on SPR sensor chips rather than by passive adsorption on ELISA well plates is a better model for the development of LFA.
2. Tracking of Immobilization and Protein Binding in Real-Time
Another advantage of using SPR instead of ELISA to optimize LFA is the real-time tracking of surface modification and immobilization steps and subsequent binding interactions between proteins. In ELISA, the only evidence that proves that there is binding is the endpoint absorbance values of a true binding interaction. It would be difficult to troubleshoot which step in ELISA has gone wrong or requires optimization.
3. Use of Complex Samples
LFAs are designed to test complex samples such as urine, saliva, sweat, serum, plasma, and whole blood. Conventional SPR is an optical technique that does not rely on a direct optical path through the sample. The change in refractive index on the surface of the gold film induces changes in the SPR signal. Therefore, conventional SPR can easily handle complex samples for LFA development.
4. Screening of Antibody-Antigen Pairs
SPR allows simultaneous and real-time screening of antibody-antigen pairs. Each channel of the SPR sensor chip can be dedicated for a different antibody-antigen pair. Screening is vital to test any steric hindrance issues and determine the dissociation constant (KD) of the biomolecular interaction, which affects the sensitivity of the assay.
5. Time Window of Binding Kinetics
SPR is a better analytical tool to evaluate the kinetics of the binding interaction of an antibody-antigen complex. In LFA, the typical reaction time window is between 5–30 min. Therefore, the association rate (kon) is a very critical parameter to determine. SPR has an experimental time frame of 30 min, which is similar to that of LFA.
Conclusions
SPR provides a more suitable testing platform than ELISA to develop LFA rapidly. Capture molecules can be immobilized via different strategies onto an SPR sensor surface rather than being passively adsorbed in ELISA. SPR allows the antibody immobilization steps and protein interactions to be monitored closely in real-time. Complex biological samples can be tested on SPR sensor chips without much sample preparation. SPR is a better tool to gather kinetics information on antibody-antigen pairs due to the availability of compatible pumps to provide controlled flow conditions.