Aptamer Biosensor vs Antibodies - Insights on Biosensing Applications

Introduction

Aptamers and antibodies are two distinct classes of binding molecules widely used in biosensing but differ fundamentally in structure and production. They represent two distinct tools in the molecular toolbox. It is important to understand which tool is right for your project.

This article provides an objective comparison of aptamers vs antibodies across three main biosensing applications: lateral flow, electrochemical sensors and surface plasmon resonance. We compare the performance of aptamers and antibodies, as well as their optimisation strategies, development timelines, commercial availability and regulatory considerations to equip you with the information needed to make an educated choice for your next project. 

Background on Aptamer Biosensors and Antibodies

Aptamers, also referred to as “chemical antibodies”, are short single-stranded DNA or RNA oligonucleotides selected for specific target binding which are chemically synthesised. In comparison, antibodies are large protein immunoglobulins generated by the immune system of animal models in vivo, or by recombinant expression in vitro. These differences give rise to unique performance characteristics.

Size of Binding Molecule

Aptamers are 5-10 times smaller (1-3 nm for aptamers compared to 10-15 nm for antibodies), and 10 times lighter (~15 kDa for aptamers compared to ~150 kDa for antibodies), allowing them to bind and fold very specifically. This is useful for diagnostics, allowing an improved sensitivity due to high packing density on sensor surfaces. 

Stability

Aptamers are exceptionally stable: they can tolerate a wide range of pH and temperature, be dried and rehydrated, or even heat-denatured and refolded to restore function, opening up use cases in these complex matrices. This is in-part thanks to their robust phosphodiester backbone which gives them a long shelf-life under ambient conditions, allowing aptamers to be shipped and stored without a cold supply chain, unlike protein reagents that risk denaturation [1]. 

Antibodies can sometimes have a higher thermal denaturation threshold but because they do not have the ability to renature, their practical stability is more limited than that of aptamers.

Molecule Type	Denatures Above	Unstable pH Range	Renature capability?
IgG Antibody	60–75 °C (140–167 °F)	<5.0 or >9.0	No
DNA Aptamer	40–80 °C / 104–176 °F	<5.0 or >9.0	Yes
RNA Aptamer	40–70 °C / 104–158 °F	<6.0 or >8.5	Yes

A breakdown of aptamer advantages:

Feature	Aptamers	Reference
Thermal recovery	Can renature upon cooling; antibodies typically cannot.	[2], [3]
Freeze-thaw tolerance	Highly resistant to freeze-thaw cycles.	[4], [5]
Chemical stability	Chemically synthesized and modifiable (e.g. PEGylation, LNA bases) for enhanced stability.	[6], [7]
Storage	Can be stored lyophilised at room temp for months to years.	[8], [9]
Batch consistency	No batch-to-batch variability (unlike biologically produced antibodies).	[10]

A breakdown of antibody disadvantages:

Feature	Antibodies	Reference
Cold chain required	Must be kept at 2–8 °C, no freezing.	[11], [12]
Aggregation risk	Denatured or shaken antibodies aggregate irreversibly.	[13], [14]
pH sensitivity	Sensitive to both acidic and basic pH shifts.	[15], [16]
Limited chemical tunability	Modifying structure can be challenging and unpredictable.	[17]

Binding Affinity & Target Suitability 

Antibodies have the strong advantage of a long-proven track record in biosensing compared to its new-comer, aptamers. Antibodies often exhibit nanomolar affinities and extremely high specificity [16]. Antibody discovery requires at least some target immunogenicity, generating antibodies against certain analytes can be infeasible, for example: toxins, small molecules, or other non-immunogenic compounds which may not elicit a useful immune response [1].

A vast repertoire of validated antibodies are biologically produced using phage display or (more traditionally) derived from animals. Antibodies are thus subject to high batch-variability and cost [18]. Even recombinant antibodies can show a lot-to-lot difference or require costly cell culture production.

On the other hand we have Aptamers with binding affinities range from 1–1000 nM9. Thanks to SELEX (Systematic Evolution of Ligands by EXponential enrichment), aptamers face none of the target limitations experienced by antibodies. SELEX is the entirely in vitro process of discovering aptamers; an evolutionary screen inspired by Darwinian evolution. It starts with a library of trillions of random sequences, keeps the few that stick to the target, amplifies them and repeats until only the strongest binders remain. As a result, given a suitable SELEX strategy, aptamers can be evolved to bind virtually to any molecule, from ions and small organics to proteins and whole cells. Aptamers can be synthesised with precise, single-site modifications to facilitate oriented immobilisation or reporter attachment [1]. Because aptamers are produced by chemical synthesis rather than biological fermentation, batch-to-batch variability is close to non-existent and are about 5-6 cheaper than antibodies to manufacture at scale [19].

Figure I - Schematic workflow of SELEX for aptamer selection

The following sections of this article evaluate how the properties of aptamers and antibodies translate into actual performance differences on three major biosensing platforms: lateral flow assays, ELISA, electrochemical sensors, and surface plasmon resonance; highlighting the scenarios where aptamers or antibodies have the edge.

Lateral Flow Assays (LFAs)

Lateral flow assays (LFAs) are widely used for rapid, low cost biosensing and diagnostics, from pregnancy tests to COVID-19 screening. Traditionally, antibodies serve as the capture and detection elements, but aptamers are emerging as a compelling alternative. 

Figure II – Schematic of the Lateral Flow Assay with Aptamers Conjugates

Stability

A key advantage of aptamers is stability: aptamers remain functional after heat exposure and drying, making them ideal for use in settings without refrigeration. This contrasts with antibodies, which can degrade in high temperatures or extreme pH environments [1].

Low Cost

Cost is another differentiator. Biologically produced antibodies are often tenfold more expensive than synthetic aptamers. Substituting aptamers can reduce per-test reagent costs. Once an aptamer sequence is known, it can be synthesised indefinitely with consistent quality, whereas antibody production requires maintaining hybridoma cell lines or animals, contributing to higher expense and variability [1].

Overcoming Past Challenges of Aptamer LFA Manufacturing

Despite these benefits, integrating aptamers into LFAs has posed some engineering challenges. Nitrocellulose membranes naturally bind proteins like antibodies but not nucleic acids. Early aptamer LFAs used streptavidin linkers, but this added complexity. Newer chemical strategies now allow direct aptamer immobilisation, paving the way for aptamer LFAs, also called ALFAs. Thanks to the discovery process of aptamers, ALFAs can be engineered to bind almost any target, including small molecules like toxins or antibiotics, an area where antibodies often fall short. For example, one ALFA for the marine toxin tetrodotoxin achieved a detection limit of ~0.3 ng/mL, rivalling top-performing antibody tests20. Similarly, an ALFA has also been developed to detect the antibiotic ampicillin in milk, achieving a high-selectivity that antibody-based methods struggle to attain [20].

While antibodies benefit from decades of availability and validated pairs (matched set of antibodies with one for target detection and the other for capture), the aptamer toolkit is growing. As the technology matures, aptamers are likely to take the lead in applications demanding ruggedness, cost-efficiency, or detection of otherwise intractable targets.

Electrochemical Aptamer Biosensors

Electrochemical biosensors detect targets by translating a biorecognition event into an electrical signal (current, voltage, or impedance change). Common implementations include amperometric sensors (measuring current from redox reactions), potentiometric sensors (measuring voltage shifts), and impedimetric sensors (measuring changes in electrical impedance at a surface).

When comparing aptamers versus antibodies as the sensing element in electrochemical biosensing platforms, one finds aptamers especially well-suited to this modality. In a traditional electrochemical immunosensor, an antibody will be coated on an electrode to capture the analyte, and detection often requires a secondary antibody conjugated to an enzyme (such as HRP) or a redox-active label. The enzyme then produces an electro-active product, or the label generates a current under the right conditions; essentially an ELISA-like process adapted to electrodes. This typically involves multiple steps and reagents [21].

Aptamer-based electrochemical sensors, by contrast, can often operate in a reagentless format: the aptamer itself is modified with a redox reporter and is immobilised on the electrode allowing the signal to be generated directly upon target binding (see Figure V). This is also called: the “E-aptamer”, or electrochemical aptamer-based sensor (E-AB sensor). The aptamer is end-labelled with a small redox molecule (e.g. ferrocene or methylene blue) and attached to the electrode surface. In the absence of the target, the aptamer is in a flexible or partially folded state that keeps the redox tag far from the electrode, hence limiting electron transfer. When the target binds, the aptamer undergoes a conformational change that brings the redox tag closer to the electrode surface, resulting in a measurable increase in current [22+23]. Thus, binding directly modulates the electrical signal. This mechanism is rapid, real-time, and single-step, no wash steps or secondary reagents are needed to produce a signal, since the aptamer itself is the transducer. This sensor architecture leverages the ability to design the highly flexible phosphodiester backbone of an aptamer to undergo a conformational folding event, something not possible with antibodies.

Figure V - Schematic of Target-Induced Folding in Electrochemical Aptamer Biosensors (E-ABs)

The flexible structure of an aptamer’s backbone can be leveraged to produce reversible and continuous sensors. Something not possible with antibodies. 

Small Size, Stability, Synthesis

In addition, the properties of aptamers, such as their size and stability allow for the engineering of practical electrochemical sensors. Their small size (~15 kDa) allows for dense surface packing and positions target binding events close to the sensor surface, critical for nano-scale platforms like field-effect transistors (FETs) and nanowire sensors, where signal strength depends on proximity. Antibodies, by contrast, are much larger (~150 kDa) and can place the target outside the effective sensing range, reducing signal quality [24]. The stability of aptamers also means that an electrochemical aptasensor can be used in harsher conditions or cleaned between uses. Aptamers can also tolerate organic solvent or non-aqueous conditions better than antibodies in some cases, which is useful for certain electrochemical detections in mixed solvents. Aptamers’ “dry” synthesis means they can be integrated into device fabrication more easily; one can print aptamer-containing inks or use solid-phase synthesised aptamers for on-chip sensor assembly without needing cold shipping of antibodies. All these factors lend aptamers a flexibility in sensor design [25]. Of course, any sensor’s performance also depends on the transducer and design – antibodies can also reach low detection limits if coupled to a good electrochemical transducer with amplification. But the aptamer’s ability to be intimately integrated into the electrode interface (often as part of a dynamic signalling architecture) gives it a unique edge in creating simple, reagent-free biosensors [22].

Surface Plasmon Resonance (SPR) Biosensors

SPR is a label-free optical sensing technique to measure bio-molecular interactions in real time, often to determine binding stoichiometries, affinities and kinetics. In an SPR sensor, one interaction partner is immobilised, and the other partner is flowed over; binding is detected by a change in the refractive index at the surface. Both antibodies and aptamers are commonly used as capture molecules in SPR assays, and each brings distinct advantages to the SPR format.

Aptamers are particularly well-suited for SPR biosensors thanks to their small size and ease of site-specific modification. They can be synthesised with a thiol group to anchor precisely onto gold surfaces, creating a densely packed and uniformly oriented layer with all binding sites accessible, ideal for efficient target capture. In contrast, antibodies are roughly ten times larger and have multiple attachment sites, often leading to random, uneven orientations when immobilised. This can block their binding regions and reduce sensor performance. While certain techniques can help orient antibodies, they add complexity and aren’t universally compatible. Aptamers naturally avoid these issues and therefore increase signal strength in SPR by capturing more target molecules from dilute samples [23].

Figure VI - Schematic of Aptamer-Functionalised Surface Plasmon Resonance (SPR) Sensor for Real-Time Analyte Detection

Stability 

In terms of regeneration and reusability, aptamers are often superior in SPR biosensors thanks to their ability to withstand hard regeneration conditions. Unlike antibodies which lose function when exposed to extreme pH, heat or solvents; aptamers, being nucleic acids, can denature and refold without losing activity. This makes them ideal for repeated use in SPR assays, where sensor surfaces must be regenerated between measurements. Studies have shown that aptamer-coated SPR chips retain full functionality after multiple regeneration cycles using acid washes or heat, whereas antibody surfaces degrade quickly and require gentler, less effective treatments. As a result, aptamer-based sensors are more cost-effective and sustainable, offering reliable performance across many binding cycles.

Small Size

However, one consideration for SPR detection is the mass of the analyte and the recognition molecule. Aptamers being smaller than antibodies means the total mass on the surface from an aptamer and target complex may be less than an antibody and target complex, potentially yielding a smaller SPR angle shift for the same number of binding events. However, this is often compensated by the higher aptamer surface density capturing more target analytes. If the target is a large protein or cell, the analyte’s mass dominates the signal and the difference between using an aptamer or an antibody is negligible in terms of SPR response. However, if the target is a small molecule, both binders will produce only a small direct SPR signal upon binding such a low-mass analyte. For these types of targets, creative signal amplification strategies are required for either binder. 

Conformational Folding

Aptamers are especially useful here, as they can be engineered for conformational changes upon binding, which can amplify SPR signals or generate distinct optical signatures. They can also be paired with plasmonic nanomaterials for Localised Surface Platform Resonance (LSPR) applications. For example, in one study an aptamer for a small antibiotic was used on an SPR sensor, and the aptamer’s folding upon binding effectively increased the local refractive index change, enabling detection of kanamycin down to sub-ng/mL levels. These structural-switching capabilities, unique to aptamers, offer powerful signal enhancement tools not easily replicated with antibodies [23].

Selectivity & Cross Reactivity

SPR is also often used to test the selectivity and cross-reactivity of a sensor to various flowing analytes. Aptamers offer exceptional selectivity in SPR assays thanks to the SELEX process, which can include counter-selection against similar molecules to minimise cross-reactivity. As a result, antibodies are more likely to bind to unintended targets compared to aptamers. In a study, aptamers in a dual-analyte SPR assay accurately distinguished E. coli and S. aureus without cross-binding, something that would have been challenging to perform with antibodies [23]. This is in part due as well to the small size of aptamers, which can be tuned to bind to sites of the target of choice. This counter-target selection of aptamers during SELEX makes them ideal for multiplexed SPR, where different sensor spots can be functionalised with different aptamers without the fear of cross-interference. Moreover, as nucleic acids, aptamers enable additional assay designs not possible with antibodies. For instance, they can be hybridised with complementary DNA to remove or organise aptamers on the surface, or to actuate binding. An interesting concept is an SPR assay where an aptamer is hybridised to a short complementary strand to keep it in an inactive conformation; when target is present, the aptamer releases the strand and binds the target, providing an extra level of specificity and an internal control via the complementary strand [25]. Such nucleic acid tricks have no parallel in antibody-based SPR.

Conclusion

As the biotechnology landscape continues to evolve rapidly, it’s increasingly important to consider alternative tools, such as aptamers, rather than assuming antibodies are always the superior choice for every biosensing application. Aptamers now rival antibodies across biosensing applications, offering superior stability, reusability, and unique signal mechanisms thanks to their nucleic acid nature. They perform particularly well in rugged formats like electrochemical and SPR sensors and excel at detecting small molecules. While antibodies remain popular for established protein assays thanks to their high affinity and widespread availability, aptamers are increasingly preferred where robustness, cost, or target flexibility are priorities. The future of biosensing lies not in choosing one over the other, but in strategically deploying each where it suits best, sometimes in combination, to optimise biosensing performance across varied platforms and applications.

At PentaBind, we are the pioneers of AI-powered aptamer design and functional aptamers. Our AI architecture used in synergy with our proprietary wet-lab workflow allows us to overcome the limitations of traditional SELEX to design multifunctional aptamers with stronger binding affinity in a single molecule, and 10-20x faster than traditional SELEX campaigns. If you are looking to design bespoke functional aptamers, find functional aptamers in your SELEX data or simply accelerate your projects' R&D, feel free to schedule a call with us.

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