The Un-LFA: 3 Reasons Why CaDI Stands Apart from Lateral Flow Assays and Stays Familiar

Lateral flow assays (LFAs) are widely used across clinical and non-clinical industries due to their low cost, portability, rapid turnaround, and ease of use. From over-the-counter pregnancy tests to infectious disease screening, LFAs have become a pillar of rapid diagnostics. However, their simplicity also introduces performance limitations, especially when high sensitivity or quantification is required. 

At Burst Diagnostics, we’ve developed CaDI to address these limitations. On a scientific level, CaDI is unlike a traditional LFA, but it retains many of the features that make LFAs appealing. The following article breaks down the key technical differences between LFAs and CaDI, exploring the modifications that elevate CaDI’s technical performance and similarities that preserve the familiar user experience, rapid time to result, and low-cost.  

1. Flow Format 

Traditional LFA Limitation: Linear Flow 

In a traditional LFAs, a sample is added to the test cartridge and flows linearly through the test strip. If the target is present in the sample, a signal will appear at both the test line and the control line. If the target is absent, a signal will appear only at the control line. Regardless, everything happens in a single, continuous flow step. This approach is simple but doesn’t allow for the capability to control timing or enable sequential steps for washing or signal amplification. Incorporating these additional washing and amplification steps would require manual user intervention, which adds workflow complexity, increases variability, and disrupts the simplicity that makes LFAs appealing. As a result, the sensitivity is limited under these constraints. 

The CaDI Advantage: Non-Linear Flow 

CaDI takes a different approach by using a microfluidic device with a non-linear flow path that enables automated control over the movement of sample and the timing/release of reagents. Two reagent pads, embedded within the device, are rehydrated as the buffer flows over them and sequentially delivered to a test strip based on the geometry, length, and width of the channels in the device. This non-linear flow path and timed release enables CaDI to execute automated washing and signal amplification steps, critical for enhanced sensitivity and quantification, without additional user intervention. The non-linear flow path may seem more complex internally but adds no complexity for the end user. The result is a platform capable of supporting more sensitive and quantitative tests, while preserving the speed and simplicity necessary for point-of-need settings. 

2. Detection Modality 

 Traditional LFA Limitation: Non-Amplified Fluorescent Detection 

Traditional LFAs often use colorimetric or fluorescent labels to generate signal. In fluorescent LFAs, signal is produced when an external light source excites fluorescent tags on detection antibodies. In colorimetric LFAs, signal is generated when the target binds to gold nanoparticles at the test and control lines, producing two visible colored lines. In both types, each binding event corresponds to a fixed amount of signal. While fluorescence increases sensitivity over colorimetric methods, it does not offer true signal amplification. Signal output remains directly proportional to the number of binding events, and performance is limited by the number of detectable labels at the test and control lines.  

The CaDI Advantage: Enzyme-Amplified Chemiluminescent Detection 

CaDI replaces fluorescent and colorimetric detection with chemiluminescent detection. Within the channels of the CaDI device, separate reagent pads containing enzyme-labeled detection antibodies and a substrate are sequentially delivered to the detection zone. When this enzyme encounters its substrate, it catalyzes a chemiluminescent reaction that produces light. Because each enzyme can catalyze many reactions over time, unlike LFAs with fixed signal, a single binding event can generate a significantly stronger signal. This repeated reaction effectively amplifies the signal, enabling lower limits of detection and a broader detection range. 

This approach also improves reproducibility and allows for precise quantification at low concentrations, which can be useful in applications such as food safety, clinical diagnostics, environmental monitoring, and beyond.   

3. Signal Output 

Traditional LFA Limitation: Excitation-Induced Background Signal 

Finally, one of the challenges with fluorescent LFAs is background interference. To generate signal, these tests require external excitation light, typically from a reader, which can also cause autofluorescence from the test strip material. Some materials naturally absorb and emit light, even without fluorescent labels, leading to unwanted background signal. This unintended light emission can create background noise in fluorescent LFAs, making it difficult to detect weak signals or quantify low concentrations accurately. 

The CaDI Advantage: Low Background Signal, Reliable Quantification 

Conversely, CaDI uses chemiluminescence, which produces light when the enzyme-substrate reaction occurs. There is no external excitation source required, so the background signal is inherently lower. This cleaner signal improves the signal-to-noise ratio, particularly at low concentrations, and allows for more reliable quantification. The low background, signal amplification, and enhanced sensitivity make CaDI well suited for applications where a quantitative immunoassay is required for decision-making. 

Shared Strengths 

Despite its technical differences, CaDI retains many of the advantages that make LFAs appealing: 

• Results in under 10 minutes 

• Compact test cartridge and reader 

• Minimal user training required 

• One-step sample addition 

• Low-cost device (<$5 COGS) 

These shared features make CaDI a portable diagnostic solution for the same decentralized settings as LFAs, including clinics, industrial sites, field environments, or at-home use. The key difference is that CaDI leverages chemiluminescent immunoassay technology to expand what’s possible in these settings, deliver more powerful diagnostics, and overcome the inherent sensitivity constraints of LFAs. 

Conclusion 

Lateral flow assays have played an essential role in improving access to diagnostics. But their limitations, particularly around linear flow, lack of signal amplification, background interference, and quantification, have made them less suitable for some emerging use cases. CaDI addresses these challenges by introducing controlled non-linear flow, signal amplification, and low background chemiluminescence. These changes improve analytical performance without compromising speed or usability. 

If you're looking to develop a rapid, low-cost, quantitative immunoassay to solve an industry challenge, our custom assay development services can help. Explore our custom assay solution page to learn more about how our team leads develops chemiluminescent immunoassays, powered by CaDI, across food safety, veterinary diagnostics, environmental testing, pharma, and beyond. 

Our custom assay development team will work with you from assay concept through full-scale manufacturing to ensure your test meets both technical requirements and real-world use case needs. Explore our case studies to learn how CaDI is already helping partners overcome sensitivity limitations of lateral flow assays.  

Contact us at info@burst-dx.com to explore timelines, project requirements, cost, feasibility, and how our custom assay solutions can help you overcome technical challenges and accelerate development.