Lateral flow assays (LFAs) are extensively employed in diagnostic applications because of their rapid response time, straightforward operation, and low cost. However, traditional LFAs are frequently limited by problems with sensitivity, specificity, and quantitative precision, often delivering qualitative results only. To overcome these issues, advanced technologies are needed to improve LFA performance and broaden their diagnostic abilities.
This article shows how combining Molecular Devices’ ScanLater® Western Blot Detection System with Bright-Dtech™ nanoparticles, a proprietary technology from Poly-Dtech, effectively overcomes the intrinsic limitations of standard LFAs.
Initially developed for highly sensitive fluorescence detection in Western blotting, the ScanLater® system can be adapted to substantially improve lateral flow assessment, particularly when combined with Bright-Dtech™ technology.
Bright-Dtech™ nanoparticles provide a distinct advantage for LFAs due to their unique lanthanide-based probes, which offer remarkable brightness, photostability, and minimal background interference: these are essential for accurate and sensitive fluorescence-based detection. These nanoparticles ensure robust signal intensity, allowing precise quantification even at low analyte concentrations.
Advantages
- Bright-Dtech™ nanoparticles coupled with ScanLater® technology provide enhanced sensitivity compared with traditional LFAs.
- Accurate quantification of fluorescence intensity enables quantitative LFAs with precise determination of analyte concentrations.
- ScanLater® technology supports multiplexed analysis of multiple test strips, increasing both efficiency and throughput in high-volume workflows.
By combining the ScanLater® system’s quantitative fluorescence measurement abilities with the improved brightness and signal stability of Bright-Dtech™ probes, this technique broadens the analytical performance of LFAs beyond their conventional limitations.
To demonstrate the potential of this combined technology, this article presents an LFA specifically designed for the quantitative detection of human lactate dehydrogenase (h-LDH). h-LDH is an important enzyme involved in cellular metabolism, and its quantification serves as a valuable biomarker of tissue damage.
Increased h-LDH levels are a sign of disruption of cell membranes and are linked to illnesses including myocardial infarction, hemolysis, liver diseases, and cancer. In oncology, LDH also functions as a prognostic marker, with elevated levels frequently reflecting tumor progression and unfavorable outcomes.
This example demonstrates the improved sensitivity, precision, and reliability achieved using this novel method, highlighting its suitability for high-precision diagnostic development and testing across a variety of study and lab applications.
Figure 1. Depiction of the LFIA assay for detecting h-LDH using a non-competitive assay format with the Bright-Dtech™-614 Eu nanoparticles (EuNPs) as signal reporters. When excited at 340 nm, these EuNPs emit a red fluorescent light at 614 nm that is detected in a time-resolved mode. Image Credit: Molecular Devices UK Ltd
Advantages of Bright-Dtech nanoparticles + ScanLater technology
- Quantitative LFA: Integrating Bright-Dtech™ nanoparticles with ScanLater® technology allows accurate quantification of fluorescence intensity, enabling precise determination of analyte concentrations through the test-line-to-control-line (TL:CL) ratio.
- High sensitivity: Bright-Dtech™ nanoparticles improve detection sensitivity thanks to their extended fluorescence lifetime, supporting reliable quantification of low analyte concentrations.
- Low background interference: ScanLater® technology’s time-resolved fluorescence (TRF) capabilities provide substantially clearer and more accurate signal detection by reducing background interference.
- Enhanced sensitivity compared with standard LFAs: The combined application of Bright-Dtech™ nanoparticles and ScanLater® technology improves analytical performance and exceeds the sensitivity of conventional LFAs.
- Multiplexed evaluation: The ScanLater® system facilitates concurrent examination of several test strips, improving throughput and efficacy in high-volume diagnostic workflows.
- Flexible applications: This tool can be adapted to a broad range of analytes and sample types such as plasma, serum, saliva, cells, water, and others, making it ideally suited for a variety of applications across research, veterinary, or even agri-food environments.
Assay principle
The team has developed a dipstick LFA for the quantitative detection of an analyte using Bright-Dtech™ nanoparticles, with h-LDH used as a model.
In this assay, Bright-Dtech™ lanthanide nanoparticles replace traditional detection probes, such as gold nanoparticles and conventional europium chelates, to achieve high-sensitivity detection with improved fluorescence characteristics.
The test strip comprises a nitrocellulose membrane with two main detection lines: the test line (TL) and the control line (CL). The TL includes immobilized antibodies specific to the target antigen. When the analyte is present in the sample, these antibodies bind to it, capturing the nanoparticle-labeled complexes and producing a fluorescent signal proportional to the analyte concentration.
The CL functions as an internal control to confirm the validity of the assay by capturing surplus nanoparticles and generating a fluorescence signal regardless of analyte presence (Figure 1).
To accurately quantify the fluorescence signal at both the TL and CL following migration, the ScanLater® module was repurposed from its original application in high-sensitivity fluorescence detection for Western blot analysis. The system can simultaneously accommodate as many as 20 strips, improving assay throughput and consistency.
The SpectraMax® iD5 Multi-Mode Microplate Reader fitted with the ScanLater system and TRF detection is especially suitable for this application, as its fine-tuned fluorescence reading parameters correspond with Bright-Dtech™ nanoparticle characteristics.
Note: The iD5 reader has been updated to the newer SpectraMax iD5e Multi-Mode Reader, which has comparable features and performance, plus optional SpectraMax aer Gas Mixer and advanced shaking features.
Fluorescent images of the lateral flow strips are obtained in TRF mode using a 0.05 ms delay between excitation (350 nm) and emission (616 nm). This setup substantially minimizes background noise from autofluorescence and other short-lived emissions, especially those that originate from the nitrocellulose membrane, providing a clearer and more dependable quantitative readout.
The fluorescence intensity at the TL and CL is measured with ImageJ (National Institutes of Health, Bethesda, MD), employing the “gel analysis” function. This function makes it possible to choose each strip and transforms band intensities into peak profiles, with the region under the curve representing fluorescence intensity.
To normalize results and enable comparisons with other assays, the test-line-to-control-line ratio (TL:CL) was calculated. This acts as a normalized measure of analyte concentration.
Figure 2. Strips for the h-LDH LFA placed on the rack of the ScanLater™ system of the TRF SpectraMax® iD5 reader. Image Credit: Molecular Devices UK Ltd
Sensitivity and dynamic range
Calibration curves for h-LDH detection were generated by dispensing 5 μL of conjugated nanoparticles and 75 μL of serial dilution of h-LDH antigen (ranging from 0 to 40 ng/ mL in migration buffer, n = 4) into the wells of a 96-well plate. The test strips were subsequently immersed and allowed to migrate for 20 minutes before being air-dried.
The strips were positioned in the ScanLater® system rack (Figure 2) and underwent scanning. The images were acquired via the ScanLater® system and processed with ImageJ software for data extraction (Figure 3).
The obtained data was modeled with a five-parameter logistic (5PL) model in SoftMax Pro™ software (Molecular Devices), with an R2 of 1. The system attained a limit of detection (LoD) of 38 picograms/mL of h-LDH, indicating sensitivity similar to commercially available ELISA tests.
Following the optimization of the assay, a preclinical test was performed by spiking h-LDH into serum. The acquired results, with a recovery rate (R%) ranging from 88% to 121%, showcase the robustness of this test and its precise quantification abilities in a complex matrix such as serum.
Figure 3. (A) Images of the EuNPs-based LFIA strips after detecting serial dilutions of h-LDH in working buffer. (B) Calibration curve plot representing the normalized signal in the TL achieved when detecting serial dilutions of h-LDH standard in working buffer with EuNPs-based LFIA (n = 4). Inset showing the calibration curve plot between 0.001 and 10 ng mL-1 of h-LDH. Image Credit: Molecular Devices UK Ltd
| Sample |
Expected [h-LDH]
(ng mL-1) |
Detected [h-LDH]
(ng mL-1) |
Recovery (%) |
| 1 |
0 |
<LoD |
– |
| 2 |
0.6 |
0.8 ±0.1 |
121 |
| 3 |
1.2 |
1.2 ±0.1 |
100 |
| 4 |
2.5 |
2.4 ±0.2 |
94 |
| 5 |
5.0 |
5.3 ±0.16 |
105 |
| 6 |
6.2 |
7.1 ±0.2 |
113 |
| 7 |
10.0 |
11.7 ±0.7 |
117 |
| 8 |
12.5 |
14.3 ±1.0 |
114 |
| 9 |
20.0 |
23.2 ±2.6 |
116 |
| 10 |
25.0 |
21.9 ±4.8 |
88 |
| Mean |
|
|
108 ±11 |
Figure 4. Recovery results for analyzing h-LDH-spiked serum samples with an EuNP-based LFA. Image Credit: Molecular Devices UK Ltd
Conclusion
In conclusion, integrating Bright-Dtech™ nanoparticles with ScanLater® technology into LFAs indicates notable progress in quantitative assessment. The excellent background-reduction capabilities of ScanLater® technology, alongside the improved sensitivity of Bright-Dtech™ nanoparticles, enable detection of low analyte concentrations with elevated precision.
The dipstick assay for human lactate dehydrogenase (h-LDH) highlights this potential, achieving a limit of detection similar to that of conventional ELISA assessments while providing the benefits of a fast and intuitive format.
Acknowledgments
Produced from materials originally authored by Caroline Cardonnel, PhD, European Applications Supervisor at Molecular Devices; Juliette Lajoux, MSc, Research Engineer, Biology at Poly-Dtech; Susana Brun, PhD, Biology Manager at Poly-Dtech; and Mohamadou Sy, PhD, Chemistry Manager at Poly-Dtech.
References and further reading
- Lajoux, J., et al. (2025). Breaking the picomolar barrier in lateral flow assays using Bright-Dtech™ 614 – Europium nanoparticles for enhanced sensitivity. Microchemical Journal, 209, p.112864. DOI:10.1016/j.microc.2025.112864. https://www.sciencedirect.com/science/article/abs/pii/S0026265X25002188?via%3Dihub.
About Molecular Devices UK Ltd
Molecular Devices is one of the world’s leading providers of high-performance bioanalytical measurement systems, software and consumables for life science research, pharmaceutical and biotherapeutic development. Included within a broad product portfolio are platforms for high-throughput screening, genomic and cellular analysis, colony selection and microplate detection. These leading-edge products enable scientists to improve productivity and effectiveness, ultimately accelerating research and the discovery of new therapeutics. Molecular Devices is committed to the continual development of innovative solutions for life science applications. The company is headquartered in Silicon Valley, California, with offices around the globe. For more information, please visit www.moleculardevices.com.
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