Fig. 1 Illustration of fluorescent nanodiamond, which could enable early stage diagnostics of diseases like HIV and SARS-CoV-2.An imperfection in the highly regular crystal structure of diamond could hold the key for ultrasensitive biosensing. Researchers at University College London (UCL) have shown that these nitrogen-vacancy (NV) centers can be used to boost the sensitivity of diagnostic tests for the early detection of diseases like HIV [Miller et al., Nature (2020), https://doi.org/10.1038/s41586-020-2917-1].
Early diagnosis is vital to manage the spread of infectious diseases and start treatment as soon as possible. In HIV, for example, viral RNA can be detected 7 days before antigens and 14 days before antibodies, so nucleic acid testing offers a promising strategy for earlier disease detection. Paper-based lateral flow tests, like those used in over-the-counter pregnancy or HIV tests, rely on a color change or fluorescent signal to indicate a positive result when a strip of sample-soaked paper detects DNA or virus proteins. These diagnostic tests rely on light emission from gold nanoparticles but background fluorescence levels from nitrocellulose in the paper substrate limit their sensitivity.
“Paper-based lateral flow tests with gold nanoparticles do not require laboratory analysis, making them particularly useful in low resource settings and where access to healthcare is limited. They are low cost, portable, and user friendly,” explains first author of the study, Ben S. Miller. “However, these tests currently lack the sensitivity to detect very low levels of biomarkers.”
NV centers in nanodiamond give rise to attractive optical properties such as high brightness, stability, lack of blinking and photobleaching, which coupled with diamond's low toxicity, biocompatibility, low cost, and ease of manufacture, are ideal for in vitro sensing (Fig. 1). Miller, along with Rachel A. McKendry and colleagues in the i-sense research group, incorporated biomolecule-functionalized nanodiamonds into paper-based lateral flow tests using a model biotin-avidin interaction.
“[Our] approach takes advantage of the quantum properties of nanodiamond to improve the signal-to-background ratio, resulting in a sensitivity improvement of 100,000 times compared to gold nanoparticles, benchmarking with a biotin-avidin model,” he says. “By… selectively modulating their (already bright) emission of light, we have been able to separate their signal from the unwanted background fluorescence of the test strip, dramatically improving sensitivity.”
The results with 600 nm fluorescent nanodiamonds are impressive: the approach has a detection limit of 0.5 particles per microliter and offers a 100,000-fold improvement in fluorescence over conventional lateral flow strips. Applying the same technique to a sandwich assay for DNA detection gives a sensitivity improvement of 7500-fold over gold nanoparticles. By adding a short 10-minute amplification step into the process, in which the number of copies of HIV RNA are multiplied, the approach can achieve single molecule detection.
“This greater sensitivity means that lower biomarker concentrations can be detected, meaning that a test using fluorescent nanodiamonds could detect a disease at an earlier stage, which could reduce the risk of onwards transmission and improve prognosis with earlier treatment,” says Miller.
The simplicity and ease of the approach are attractive for point-of-care diagnostic tests that can be carried out in clinical settings or even at home, without the need for lab processing. The team are now working on adapting their flexible approach to test for SARS-CoV-2.
“Our proof-of-concept study shows how quantum technologies can be used to detect ultralow levels of virus in a patient sample, enabling much earlier diagnosis,” says McKendry. “The use of fluorescent nanodiamonds is applicable to numerous diagnostic test formats and diseases, where the improved sensitivity has the potential to allow earlier diagnosis, improving prognosis and reducing disease spread,” adds Miller.
The main disadvantage of the current approach, compared with gold nanoparticles, is that a fluorescence reader is required. But the team are now working on a solution so that a hand-held device such as a smartphone could read out the results.
“We believe that this transformative new technology will benefit patients and protect populations from infectious diseases,” says McKendry.
As the current COVID-2019 pandemic has demonstrated, biosensing with high selectivity and high sensitivity is of critical importance for early disease detection, points out Yury Gogotsi of Drexel University. Rapid and inexpensive point-of-care tests could transform healthcare.
“The authors demonstrate here that fluorescent nanodiamonds can be used as an ultrasensitive label for in vitro diagnostics and demonstrate extreme sensitivity, including single-copy detection of HIV RNA,” he says. “Strictly speaking, these particles are not quite ‘nano’, as they are larger than 100 nm, but fluorescence of diamond at larger sizes become brighter, as there are more fluorescent centers per particle. This is clearly a major advantage.”
While Gogotsi cautions that there is still long way to go before clinical implementation, the proposed approach may allow sensitive disease diagnostics with the use of a smartphone for detection, which would be of great advantage, especially in low-resource settings outside of hospitals.
This article originally appeared in Nano Today 36 (2021) 101067.