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Plasmonic ELISA Offers High-Sensitivity Protein Assay Readable by the Naked Eye


Researchers at Imperial College London have developed a high-sensitivity immunoassay that can be easily read with the naked eye.

Detailed in a paper published this week in the online edition of Nature Nanotechnology, the technique could provide a quick and inexpensive way of testing for various disease biomarkers and could prove particularly useful in resource-constrained settings where more expensive and equipment-intensive methods are not available or feasible, Molly Stevens, professor of biomedical materials and regenerative medicine at Imperial College and author on the paper, told ProteoMonitor.

The technique, which the researchers termed plasmonic ELISA, combines a standard ELISA with gold nanoparticles to generate colored solutions in the presence of a target analyte.

In a traditional ELISA the assay's enzyme label is converted into a colored molecule upon capture of the target protein. The intensity of color can then be read using a plate reader, allowing researchers to quantify the amount of target present.

The plasmonic ELISA method, by contrast, links the assay's enzyme label to the growth of gold nanoparticles so that when the target is absent the reduction of gold ions with hydrogen peroxide occurs rapidly, forming gold nanoparticles that take on a red color. In the presence of the target, on the other hand, the enzyme label interacts with hydrogen peroxide, resulting in a form of nanoparticle growth that turns the sample blue. Unlike the color change generated by conventional ELISAs, these red and blue colors can be easily distinguished by the naked eye.

This, Stevens noted, makes the technique ideal for use in areas without access to equipment like ELISA plate readers. In particular, she said, the researchers "think the most immediate application is for infectious disease detection in resource-constrained countries due to the simple color readout that does not require specialized equipment."

In addition to the technique's simplicity, it also offers high sensitivity, making it potentially competitive with high-sensitivity immunoassay technology offered by companies like Quanterix and Singulex as well as nucleic acid-based methods. In the Nature Nanotechnology paper the researchers used the method to detect prostate specific antigen and HIV-1 capsid antigen p24 in whole serum at concentrations as low as 1x10-18 grams per ml. At these levels, they wrote, they were able to detect p24 at levels lower than those detectable by nucleic acid-based techniques – the current gold standard for HIV testing.

Given these results, the method "is very promising for the diagnosis of HIV infection in patients with a very low viral load," Stevens said.

A significant drawback of the plasmonic ELISA method is that while it allows researchers to detect the presence of a target with the naked eye, it doesn't allow for accurate quantitation of the target, making it unsuitable for assays where it is important to know the precise concentration of the analyte under investigation – such as viral load monitoring in HIV patients.

Nonetheless, Stevens said, even with this limitation, the method still has a number of potential uses. Infectious disease testing is perhaps foremost among them, but, she suggested, it could also be used as a simple, inexpensive screening technique for other diseases.

As demonstrated by the study's detection of PSA, "the extreme sensitivity shown in our method could be useful to detect [prostate] cancer recurrence in patients that have undergone total removal of the prostate due to the presence of a tumor," Stevens said.

More generally she noted that the technique could be used as a simple screening tool up front of more complicated quantitative methods. "It could be useful to use our technique for the screening of disease biomarkers in large populations of patients," she said. "This would potentially allow the detection of early signs of disease that could then be corroborated with better established techniques."

Stevens said that she and her colleagues are now working on adapting the technique for point-of-care diagnoses and "making the test more accessible to non-experts."

She added that they "are also keen to work with global health not-for-profit organizations in order to test the suitability of the method in large populations of patients and enable its translation to resource-constrained countries."

The two diseases focused on in the Imperial College study – prostate cancer and HIV – have also been areas of focus for the immunoassay firm Quanterix, which has developed its Simoa single-molecule immunoassay system as a platform for high-sensitivity protein detection.

Indeed, last month, Quanterix scientists published a paper in the Journal of Virological Methods on using the Simoa platform to identify HIV-positive patients by detecting the same p24 protein measured by the Imperial College researchers (PM 10/12/2012).

In the study, the Quanterix team analyzed serial samples from 10 HIV-infected individuals, detecting infection at the same time as nucleic acid testing and seven to 10 days earlier than standard immunoassays.

In a statement issued upon release of the study, David Wilson, vice president of product development at Quanterix and lead author on the paper, suggested that the lower cost of the Simoa system could make it an attractive alternative to nucleic acid-based tests for HIV screening.

The Imperial College plasmonic ELISA technique could prove cheaper and better suited to resource-constrained settings still, given that it doesn't require specialized equipment like Quanterix's Simoa reader. Unlike plasmonic ELISA, however, the Simoa platform does provide quantitative results, making it more suitable for clinical applications where such information is needed.

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