We use smartphones to find directions when we’re lost, to send photos and videos to family and friends in seconds, even to sleuth out the best place to go for lunch in a three-mile radius. However, scientists at the University of Houston’s UH Cullen College of Engineering are harnessing the pocketable power of smartphones for a task with much more gravitas: diagnosing diseases in real time.
The UH researchers are developing a disease diagnostic system that offers results that could be read using only a smartphone equipped with a $20 lens attachment.
The system is the brainchild of Dr. Jiming Bao, a UH assistant professor of electrical and computer engineering, and Dr. Richard Willson, Huffington-Woestemeyer Professor of Chemical and Biomolecular Engineering, and Professor of Biochemical & Biophysical Sciences at UH, and also affiliated with the Centro de Biotecnología FEMSA, Departamento de Biotecnología e Ingeniería de Alimentos, Tecnológico de Monterrey at Monterrey, Mexico. The invention was created with the help of funding grants from the National Institutes of Health and The Welch Foundation, and was featured in February in the American Chemical Society journal ACS Photonics.
The ACS Photonics paper, titled “Transmissive Nanohole Arrays for Massively-Parallel Optical Biosensing” is co-authored by Drs. Bao and Willson, with colleagues Yanan Wang, Archana Kar, Andrew Paterson, Katerina Kourentzi, Han Le, and Paul Ruchhoeft.
The researchers explain in the article abstract that a high-throughput optical biosensing technique is proposed and demonstrated that combines optical transmission of nanoholes with colorimetric silver staining. The size and spacing of the nanoholes are chosen so that individual nanoholes can be independently resolved in massive parallel using an ordinary transmission optical microscope, and, in place of determining a spectral shift, the brightness of each nanohole is recorded to greatly simplify the readout.
Each nanohole then acts as an independent sensor, and the blocking of nanohole optical transmission by enzymatic silver staining defines the specific detection of a biological agent. Nearly 10,000 nanoholes can be simultaneously monitored under the field of view of a typical microscope. As an initial proof of concept, biotinylated lysozyme (biotin-HEL) was used as a model analyte, giving a detection limit as low as 0.1 ng/mL.
Because of their small size, nanohole biosensors can be integrated with microfluidic channels to create portable lab-on-a-chip devices, but while EOT nanoholes are compact and sensitive, there are several drawbacks that have prevented either the peak-shift or the light-transmission EOT-based nanohole techniques from being widely used in laboratories or clinical diagnostics that the coauthors go on to describe. In their paper, they report how these issues were addressed and overcome in their research, concluding that a new nanohole-based biosensing technique combining direct nanohole transmission imaging and chemical staining was achieved in this initial demonstration, and suggest that the technique is general and can be applied to a wide range of biological agents from proteins to large pathogens. They predict that when combined with microfluidics, the technique should become more compact and efficient and may lead to more practical, real-life clinical diagnostics applications.
This new technique, like essentially all diagnostic tools, relies on specific chemical interactions that form between something that causes a disease, such as a virus or bacterium, and a molecule that bonds with that one thing only, like a disease-fighting antibody. For example, a bond formed between a strep bacterium and an antibody that interacts only with strep can support a confident diagnosis.
The difficult part is finding a way to detect these chemical interactions quickly, cheaply and easily. The solution proposed by Drs. Bao and Willson involves a simple glass slide and a thin film of gold with thousands of holes poked in it.
However, creating this slide is itself an achievement. This task, led by Dr. Bao, and described in a UH release, starts with a standard slide covered in a light-sensitive material known as a photoresist. He next uses a laser to create a series of interference fringes – essentially lines – on the slide; then rotates it 90 degrees and creates another series of interference fringes. The intersections of these two sets of lines creates a fishnet pattern of UV exposure on the photoresist. The photoresist is then developed and washed away.
Next, the slide is exposed to evaporated gold, which attaches to photoresist and the surrounding clean glass surface. Dr. Bao then performs a procedure called “lift-off,” which essentially washes away the photoresist pillars and the gold film attached to them, with the end result being a glass slide covered by a film of gold with ordered rows and columns of transparent holes where light can pass through. These “nanoholes,” measuring some 600 nanometers each, are key to the system. Drs. Willson and Bao’s device diagnoses an illness by blocking the light with a disease-antibody bond – aided by a few additional ingredients.
That is where Dr. Willson comes in. An internationally known biomolecular engineer, Willson starts by placing disease antibodies in the holes, where they are coaxed into sticking to the glass surface. Next, he flows a biological sample over the slide. If the sample contains the bacteria or virus being sought out, it will bond with the antibody in the hole.
This bond alone, though, doesn’t block the light. “The thing that binds to the antibody is probably not big and grey enough to darken this hole, so you have to find a way to darken it up somehow,” Dr. Willson notes in the release. He achieves the desired result by flowing a second round of antibodies that bond with the bacteria over the slide. Attached to these antibodies are enzymes that produce silver particles when exposed to certain chemicals. With this second set of antibodies now attached to any bacteria in the holes, Dr. Willson then exposes the entire system to the chemicals that encourage silver production.
After about 15 minutes he rinses off the slide, and thanks to chemical properties of the gold, the silver particles in the holes will remain in place, completely blocking light.
Now we get to the smartphone’s role. One of the advantages of this system is that results can be read with simple tools. A basic microscope used in elementary school classrooms, Dr. Willson says, provides sufficient light and magnification to show whether the holes are blocked. With a few small tweaks, a similar reading could almost certainly be made with a phone’s camera, flash and an attachable lens.
This system, then, promises readouts that are affordable and easy to interpret. “Some of the more advanced diagnostic systems need $200,000 worth of instrumentation to read the results,” Dr. Willson Notes. “With this, you can add $20 to a phone you already have and you’re done.”
Dr. Willson cautions that there are still major technical hurdles to clear before the system can be rolled out, one of the biggest challenges being to find a way to drive the bacteria and viruses in the sample down to the surface of the slide to ensure the most accurate results. However, if those problems are overcome, the system would be an excellent tool for healthcare providers in the field.
At the site of an industrial accident, for instance, the holes on a single slide could be populated with molecules that bond with 10 potential contaminants, allowing response teams to quickly assess the situation. In economically disadvantaged areas, such a system could be used to screen large groups of people for widespread and serious health problems, like diabetes.
“There are a lot of situations where an affordable diagnostic tool that is simple to use and simple to interpret could be very useful,” says Dr. Willson. “If both your disposables and your reader are cheap, that makes it a lot easier to extend your system out into the real world.”
The ACS Photonics paper is available at:
University of Houston
University of Houston
Jiming Bao Research Group