Detecting HIV diagnostic antibodies with DNA nanomachines

A nanoscale machine composed of synthetic DNA can be used for the rapid, sensitive and low-cost diagnosis of many diseases, including HIV

New research may revolutionize the slow, cumbersome and expensive process of detecting the antibodies that can help with the diagnosis of infectious and auto-immune diseases such as rheumatoid arthritis and HIV. An international team of researchers have designed and synthetized a nanometer-scale DNA "machine" whose customized modifications enable it to recognize a specific target antibody. Their new approach, which they described this month in Angewandte Chemie, promises to support the development of rapid, low-cost antibody detection at the point-of-care, eliminating the treatment initiation delays and increasing healthcare costs associated with current techniques.

The binding of the antibody to the DNA machine causes a structural change (or switch), which generates a light signal. The sensor does not need to be chemically activated and is rapid - acting within five minutes - enabling the targeted antibodies to be easily detected, even in complex clinical samples such as blood serum.

"One of the advantages of our approach is that it is highly versatile," said Prof. Francesco Ricci, of the University of Rome, Tor Vergata, senior co-author of the study. "This DNA nanomachine can be in fact custom-modified so that it can detect a huge range of antibodies, this makes our platform adaptable for many different diseases".

"Our modular platform provides significant advantages over existing methods for the detection of antibodies," added Prof. Vallée-Bélisle of the University of Montreal, the other senior co-author of the paper. "It is rapid, does not require reagent chemicals, and may prove to be useful in a range of different applications such as point-of-care diagnostics and bioimaging".

"Another nice feature of our this platform is its low-cost," said Prof. Kevin Plaxco of the University of California, Santa Barbara. "The materials needed for one assay cost about 15 cents, making our approach very competitive in comparison with other quantitative approaches."

"We are excited by these preliminary results, but we are looking forward to improve our sensing platform even more" said Simona Ranallo, a PhD student in the group of Prof. Ricci at the University of Rome and first-author of the paper. "For example, we could adapt our platform so that the signal of the nanoswitch may be read using a mobile phone. This will make our approach really available to anyone! We are working on this idea and we would like to start involving diagnostic companies."

University of Montreal

Nanotechnology World Association

Nanotechnology produces cheaper and portable measuring instruments

Researching, creating and applying nanoparticles go hand in hand in bionanotechnology. However, it is high time for a periodic table of nanoparticles. “Scientists working in bionanotechnology find themselves in the same position as the alchemists of the early Renaissance,” says Professor Aldrik Velders in his inaugural lecture as Professor of Bionanotechnology at Wageningen University on 17 September.

Let's start by correcting one misunderstanding: nanoparticles are not extremely small. In fact, they vary in size - from a couple of atoms to millions of atoms or thousands of molecules - and make up an intermediate form between loose molecules and relatively large grains. Proteins are also a type of nanoparticle. “In light of this, chemists see nanoparticles as actually being quite large,” explains Professor Velders in his inaugural speech entitled 'Much ado about nano'. “Nanoparticles form a whole new world, complete with its own peculiarities. With this in mind, it is important that all nanoparticles which form spontaneously or are formed by human intervention are properly catalogued. This is why we are working on technology which will allow us to catalogue them effectively, adopting an optical approach via absorption and fluorescence and using magnetic resonance spectroscopy such as the technology used in MRI scans. We also do not have a periodic table of nanoparticles like the one we have for all chemical elements. We currently only know a few classes of particles, and we have very few predictive values. We also have very little understanding of how nanoparticles behave in biological systems - this despite the fact that the basis of life is found within the interaction of molecules and that nanoparticles could hold a range of useful applications.”

Creating nanoparticles

 

Aside from carrying out pure research, Velders is also active in creating therapeutic and diagnostic nanoparticles. “We can create nanoparticles which change colour when they are in the vicinity of other molecules. We are currently working on this project together with Leiden University Medical Center. Amongst other things, we aim to create applications for robot-assisted surgery. It is also possible to make hard and soft nanoparticles, as well as to insert extra molecules into the soft nanoparticles. We are currently researching how we can insert metal complexes into a large nanoparticle, or a 'micelle’, which can then be implemented as a sort of Trojan horse. This technique can be used in the first instance for diagnostic purposes; later, it could be used to administer medication into a cell. To give you an idea of the scale of this: if an atom were a large as a football, the micelle would be as large as the Main Auditorium of the university, and the cell would be a city as large as Wageningen. Alongside this, we are studying the coming together and break up of nanoparticles under the influence of biomarkers in blood samples and other areas. Changes in light absorption or fluorescence indicate that somebody has a certain disease.”

A lab on a chip

 

Velders is also involved in researching how nanotechnology can be applied. This mostly involves the development of diagnostic sensors, as he explains. “By developing miniature instruments, we can produce cheaper and portable devices that can be used everywhere. We can create a lab on a chip. This is useful for analyses of ditchwater on the campus, for instance, or for monitoring malaria infections in the field in Sub-Saharan Africa.” A significant development in this respect is that Velders and his group are now able to create microchannels in small blocks that are made of polydimethylsiloxane (PDMS), a type of rubber. They use the same plastic as Lego bricks to do this. By creating a flow of substances - in some cases cooled, heated, and/or illuminated - through the small channels, you can trigger reactions in the PDMS blocks or carry out measurements without the need for large and expensive apparatus.

NMR antennae

 

“We are also in the process of developing very small NMR antennae. Using NMR, we can observe energy in the form of radio frequencies. These are specific to an element or atom, so this can also tell us how atoms will look later in the same molecule or nanoparticle. Every element has its own specific frequency. Our nanospools can listen to all frequencies simultaneously instead of just one, which is usually the case. Our nanospools are also a lot cheaper. A normal spool can easily cost ten thousand euros, whereas our most recently development antennae cost less than one euro.”

Finally, Velders will soon become involved with the catching and removing of antibiotic-resistant bacteria from the waste water of hospitals in order to prevent these bacteria from spreading. “We are developing technology that will allow us to attach these cells to nanoplates. We can convert expensive hospital technology to purify waste water; so from nanotechnology used in refined medicines to nanotechnology used in mud.”

Wageningen University

Nanotechnology World Association

Spinning a new version of silk

Microscope images of lab-produced fibers confirm the results of the MIT researchers' simulations of spider silk. At top are optical microscope images, and, at bottom, are scanning electron microscope images. At left are fibers 8 micrometers across, and, at right, are thinner, 3 micrometer fibers.
Courtesy of the researchers

Simulations and experiments aim to improve on spiders in creating strong, resilient fibers.

After years of research decoding the complex structure and production of spider silk, researchers have now succeeded in producing samples of this exceptionally strong and resilient material in the laboratory. The new development could lead to a variety of biomedical materials — from sutures to scaffolding for organ replacements — made from synthesized silk with properties specifically tuned for their intended uses.

The findings are published this week in the journal Nature Communications by MIT professor of civil and environmental engineering (CEE) Markus Buehler, postdocs Shangchao Lin and Seunghwa Ryu, and others at MIT, Tufts University, Boston University, and in Germany, Italy, and the U.K.

The research, which involved a combination of simulations and experiments, paves the way for “creating new fibers with improved characteristics” beyond those of natural silk, says Buehler, who is also the department head in CEE. The work, he says, should make it possible to design fibers with specific characteristics of strength, elasticity, and toughness.

The new synthetic fibers’ proteins — the basic building blocks of the material — were created by genetically modifying bacteria to make the proteins normally produced by spiders. These proteins were then extruded through microfluidic channels designed to mimic the effect of an organ, called a spinneret, that spiders use to produce natural silk fibers.

No spiders needed

While spider silk has long been recognized as among the strongest known materials, spiders cannot practically be bred to produce harvestable fibers — so this new approach to producing a synthetic, yet spider-like, silk could make such strong and flexible fibers available for biomedical applications. By their nature, spider silks are fully biocompatible and can be used in the body without risk of adverse reactions; they are ultimately simply absorbed by the body.

The researchers’ “spinning” process, in which the constituent proteins dissolved in water are extruded through a tiny opening at a controlled rate, causes the molecules to line up in a way that produces strong fibers. The molecules themselves are a mixture of hydrophobic and hydrophilic compounds, blended so as to naturally align to form fibers much stronger than their constituent parts. “When you spin it, you create very strong bonds in one direction,” Buehler says.

The team found that getting the blend of proteins right was crucial. “We found out that when there was a high proportion of hydrophobic proteins, it would not spin any fibers, it would just make an ugly mass,” says Ryu, who worked on the project as a postdoc at MIT and is now an assistant professor at the Korea Advanced Institute of Science and Technology. “We had to find the right mix” in order to produce strong fibers, he says.

Closing the loop

This project represents the first use of simulations to understand silk production at the molecular level. “Simulation is critical,” Buehler explains: Actually synthesizing a protein can take several months; if that protein doesn’t turn out to have exactly the right properties, the process would have to start all over.

Using simulations makes it possible to “scan through a large range of proteins until we see changes in the fiber stiffness,” and then home in on those compounds, says Lin, who worked on the project as a postdoc at MIT and is now an assistant professor at Florida State University.

Controlling the properties directly could ultimately make it possible to create fibers that are even stronger than natural ones, because engineers can choose characteristics for a particular use. For example, while spiders may need elasticity so their webs can capture insects without breaking, those designing fibers for use as surgical sutures would need more strength and less stretchiness. “Silk doesn’t give us that choice,” Buehler says.

The processing of the material can be done at room temperature using water-based solutions, so scaling up manufacturing should be relatively easy, team members say. So far, the fibers they have made in the lab are not as strong as natural spider silk, but now that the basic process has been established, it should be possible to fine-tune the materials and improve its strength, they say.

“Our goal is to improve the strength, elasticity, and toughness of artificially spun fibers by borrowing bright ideas from nature,” Lin says.  This study could inspire the development of new synthetic fibers — or any materials requiring enhanced properties, such as in electrical and thermal transport, in a certain direction.

“This is an amazing piece of work,” says Huajian Gao, a professor of engineering at Brown University who was not involved in this research. “This could lead to a breakthrough that may allow us to directly explore engineering applications of silk-like materials.”

Gao adds that the team’s exploration of variations in web structure “may have practical impacts in improving the design of fiber-reinforced composites by significantly increasing their strength and robustness without increasing the weight. The impact on material innovation could be particularly important for aerospace and industrial applications, where light weight is essential.”

The research was supported by the National Institutes of Health, the National Science Foundation, the Office of Naval Research, the National Research Foundation of Korea, and the European Research Council.

Source link: http://newsoffice.mit.edu/2015/simulations-improve-spider-silk-0528

Build-A-Nanoparticle

An engineered Silicon-Silver nanoparticle of ~10 nanometers in size.
Image: OIST

Nanoparticles, which range from 1-100 nanometers in size, are roughly the same size as biomolecules such as proteins, antibodies, and membrane receptors.  Because of this size similarity, nanoparticles can mimic biomolecules and therefore have a huge potential for application in the biomedical field. In a paper published in Scientific Reports on October 30^th, a group of researchers from the OIST Nanoparticles by Design Unit lead by Prof. Mukhles Sowwan announced that they have succeeded in designing and creating multicomponent nanoparticles with controlled shape and structure.

Multicomponent nanoparticles, which are nanoparticles containing two or more materials, are even more powerful since they bring together the unique properties of each material to make a single nanoparticle with various functionalities. For example, a single-component nanoparticle may be able to transport drugs but may not be able to differentiate between healthy and diseased cells. In contrast, a multicomponent nanoparticle could also include characteristics of another material that can distinguish between healthy and diseased cells to make drug delivery more efficient.

The OIST researchers produced Silicon-Silver nanoparticles using advanced equipment custom-designed specifically for producing multicomponent nanoparticles. Silicon and Silver are an interesting combination because each element has different optical properties that give out different signals. A single nanoparticle simultaneously sending out multiple signals is attractive for bioimaging and biosensoring: for example, Silver would show whether a certain reaction is happening or not, while Silicon could give out information about where the nanoparticles are located.

Especially exciting is that Prof. Sowwan and his team that includes scientists from Ireland, Greece, India, United Kingdom, Peru, South Korea, Palestine, France, Spain, and Japan, can customize not only the shape and structure of the nanoparticles but also the nanoparticles’ characteristics. Engineering a particle that is 10 million times smaller than the size of a football is not easy: although nanoparticles like these have been made elsewhere in the past using different methods, they lack the level of control and purity offered at the Nanoparticles by Design Unit. With this technique, the size, structure, and  crystallinity – the orderliness of atoms –  of each nanoparticle can be customized. In this particular study, Sliver was used to control the crystallinity of Silicon. By controlling the crystallinity, optical, electrical, and chemical properties of the nanoparticle can be fine-tuned. “This is engineering. We control how we want the nanoparticles to be,” said Prof. Sowwan. 

Source: http://www.oist.jp/news-center/news/2013/11/6/build-nanoparticle

Seeing through silicon

New microscopy technique allows scientists to visualize cells through the walls of silicon microfluidic devices.

Scientists at MIT and the University of Texas at Arlington (UTA) have developed a new type of microscopy that can image cells through a silicon wafer, allowing them to precisely measure the size and mechanical behavior of cells behind the wafer.

The new technology, which relies on near-infrared light, could help scientists learn more about diseased or infected cells as they flow through silicon microfluidic devices.

“This has the potential to merge research in cellular visualization with all the exciting things you can do on a silicon wafer,” says Ishan Barman, a former postdoc in MIT’s Laser Biomedical Research Center (LBRC) and one of the lead authors of a paper describing the technology in the Oct. 2 issue of the journal Scientific Reports.

Other lead authors of the paper are former MIT postdoc Narahara Chari Dingari and UTA graduate students Bipin Joshi and Nelson Cardenas. The senior author is Samarendra Mohanty, an assistant professor of physics at UTA. Other authors are former MIT postdoc Jaqueline Soares, currently an assistant professor at Federal University of Ouro Preto, Brazil, and Ramachandra Rao Dasari, associate director of the LBRC. 

Silicon is commonly used to build “lab-on-a-chip” microfluidic devices, which can sort and analyze cells based on their molecular properties, as well as microelectronics devices. Such devices have many potential applications in research and diagnostics, but they could be even more useful if scientists could image the cells inside the devices, says Barman, who is now an assistant professor of mechanical engineering at Johns Hopkins University. 

To achieve that, Barman and colleagues took advantage of the fact that silicon is transparent to infrared and near-infrared wavelengths of light. They adapted a microscopy technique known as quantitative phase imaging, which works by sending a laser beam through a sample, then splitting the beam into two. By recombining those two beams and comparing the information carried by each one, the researchers can determine the sample’s height and its refractive index — a measure of how much the material forces light to bend as it passes through.

Traditional quantitative phase imaging uses a helium neon laser, which produces visible light, but for the new system the researchers used a titanium sapphire laser that can be tuned to infrared and near-infrared wavelengths. For this study, the researchers found that light with a wavelength of 980 nanometers worked best. 

Using this system, the researchers measured changes in the height of red blood cells, with nanoscale sensitivity, through a silicon wafer similar to those used in most electronics labs.

As red blood cells flow through the body, they often have to squeeze through very narrow vessels. When these cells are infected with malaria, they lose this ability to deform, and form clogs in tiny vessels. The new microscopy technique could help scientists study how this happens, Dingari says; it could also be used to study the dynamics of the malformed blood cells that cause sickle cell anemia.

The researchers also used their new system to monitor human embryonic kidney cells as pure water was added to their environment — a shock that forces the cells to absorb water and swell up. The researchers were able to measure how much the cells distended and calculate the change in their index of refraction. 

“Nobody has shown this kind of microscopy of cellular structures before through a silicon substrate,” Mohanty says.

“This is an exciting new direction that is likely to open up enormous opportunities for quantitative phase imaging,” says Gabriel Popescu, an assistant professor of electrical engineering and computer science at the University of Illinois at Urbana-Champaign who was not part of the research team. 

“The possibilities are endless: From micro- and nanofluidic devices to structured substrates, the devices could target applications ranging from molecular sensing to whole-cell characterization and drug screening in cell populations,” Popescu says.

Mohanty’s lab at UTA is now using the system to study how neurons grown on a silicon wafer communicate with each other.

In the Scientific Reports paper, the researchers used silicon wafers that were about 150 to 200 microns thick, but they have since shown that thicker silicon can be used if the wavelength of light is increased into the infrared range. The researchers are also working on modifying the system so that it can image in three dimensions, similar to a CT scan. 

The research was funded by the National Institute of Biomedical Imaging and Bioengineering and Nanoscope Technologies, LLC.

Source: http://web.mit.edu/newsoffice/2013/seeing-through-silicon-1002.html