Technique quickly identifies bacteria for food safety

Researchers at Purdue University have used a new technique to rapidly detect and identify bacteria, including dangerous E. coli, without the time-consuming treatments usually required.

The technique, called desorption electrospray ionization, or DESI, could be used to create a new class of fast, accurate detectors for applications ranging from food safety to homeland security, says R. Graham Cooks, the Henry Bohn Hass Distinguished Professor of Chemistry in Purdue’s College of Science.

Using a mass spectrometer to analyze bacteria and other microorganisms ordinarily takes several hours and requires that samples be specially treated and prepared in a lengthy series of steps. DESI eliminates the pretreatment steps, enabling researchers to take “fingerprints” of bacteria in less than a minute using a mass spectrometer.

“This is the first time we’ve been able to chemically analyze and accurately identify the type of bacteria using a mass spectrometer, without any prior pretreatment, within a matter of seconds,” says Cooks.

New findings show how the Purdue researchers used the method to detect living, untreated bacteria, including E. coli and Salmonella typhimurium, both of which cause potentially fatal infections in humans.

This illustration depicts the use of a technique developed at Purdue to identify bacteria in its ambient environment using mass spectrometry. The technique, called desorption electrospray ionization, or DESI, could be used to create a new class of fast, accurate detectors for applications ranging from food safety to homeland security. Image courtesy of Purdue University
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“There is always an advantage to the analysis of living systems because the bacteria retain their original properties,” says Cooks.

The findings are detailed in a paper appearing January 7 in the journal Chemical Communications. The paper was written by chemistry graduate students Yishu Song, Nari Talaty and Zhengzheng Pan; Andy W. Tao, an assistant professor of biochemistry; and Cooks.

Mass spectrometry works by turning molecules into ions, or electrically charged versions of themselves, inside the instrument’s vacuum chamber. Once ionized, the molecules can be more easily manipulated, detected, and analyzed based on their masses. The key DESI innovation is performing the ionization step in the air or directly on surfaces outside of the mass spectrometer’s vacuum chamber. When combined with portable mass spectrometers also under development at Purdue, DESI promises to provide a new class of compact detectors.

Purdue researchers are focusing on three potential applications for detecting and identifying pathogens: food safety, medical analysis, and homeland security. Such a detector could quickly analyze foods, medical cultures and the air in hospitals, subway stations and airports, Cooks says.

The researchers are able to detect one nanogram, or a billionth of a gram, of a particular bacterium. More importantly, the method enables researchers to identify a particular bacterium down to its subspecies, a level of accuracy needed to detect and track infectious pathogens. The identifications are based on specific chemical compounds, called lipids and fatty acids, in the bacteria.

“We can determine the subspecies and glean other information by looking at the pattern of chemicals making up the pathogen, a sort of fingerprint revealed by mass spectrometry,” claims Cooks. “Conventional wisdom says quick methods such as ours will not be highly chemically or biologically specific, but we have proven that this technique is extremely accurate.”

The procedure involves spraying water in the presence of an electric field, causing water molecules to become positively charged hydronium ions, which contain an extra proton. When the positively charged droplets come into contact with the sample being tested, the hydronium ions transfer their extra proton to molecules in the sample, turning them into ions. The ionized molecules are then vacuumed from the surface into the mass spectrometer, where the masses of the ions are measured and the material analyzed.

Such a system could alert employees in the food and health-care industries to the presence of pathogens and could provide security personnel with a new tool for screening suspicious suitcases or packages.

Song will further the research, conducting experiments to look for bacterial contaminants in foods. Ongoing work by Talaty with international E. coli expert Barry Wanner, a professor in Purdue’s Department of Biological Sciences, will apply the method to living bacteria in so-called biofilms.

DESI has been commercialized by Indianapolis-based Prosolia Inc.
- Emil Venere

Company develops 50V CMOS process with embedded flash

Austriamicrosystems’ foundry business unit announced a 50V High-Voltage CMOS process with embedded Flash. The company says this is the consequent next step in extending austriamicrosystems’ position in High-Voltage CMOS technology.

The new Flash process technology is, like the High-Voltage CMOS technology, a modular extension of austriamicrosystems’ 0.35µm CMOS process. Complete compatibility to the base process allows re-use of IP Blocks and adding Flash memory to it on a single chip. The company says its High-Voltage CMOS process is suited for harsh environment, making it ideal for designs in power management, automotive or medical applications.

Draper inertial stellar compass fully operational

Draper Laboratory in Cambridge, Mass., announced that its Inertial Stellar Compass (ISC) is now fully operational on board the TacSat-2 spacecraft, representing the first use of a MEMS gyro in a complete spacecraft attitude determination system.

TacSat-2 was launched on December 16 from Wallops Flight Facility. Following basic spacecraft commissioning activities, the ISC was first turned on December 27, and two days of preliminary functional tests show the instrument to be working perfectly.

Draper Laboratory’s Inertial Stellar Compass (ISC) is now fully operational on board the TacSat-2 spacecraft. Photo courtesy of Draper Laboratory
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The ISC combines a star camera and MEMS gyros with a microprocessor to provide a full 3-axis attitude determination system in a low power (3.6W) and low-mass (2.9kg) package, less than one-half the power and mass of conventional systems. It was developed at Draper Laboratory and uses Draper’s MEMS Tuning Fork Gyro package.

The lab says the fully autonomous, self-initializing instrument has operated flawlessly since being powered up on Dec. 27. Requiring no more than power and an occasional clock update from the host spacecraft, the ISC initializes upon startup, acquires and identifies stars from its own star catalog, and uses its “lost-in-space” algorithms to determine the direction in which it is pointing. If all continues to go well aboard the TacSat-2 spacecraft, a series of dedicated tests are planned for the coming weeks that will acquire extensive data to verify ISC performance under a wide variety of conditions.

The ISC development was funded by NASA’s New Millennium Program (NMP), which is managed by the Jet Propulsion Laboratory in Pasadena, Calif. The TacSat-2 spacecraft was developed by the Air Force Research Laboratory (AFRL) and is operated out of the AFRL command center at Kirtland AFB in Albuquerque, N.M.

NanoSensors initiates first prototype production of product to detect E. coli

NanoSensors Inc., a Santa Clara, Calif., nanotechnology development company that develops instruments and sensors to detect explosives and chemical and biological agents, announced that it has engaged a third-party contractor to manufacture units of a test version of its biosensor product that will be used for third-party field testing.

The company expects to receive delivery of the product units and commence testing during the first quarter of 2007 in order to obtain feedback on the performance of the biosensor. The product is based on the company’s recently licensed nanoporous silicon-based biosensor technology to detect E. coli.

Although NanoSensors has not entered into testing agreements with third parties, it is seeking to enroll between 6 and 10 users in its product testing program in order to subject the product design to simulated field conditions and to further assess the commercial viability of the current design. It is expected that the testing cycle will continue for a period of approximately three months and the company intends to deploy the biosensors in a number of different testing conditions. After the testing period is complete, the company said it intends to incorporate user feedback into the product design in order to improve product functionality, as may be appropriate.

The proposed biosensor has been designed to consist of two core functional parts: a disposable housing unit in which the actual sensor device is mounted and a separate, external data acquisition unit.

Based on this design, the disposable housing unit that contains the sensor transmits signals across electrical leads to the data acquisition unit, which accepts the output signal from the disposable housing unit and converts the signal to the appropriate format to display the results.

Carbon nanotubes provide critical link to block HIV

Researchers led by Hongjie Dai at Stanford University are using carbon nanotubes to solve the challenge of efficient and targeted delivery of RNA into cells. Solving the problem promises a new type of gene therapy that involves binding short DNA fragments (interfering RNA) to specific genes to block their “translation” into the corresponding, disease-related protein.

The use of carbon nanotubes has allowed the researchers to successfully introduce RNA fragments that “switch off” the genes for special HIV-specific receptors and co-receptors on the cells’ surface into human T-cells and primary blood cells. This leaves few entry points for the HIV viruses. The researchers report in the journal Angewandte Chemie that this allows for much better silencing effect to the cells than current transport systems based on liposomes.

T-cells are one of the types of white blood cells important for a good immune defense; they detect and destroy virus-affected cells. However, they themselves are among the targets attacked by HIV. In order to enter into a T-cell, the virus must first dock to a receptor known as CD4. Also involved is the co-receptor CXCR4. The use of short interfering RNA strands allows the CD4 and CXCR4 genes of the T-cell to be shut off. The T-cell then strops producing these receptors and the virus cannot find any points of attack on the surface of the cell. This could significantly slow down an HIV infection, as previous work has shown.

But how to get the RNA fragments into the T-cells? The shells of nonpathogenic viruses can be used to smuggle genetic material into cells, but this is dangerous in therapeutic applications because they can trigger allergies. Liposomes, tiny bubbles of fat, are safe but have proven to be ineffective for use in T-cells.

Carbon nanotubes are known for their abilities to be absorbed by cells and to smuggle in other molecules at the same time. The researchers attached phospholipids-molecules from which cell membranes are also made-to chains of polyethylene glycol. The phospholipids nestle securely onto the outer wall of the carbon nanotubes while the polyethylene glycol chains protrude into the surrounding solution.

The required RNA molecules were fastened to the ends of these chains. Once inside the cell, the RNA could easily be split off by means of a sulfur-sulfur bridge.

EPA issues nanotechnology White Paper

The Environmental Protection Agency has issued its Nanotechnology White Paper, EPA 100/B-07/001, to inform EPA’s management of the science issues and needs associated with nanotechnology, to support related EPA program office needs, and to communicate the identified nanotechnology science issues to stakeholders and the public.

The white paper provides background information regarding nanotechnology and various environmental issues and discusses the risk assessment of nanomaterials, the environmentally responsible development of nanoscale materials, and the EPA’s research needs regarding nanomaterials.

Nanoparticles shed light on disease-causing proteins

The problem with current protein profiling methods is that the small samples are so sensitive that “we can’t effectively use existing technologies to study them,” says Andy Tao, a Purdue University biochemist. In an effort to discover a better way to ascertain the presence, concentration, and function of proteins involved in disease processes, Tao and his colleagues bound a complex nanomolecule, called a dendrimer, with a glowing identification tag delivered to specific proteins in living venom cells from a rattlesnake.

The researchers hope the new method will also facilitate better, more-efficient diagnosis in living cells and patients. Because molecular interactions and protein functions are disturbed when samples are collected, researchers can’t obtain an accurate picture of biochemical mechanisms related to illnesses such as cancer and heart disease.

Andy Tao uses a linear ion trap mass spectrometer to analyze several hundred proteins per hour. Photo courtesy Purdue Agricultural Communication/Tom Campbell
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Tao and his research team used dendrimers because they can pass through cell walls efficiently with little disturbance to the cells and then label specific proteins with isotopic tags while cells are still alive. This allows the scientists to determine the activities of proteins that play roles in specific diseases. Proteins carry genetic messages throughout the cell causing biochemical changes that can determine whether a cell behaves normally or abnormally. Proteins are also important in directing immune responses.

The team, which includes Purdue postdoctoral student Minjie Guo and Purdue graduate student Jacob Galan, report on their new strategy to discover proteins and protein levels, called soluble polymer-based isotopic labeling (SoPIL), in the current issue of the journal Chemical Communications. The study is also featured in the journal’s news publication Chemical Biology.

The dendrimers would carry one of the stable isotopic or fluorescent labels to identify the presence or absence of a protein that can be further developed for use as a disease indicator, or biomarker.

Snake venom cells were used because they have a very high concentration of proteins similar to some found in human blood, Tao says. The proteins apparently are part of the biochemical process that affects blood clotting or hemorrhage. Understanding how the proteins behave could help determine predisposition to heart disease and cancer and also be useful in diagnosis and drug development.

In future research, Tao plans to investigate how dendrimers are able to enter the cell so easily, what happens to them once they are in the cell, and whether there are any long-term effects.

Molecular memory breakthrough using nanowires

A team of UCLA and California Institute of Technology chemists reported in the January 25 issue of the journal, Nature, the successful demonstration of a large-scale, “ultra-dense” memory device that stores information using reconfigurable molecular switches. This research represents an important step toward the creation of molecular computers that are much smaller and could be more powerful than today’s silicon-based computers. The 160-kilobit memory device uses interlocked molecules manufactured in the UCLA laboratory of J. Fraser Stoddart, director of the California NanoSystems Institute (CNSI).