When a patient develops sepsis, a potentially fatal inflammatory response to an infection, doctors race to identify what bacterial strain is to blame so they can deliver the best antibiotic. This hunt can take days, however, which can be the difference between life and death. Now, researchers have developed a chip that detects bacteria in a blood sample within a couple of hours, potentially streamlining the diagnosis and treatment of sepsis (Anal. Chem. 2016, DOI:10.1021/acs.analchem.6b00323).
In standard sepsis diagnosis, technicians culture blood samples to increase the concentration of bacteria before testing for the pathogens. Scientists are developing testing methods based on the polymerase chain reaction (PCR) that work directly on unconcentrated blood samples, but these still require preparation steps like removing blood cells and other blood components that could inhibit the PCR reaction. To simplify and accelerate bacteria detection, Thomas Laurell of Lund University developed a microfluidic chip that integrates bacteria enrichment and PCR on a single platform, and requires just a drop of blood.
The device has three distinct regions connected via microfluidic channels. The first area removes red and white blood cells, the next concentrates the bacteria, and a third amplifies and detects bacterial DNA using combined PCR and fluorescence. In the first two steps, acoustic forces focus and trap cells and bacteria. A piezoceramic transducer, which vibrates like a cell phone, generates standing waves along the device’s channel. As the blood cells are pumped through the channel, they encounter a wave node, which focuses them into the center of the stream, allowing them to be diverted into a waste compartment.
The bacteria are too small to even notice the sound wave, Laurell says, so they continue on to the next part of the device, where they run into a honeycomb of polystyrene particles. Here, the sound waves scatter off the particles creating an acoustic force that attracts the bacteria to the particles. “It’s like an acoustic magnet,” Laurell says. After the researchers wash away the remaining blood components, they turn off the sound, releasing the bacteria from the particles. The bacteria then flow into the third stage with the necessary reagents for PCR and detection. This part of the chip sits inside a thermocycler that carries out the PCR amplification and the subsequent fluorescent bacterial detection.
To test the device, Laurell’s team spiked whole blood with Pseudomonas putida bacteria at varying concentrations and ran the samples through the device. They successfully detected bacterial DNA in samples containing as little as 1,000 bacteria per mL. Next, the researchers tested samples from septic patients identified using the standard blood culture approach. The device successfully identified Escherichia coli from half of those patients. This is not good enough yet, obviously, but Laurell says he’d like to increase the sensitivity enough to be able to detect as little as 1 to 100 bacteria/mL.
“One of the major challenges in the field has been a lack of integration. Current systems are either bulky or complex, so they aren’t suited for point-of-care applications,” says Weian Zhao of the University of California, Irvine. “What I like about this paper is they incorporate sample processing, bacteria enrichment, and PCR into one system.”
Source: Chemical and Engineering News