Challenges:
There are two methods for cell lysis: physical or chemical. Physical methods require extra parts, but are not inhibited by poor mixing of fluids, as with chemical methods. More than just cell lysis, other reagents to degrade protein and DNA must be mixed with culture. The culture media may also need to be removed. How cell lysis prepare the crude cell extract for downstream processing is also a consideration. Below designs are discussed.
Current Design:
There are two design platforms: physical lysis and chemical lysis. Ideally, we want the least number of moving parts for durability and cost of manufacturing, so chemical lysis may be prefered.
There are two methods for cell lysis: physical or chemical. Physical methods require extra parts, but are not inhibited by poor mixing of fluids, as with chemical methods. More than just cell lysis, other reagents to degrade protein and DNA must be mixed with culture. The culture media may also need to be removed. How cell lysis prepare the crude cell extract for downstream processing is also a consideration. Below designs are discussed.
Current Design:
There are two design platforms: physical lysis and chemical lysis. Ideally, we want the least number of moving parts for durability and cost of manufacturing, so chemical lysis may be prefered.
- Sonication, laser and magnetic beads: These are the top three mentioned methods of cell lysis in microfluidics. Sonication uses sound waves to break apart cell membrane. A laser would also do the same thing, but may have the extra feature of detecting density of the culture. Magnetic beads, have been used to keep the culture in suspension, cell lysis, and nucleic acid purification. We would love to use this tech, but our Austrian team has attempted use of of magnetic beads and found them to be VERY difficult. We also have concerns about using this tech in space, as it is a moving part. I have heard recently that outcomes depend heavily on the brand of beads used.
- Surfactant: There are a variety of chemicals that can lyse cell membranes. The key is to have a chemical that will not interfere with downstream processing or analytic steps. Secondly, fluids do not easily mix in microfluidic devices, so complete penetration of the surfactant to all cells at the proper concentration is an additional challenge. Although laminar flow reduces mixing, it does allow for solutes to be filtered without a membrane by manipulating solvent to diffusion constants. We plan to exploit that feature. The culture stream can run parallel to a stream with surfactant, DNase and a dielectric positive charge. Triton 1X has been shown to lyse cells and keep proteins neutrally charged. This will allow for LB to be filtered and nucleic acids isolated. This separation must be tested first, before we choose this specific technology.