Diagnosis on Earth is a fairly simple procedure, done at any hospital or medical center. In space flight, however, where doctors and even basic medical equipment may be lacking, falling ill is a serious matter for both astronaut and ground control. Using horseshoe crab blood as a reactive agent, the Lab-On-a-Chip Application Development-Portable Test System (LOCAD-PTS) is a handheld device developed by NASA researchers to help identify microorganisms. Ginger N. Flores is the LOCAD project manager.
NASA Tech Briefs:How does LOCAD-PTS work, and why was it developed?
Ginger N. Flores: It is, basically, a portable, handheld device that uses interchangeable cartridges that can detect a variety of biological and, in the future, chemical, molecules. The handheld device itself consists of a spectrophotometer, a heating block, pump, and associated electronics and microprocessors. The cartridges are about half the size of a credit card. They each contain four channels that are linked to four "sample wells" that receive the sample. In each channel a dried reagent or formulation for the detection of a particular molecule. For example, the channels of the cartridges we sent just recently to the ISS contain a formulation called Limulus amebocyte lysate, or LAL. LAL reacts with lipopolysaccarides of bacteria, also known as endotoxins. And we couple the LAL reaction to the generation of a yellow dye, and so the more intensity or the more yellow color that is detected by the device, the more litho-polysaccharide is present. That's how it works in a nutshell. Future cartridges will contain different formulations and be able to detect for different biological or chemical molecules.
As to why it was developed, LOCAD-PTS is very sensitive, and it's a very good, overall test for microbial cleanliness. It is part of the Advanced Environmental Monitoring and Control project (AEMC), and that is funded under the Exploration Technology Development Program (EDTP) by the overall exploration systems mission directorate. LOCAD-PTS has been derived from a commercial, off-the-shelf technology produced by Charles River Laboratories and then modified for space flight by the LOCAD team here at Marshall. The collaboration also includes scientists at the Carnegie Institute of Washington, and they coupled with our principle investigator to test the device in parabolic flight and other extreme environments. They've taken it to the Arctic, and also, they've used it in the Aquarius underwater habitat in the Florida Keys during NASA's NEMO mission.
NTB: How does it use horseshoe crab blood, and why was that substance chosen?
Flores: The horseshoe crab has a very primitive, but very effective, immune system, which protects the crab from microbial infection. The active player in that immune system is the blood cells, which are known as amebocytes. Back in the 1960s, it was found that an extract from those blood cells, which is LAL, could be used for very sensitive detection of the lipopolysaccarides/endotoxins. This is called the LAL assay' it's pretty widespread in molecular biology and also it is important in the medical field for the monitoring of cleanliness, specifically of injectable fluids. But it can be time-consuming; it can take hours or even days, and it can also be hazardous, because you have to use pure endotoxins as controls. So Charles River Laboratories developed the PTS part of LOCAD-PTS to reduce the assay time and also to miniaturize the whole process so that you can take it outside of the lab. The LAL test is proven. This is the reason that the LAL cartridge was the first to be chosen to do the first test that we will be performing on ISS.
NTB: Does LOCAD-PTS have a shelf life, or can it be used repeatedly?
Flores: It does have a shelf life. It has been shown to be stable for minimum of two years, and we are monitoring it on going with ground tests to see if that shelf life can be extended. They cannot be used repeatedly; a cartridge is disposable and for a one-time use. But due to their size - they are very small, very lightweight - you can take as many as you need into the field or into space to perform multiple analyses.
NTB: How does it differ from a conventional Petri dish?
Flores: A Petri dish contains a microbial growth media, and it cultures living, or viable, microorganisms. So after a few days growth, the colonies, which are also called colony-forming units (CFUs), and then they are analyzed to attain an indication of the number of the microorganisms present. It is estimated that up to about 95% of microorganisms on earth and, potentially, on the ISS, however, are "un-culturable." They are extremely difficult to grow in the lab, and have to have specific growth media.
The LOCAD-PTS is a non-culture-based technique that makes it very useful, that's very complementary to the current culture-based method. And also, and this is interesting-it makes no differentiation between dead and live cells. Petri dishes just detect live cells. Detection of dead biological material can be just as important to detect the presence of endotoxins on surfaces that need to be free of all biological material. An example would be, in the future, when we detect for life on other planets as part of our planetary protection program. We look to LOCAD-PTS as a payload on stations, as a pathfinder that can take us forward as we have other challenges in our exploration missions.
NTB: LOCAD-PTS is currently on the ISS; what, if any, are the results?
Flores: Operations for LOCAD-PTS will not begin until spring of 2007, so right now it is just stowed. Right now we are taking baby steps. Crew health diagnostics on the shuttle or ISS would be a crosscutting application that we would like to pursue. Right now, we will just be swabbing for surfaces that crewmembers do touch, so that could be an application for the future. Our main concern, when we do operate, will be that all the components of the system function normally, that we can acquire the sample, that we do not have any issues in microgravity, etc. We have performed numerous tests in our parabolic environment, and have not seen any issues to date. We want to make sure that our analysis is consistent, and we will be doing side-by-side correlation with the current culture-base method. And we expect a positive correlation between LAL and endotoxin values, between the LAL-endotoxin values and CFUs. But we do not expect it to be a very high one, because there are fundamental differences between what each method measures.
The project is a good example of how you can streamline an investigation for space. And I think that the strength of the project has been the collaborative side: we essentially have had involvement from the commercial industry, from academia, from NASA agencies, so I believe that this is a nice example of low-cost, streamlined projects leveraged off of prior work.
NTB: Can the LOCAD-PTS target a specific microbe?
Flores: It can. That's one of the advantages, and shows the versatility of the instrument. You can design the formulations for specific bacteria, fungi, yeast, or mold, in addition to families of such. For example, you could design a cartridge that detects specifically for salmonella or E. coli.
NTB: Could the NASA device be applied in terrestrial medicinal diagnoses?
Flores: Yes, and it already is. The PTS is already available commercially, and it is a great instrument for use in hospital wards, maintaining food hygiene, in genetics where you have to verify endotoxin levels in your preparation. Overall, it is a tool that is used in the pharmaceutical industry for cleanliness, and also manufacturing.
And the NASA-modified instrument could be applied on Earth as well. In fact, some of the NASA modifications that we've made in their commercial unit have been adopted, where we have made it more robust for space flight and made some material changes and internal changes to the unit.