nai

Project Progress

1. Arctic Mars Analog Svalbard Expedition (AMASE) 2007

AMASE 2007 was the latest of a series of expeditions whose primary goals are to test portable instruments for their robustness as field instruments for life detection (for robotic and future human missions to Mars), to assess the Mars analog environments for signs of life, to refine protocols for contamination reduction, and to understand the effects of transport on sample integrity by assessing bioloads immediately in the field and then comparing these with laboratory measurements made after transportation. A wide variety of science instruments and platforms was deployed on AMASE 2007, including the two instrument prototypes for the Mars Science Laboratory (MSL) mission: the Sample Analysis at Mars (SAM) Gas Chromatograph-Mass Spectrometer and the CHEmistry and MINeralogy (CheMin) X-Ray Diffraction/X-Ray Fluorescence (XRD/XRF) instrument. Four instruments from the European Space Agency (ESA) ExoMars mission were also deployed (through funding to AMASE Expedition Leader Hans Amundsen, Earth and Planetary Exploration Services — EPX A/S, Norway). These included a remote Raman spectrometer, ground-penetrating radar, infrared spectrometers, and the panoramic camera system. Other instruments included a portable Raman spectrometer, an ultraviolet excitation spectrometer (laser-induced native fluorescence), a digital color microscope, portable lab-on-a-chip test systems, and a complete polymerization chain reaction (PCR) system.

Steele and his team continued to test a “cliffbot” rover system for accessing steep terrain. The cliffbot is equipped with a robotic arm containing a scoop and microscopic imager. The rover also was fitted with a prototype sample containment system to test preliminary designs for Mars sample return. NAI funding aided in preparing instruments for field deployment and supporting Postdoctoral Fellow Mihaela Glamoclija’s participation in the expedition.

2. Oligonucleotide microarrays for space flight applications

Microorganisms interfere with or are crucial to goals and systems for space flight, and the development of effective microbial monitoring technologies is consequently critical for mission safety and success. Recognizing the need for early microbial identification, Steele and Doctoral Student Verena Starke, in collaboration with Marshall Space Flight Center, Affymetrics, and the National Institute of Health have developed molecular-based technology for microbial monitoring. Steele’s team designed a resequencing microarray with Affymetrix for the identification of microorganisms relevant to human space flight (Figure 1). In molecular biology microarrays have become an ubiquitous tool, due to their potential to simultaneously interrogate large quantities of genetic information in a single experiment. The custom array design interrogates 16S or 18S rRNA gene sequences in 157 organisms, probing 527829 base pairs of a sequence on a single array. DNA is hybridized onto the array, and, depending on the DNA present, characteristic patterns of fluorescent response are expected as a result. Steele and his team have developed a novel algorithm for the classification of microbes. The algorithm “SMAC” (Singular-value Microarray Analysis of Concentrations) developed for this work is based on a linearity assumption, namely that the fluorescent pattern measured on the microarray is a superposition of characteristic patterns for organisms. This linear model is subsequently used to estimate organism concentrations through a least square procedure, which involves estimating the pseudo-inverse of the pattern matrix. Micorarray analysis correctly detected individual organisms as well as mixtures of organisms. Additionally, proper estimates of relative concentrations were generated. Discrimination among organisms was excellent, even for those with high sequence similarity such as species from the same family. The size of the array will be drastically reduced in future implementations, because sensitivity and specificity did not degrade when using only 1% of the current probes. Alternatively, many more organisms could be probed simultaneously from a single chip. These results indicate that microarrays are an effective tool for microbial identification and monitoring.

3. Testing microarrays under space flight conditions

Antibody microarray technology has been identified as a potential method for life detection as part of the European Space Agency’s ExoMars mission to Mars, currently scheduled for launch in 2013. The antibody-microarray-based Life Marker Chip (LMC) experiment, which houses a number of antibody microarrays, each designed to detect up to 25 different molecular targets relevant to life detection on Mars, has been proposed as part of this mission. Steele and his team are working in collaboration with Marks Sims and David Cullen of the U.K.'s Cranfield and Leicester Universities. In preparation for the ExoMars mission, representative components of the LMC were launched from Baikonur in September 2007 within the BIOPAN-6 space exposure platform aboard the FOTON-M3 spacecraft. During the 12-day mission, FOTON underwent over 180 low-Earth orbits at an altitude of 260-305 km, and for 10 days BIOPAN was opened to directly expose the LMC components to the radiation conditions of space. The LMC team tested the ability of three types of shielding-“Infinite” (4 mm aluminum + 2 mm stainless steel), “ExoMars equivalent” (4 mm aluminum) and “Zero shielding” (Kapton foil)—to protect microarray integrity during the spaceflight. The microarrays consisted of fluorescent markers, antibodies, and other proteins printed onto 4.5-mm2 epoxy-coated glass and silicon chips and were fitted into the LMC housing before integration in the BIOPAN-6 platform. Upon landing, the integrity of each microarray was analyzed by fluorescence binding assays and compared with identical control microarrays shipped to Baikonur but not flown. Information gained from this study is still underway and will be used to guide the selection and development of the components to be used within the LMC experiment proposed for the ExoMars mission.

4. Protein microarray optimization

The bulk of work this year focused on antibody microarray optimization for use in life-detection technology. Research Staff member Lauren Kerr concentrated on optimizing protocols such as extraction of biomolecules of interest from rock substrates, design and preparation of specific microarrays for the detection of cyanobacterial markers from cryptoendolithic communities, and hybridization of target proteins for the microarray. A variety of buffers to stabilize antibodies on the array surface to maintain the long-term stability of arrays have been tested. These microarrays were then tested within a field deployable system on the Arctic Mars Analogue Svalbard Expedition (AMASE).

5. Lab-On-a-Chip Application Development Portable Test System (LOCAD-PTS)

The Lab-On-a-Chip Application Development Portable Test System (LOCAD-PTS) detects biological molecules found in the cell walls of bacteria and fungi. It was launched to the International Space Station (ISS) in December 2006 aboard Space Shuttle STS-116. LOCAD-PTS has been operated onboard the ISS on eleven separate occasions since March 2007, with the most recent operating session in June 2008.

The aims of LOCAD-PTS are to address goals of NASA’s Exploration Systems Mission Directorate (ESMD) regarding the maintenance of crew health during flight, and the Science Mission Directorate (SMD) and NAI regarding the development of technology and procedures necessary to monitor and document biological contamination during future human exploration missions. NAI funding has been supporting the series of laboratory tests by Steele and Collaborator Jake Maule needed to support flight activities (Figure 2).

LOCAD-PTS is a hand-held device that employs multiple interchangeable cartridges for the detection of three microbial markers: (i) endotoxin, (ii) beta-1, 3-glucan, and (iii) lipoteichoic acid. As of August 2008, the first two types of cartridge are aboard the ISS, with the third type (for detection of lipoteichoic acid, found in gram-positive bacteria) scheduled for launch to the ISS aboard flight ULF2 in November 2008.

LOCAD-PTS is the first demonstration of complete biochemical analysis of environmental samples onboard a space station — from sampling to onboard data – and serves as a foundation for a future generation of more complex tests. The ease with which crewmembers – pilots and scientists – performed procedures was especially important, given the relatively high levels of hand-eye coordination and control were required.