CMU Science and Technology
Center
NASA-ASTEPNASA-ASTEP
Detect and confirm sparse life on MarsAtacoma Desert Robot Study as model
CMU Science and Technology
Center
Science objectivesScience objectives
•To explore the limits of sparse microbe detection with minimal sample manipulation using a sensitive fluorescence instrument and innovative fluorescence reagents.
•To test the concept of multicolor biomarker correlation to minimize false positive identification of microbes.
CMU Science and Technology
Center
Fluorescence imagingFluorescence imaging
camera
LED illuminators
light skirt
Xe flash illuminators
robot frame
black box
CMU Science and Technology
Center
Wavelength selectionWavelength selection
A1 A2 A3
B1 B2 B3
C1 C2 C3
push pull
push pull
push pull
autofocus element
system lens
protective sapphire plate
CCD
filter changers
shutter
CMU Science and Technology
Center
Chlorophyll emission spectraChlorophyll emission spectra
Bacteriochlorophylls
0.0
1.0
250 350 450 550 650 750 850 950
C B
A
Eukaryotic Chlorophylls
0.0
1.0
250 350 450 550 650 750 850 950
wavelength (nm)
A
B
Figure 2: Absorption spectra of bacteriochlorophylls a, b, and c (upper panel) and eukaryotic chlorophylls a and b (lower panel).
CMU Science and Technology
Center
Detection of chlorophyll fluorescenceDetection of chlorophyll fluorescence
Figure 5. Pennsylvania sandstone with photosynthetic biofilm. Biofilm is less than 100 micrometers thick at greatest depth. Left, visible light. Center and right images are fluorescence images, emission at 820nm (near-infrared fluorescence) for both. Center, 365 nm excitation;30sec exposure; right, 665nm excitation, 3sec exposure. The 820 nm emission captures the near-infrared tail of the chlorophyll emission. Notice the much-improved contrast between background signal and chlorophyll fluorescence obtained with longer wavelength emission. This figure illustrates the sensitivity of detection possible even with weak excitation and shows that wavelength selection capability is essential for optimal contrast.
CMU Science and Technology
Center
Properties of available dyesProperties of available dyes
Dye Absorption max (nm) Emission max (nm) Enhancement on bindingDNA probesSYTO 9 470 530 ++++SYTO BC 480 500 ++++DRAQ56 506 700
Protein probesFluorescamine 380 464 ++++OPA/2ME 334 455 +++NDA/CN 419 493 +++CBQCA/CN 465 560 +++SYPRO Orange 470 570 +++SYPRO Red 550 630 ++++
Lipid probesDPH 353 449 +++Nile Red 559 637 +++DiI(7) 750 780 +
Carbohydrate probeCalcofluor White 360 480 +++
CMU Science and Technology
Center
Fluorescent biofilm indicatorFluorescent biofilm indicator
Figure 1: E. coli at edge of biofilm visualized using SYPRO Red dye.
CMU Science and Technology
Center
Relevant microorganismsRelevant microorganisms
Class Organism
Cyanobacteria Synechocystis sp.
Nostoc sp.
Eubacteria Escherichia coli
Bacillus megatherium
Burkholderia cepacia
Nitrosomonas europaea
Extremophiles Thermus thermophilus
Halobacterium salinarum
Shewanella frigidimarina
Acidovorax facilis
CMU Science and Technology
Center
Deliverable year 1Deliverable year 1
1: In Carnegie Mellon laboratories, completed testing of original fluorescent reagents on various soil samples and evaluation of reagents and instrumentation concepts.
2: Produce formal designs and construction of multicolor fluorescence detection system
3: Develop dye spritzing system.
CMU Science and Technology
Center
Deliverables year 2Deliverables year 2
1: Design and construct “drop box” containing multicolor fluorescence imaging unit. 2: Produce and evaluate the capabilities of the fluorescent reagents and drop-box imaging system placed manually on sites in Pennsylvania and the Southwest United states. Evaluation requires laboratory analysis of material from the sites.
3: Produce improvements and substitutes for reagents that do not function properly and make improvements to imaging system, excitation system and drop box.
4: Identify partner for robotic field study and produce machine plans for attachment of drop box imaging system to a rover (DONE!!).
CMU Science and Technology
Center
Deliverables year 3Deliverables year 3
1: Mount drop box imaging system on rover.
2: Produce results of field study for system on rover with laboratory analysis of material from sites visited.
3: Improved system instruments and reagents
4: Documentation and dissemination of results of studies.
CMU Science and Technology
Center
PersonnelPersonnel
Investigators Institution Role Work Commitment
FY03 FY04 FY05
Alan Waggoner Carnegie Mellon/STC
PI [10] [10] [10]
Victor Weedn Carnegie Mellon/STC
Co-I 10 10 10
Byron Ballou Carnegie Mellon/STC
Co-I 15 15 15
Lauren Ernst Carnegie Mellon/STC
Co-I 30 30 30
Ned Minkley Carnegie Mellon/BHE
Co-I 12 12 12
Dave Wettergreen Carnegie Mellon/RI Collaborator
Nathalie Cabrol NASA Ames/SETI Collaborator
Jeff Moersch Univ of Tennessee Collaborator
Rocco Mancinelli NASA Ames/SETI Collaborator