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Evaluation of cell lysis procedures and use of a micro fluidic system for an automated DNA-based cell identification in interplanetary missions

Hall, J. A. and Felnagle, E. and Fries, M. and Spearing, S. and Monaco, L. and Steele, A. (2006) Evaluation of cell lysis procedures and use of a micro fluidic system for an automated DNA-based cell identification in interplanetary missions. Planetary and Space Science, 54 (15). pp. 1600-1611. ISSN 0032-0633. doi:10.1016/j.pss.2006.06.021.

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A Modular Assay System for Solar System Exploration (MASSE) is being developed to include sample handling, pre-treatment, separation and analysis of biological target compounds by both DNA and protein microarrays. To better design sensitive and accurate initial upstream sample handling of the MASSE instrument, experiments investigating the sensitivity and potential extraction bias of commercially available DNA extraction kits between classes of environmentally relevant prokaryotes such as gram-negative bacteria (Escherichia coli), gram-positive bacteria (Bacillus megatarium), and Archaea (Haloarcula marismortui) were performed. For extractions of both planktonic cultures and spiked Mars simulated regolith, FTA® paper demonstrated the highest sensitivity, with detection as low as ~1×10^1 cells and ~3.3×10^2 cells, respectively. In addition to the highest sensitivity, custom modified application of FTA® paper extraction protocol is the simplest in terms of incorporation into MASSE and displayed little bias in sensitivity with respect to prokaryotic cell type. The implementation of FTA paper for environmental microbiology investigations appears to be a viable and effective option potentially negating the need for other pre-concentration steps such as filtration and negating concerns regarding extraction efficiency of cells. In addition to investigations on useful technology for upstream sample handling in MASSE, we have also evaluated the potential for μTAS to be employed in the MASSE instrument by employing proprietary lab-on-a-chip development technology to investigate the potential for microfluidic cell lysis of different prokaryotic cells employing both chemical and biological lysis agents. Real-time bright-field microscopy and quantitative PMT detection indicated that that gram positive, gram negative and archaeal cells were effectively lyzed in a few seconds using the microfluidic chip protocol developed. This included employing a lysis buffer with components including lysozyme, Protease, Proteinase K, Tween-20 and TritonX-100. The effectiveness of antibiotics and other chemical lysis agents were also screened and demonstrated partial effectiveness on all three cell types. This work demonstrates a step wise approach to evaluating the efficacy and sensitivity of commercial macro-scale technology and state-of-the-art developmental microfluidic technology under consideration for incorporation into the remotely operated MASSE instrument currently under development at the Carnegie Institution of Washington.

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Additional Information:© 2006 Published by Elsevier Ltd. Received 13 December 2005; accepted 19 June 2006. Available online 27 September 2006. The authors wish to acknowledge Marilyn Fogel for helpful experimental comments, Rachel Schelble for manuscript editing in addition to the NSF Research Experience for Undergraduates program.
Subject Keywords:life detection; cell lysis; FTA paper; microfluidic
Issue or Number:15
Record Number:CaltechAUTHORS:20110629-094637764
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Official Citation:J.A. Hall, E. Felnagle, M. Fries, S. Spearing, L. Monaco, A Steele, Evaluation of cell lysis procedures and use of a micro fluidic system for an automated DNA-based cell identification in interplanetary missions, Planetary and Space Science, Volume 54, Issue 15, First Results on Titan from VIMS Observations Onboard the Cassini/huygens Mission/Astrobiology: The Detection of Life in the Solar System, December 2006, Pages 1600-1611, ISSN 0032-0633, DOI: 10.1016/j.pss.2006.06.021. (
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:24247
Deposited By: Tony Diaz
Deposited On:29 Jun 2011 18:54
Last Modified:09 Nov 2021 16:22

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