Detecting Nonvolatile Life- and Nonlife-Derived Organics in a Carbonaceous Chondrite Analogue with a New Multiplex Immunoassay and Its Relevance for Planetary Exploration

Abstract

Potential martian molecular targets include those supplied by meteoritic carbonaceous chondrites such as amino acids and polycyclic aromatic hydrocarbons and true biomarkers stemming from any hypothetical martian biota (organic architectures that can be directly related to once-living organisms). Heat extraction and pyrolysis-based methods currently used in planetary exploration are highly aggressive and very often modify the target molecules, making their identification a cumbersome task. We have developed and validated a mild, nondestructive, multiplex inhibitory microarray immunoassay and demonstrated its implementation in the SOLID (Signs of Life Detector) instrument for simultaneous detection of several nonvolatile life- and nonlife-derived organic molecules relevant in planetary exploration and environmental monitoring. By utilizing a set of highly specific antibodies that recognize D- or L-aromatic amino acids (Phe, Tyr, Trp), benzo[a]pyrene (B[a]P), pentachlorophenol, and sulfone-containing aromatic compounds, respectively, the assay was validated in the SOLID instrument for the analysis of carbon-rich samples used as analogues of the organic material in carbonaceous chondrites or even Mars samples. Most of the antibodies enabled sensitivities at the 1–10 ppb level and some even at the part-per-trillion level. The multiplex immunoassay allowed the detection of B[a]P as well as aromatic sulfones in a water/methanol extract of an Early Cretaceous lignite sample (ca. 140 Ma) representing type IV kerogen. No L- or D-aromatic amino acids were detected, reflecting the advanced diagenetic stage and the fossil nature of the sample. The results demonstrate the ability of the liquid extraction by ultrasonication and the versatility of the multiplex inhibitory immunoassays in the SOLID instrument to discriminate between organic matter derived from life and nonlife processes, an essential step toward life detection outside Earth.

Simulating Mars Drilling Mission for Searching for Life: Ground-Truthing Lipids and Other Complex Microbial Biomarkers in the Iron-Sulfur Rich Río Tinto Analog

“…Powdered subsurface M and G samples were analyzed by a fluorescent sandwich microarray immunoassay (FSMI) with the LDChip immunosensor (i.e., LDChip) (Rivas et al, 2008;Parro et al, 2008;. The LDChip is a shotgun antibody microarray produced for the simultaneous detection of potential microbial biomarkers from environmental samples (Parro et al, 2018;Sanchez-García et al, 2019) and/or for detecting possible traces of life in the field of the planetary MARS SIMULATED DRILLING FOR LIFE DETECTION exploration (Blanco et al, 2017;Moreno-Paz et al, 2018) as part of the SOLID instrument concept (Parro et al, 2008(Parro et al, , 2011.…”

The Complex Molecules Detector (CMOLD): A Fluidic-Based Instrument Suite to Search for (Bio)chemical Complexity on Mars and Icy Moons

“…Here, we advocate leveraging the strong SOLID design and testing heritage (Parro et al, 2005(Parro et al, , 2008(Parro et al, , 2011a(Parro et al, , 2011bMoreno-Paz et al, 2018) to detect nonvolatile organic molecules, as either free monomers and polymers or being part of supra-macromolecular structures (including membranes, vesicles, and nano-micro particles) in liquid suspension. We have developed the SOLID instrument and it is one of the payload instruments in NASA's Icebreaker proposal mission to Mars (McKay et al, 2013).…”

“…The most obvious identified challenge of biosensors, particularly immunosensors, is that they are highly specific. However, they may also have a relatively high versatility to recognize structures that are similar and universal for different life-forms, such as the d or l chemical group on the aromatic amino acids (Kassa et al , 2011 ; Moreno-Paz et al , 2018 ), or hydrophobic stretches of proteins (regardless the aa sequence). Also, it is possible to produce antibodies (Abs) or aptamers (Aps) to detect xenobiotic (nonlife derived) molecules.…”

“…Potential molecular targets for CMOLD may be those selected to represent meteoritic input, fossil organic matter, and extant (living, recently dead) organic matter, as reviewed in the work of Parnell et al ( 2007 ), with a special focus on the potential polymeric material they can form (mainly peptides). The current list of compounds that can be detected with SOLID-LDChip comprises many of these: PAHs and derivatives such as bezo[a]pyrene and pentachlorophenol, aromatic d/l amino acids, or xenobiotics (Moreno-Paz et al , 2018 ); irradiated proteins, peptides, and other molecules (Blanco et al , 2018 ); and a variety of polymeric biomarkers from exopolysaccharide, lipopolysaccharide, humic substances, lipoteichoic acids, and peptidoglycans from bacterial cell walls, proteins from primitive metabolism, nucleic acids, and their derivatives (oxidized as 8-oxo-guanine, or thymidine dimers formed after irradiating nucleic acids with UV radiation), aromatic compounds, or short peptides (see tables from Fernandez-Calvo et al , 2006 ; Rivas et al , 2008 ; Parro et al , 2011b , 2018 ).…”

Fast Degradation of Hydrogen Peroxide by Immobilized Catalase to Enable the Use of Biosensors in Extraterrestrial Bodies

“…To address this limitation, a number of liquid-based analytical techniques have been proposed (Blanco et al, 2017;Ribette et al, 2019). Out of those, immunoassays, electrochemical sensors (Thomson et al, 2020), and microfluidic capillary electrophoresis technologies hold the most promise in terms of power requirement, weight, sensitivity, and ability to detect biomarkers in model systems (Derveni et al, 2012;Sims et al, 2012;Willis et al, 2012;Butterworth et al, 2015;Mathies et al, 2017Mathies et al, , 2019Moreno-Paz et al, 2018;García-Descalzo et al, 2019;Lezcano et al, 2019). Unfortunately, even low concentrations of H 2 O 2 present in the sample could interfere with all of these platforms because it may induce structural changes in the antibodies (required for selectivity in immunoassays) or through oxidizing the dyes used in fluorescence-based systems (Stockton et al, 2013).…”