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4.6.1 Combining Metabolomics with Other Omics Technolgies
LC-MS has been successfully utilized for metabolomics, but previous sample preparation methods made metabolomics incompatible with proteomic and lipidomic
analysis. In order to maximize the data extracted from a sample, a new method
known as sample preparation for multi-omics technologies (SPOT) has been developed for high-throughput multi-omics analysis by various collaborations at
Vanderbilt University [25, 91]. This technology allows for proteomic, transcriptomic, and metabolomic analysis from the same sample, with common sample preparation methodology. This common preparation allows for high-throughput sample
analysis, which would be optimal for applications such as rapid threat assessment.
This LC-MS based method allows for temporally resolved data sets in addition to
multi-omics analyses, optimal for addressing complex bio-logical questions.
This novel multi-omics sample preparation method utilizes cells but can be
applied to tissue samples as well. Cells are lysed, undergo a freeze-thaw cycle, and
then are sonicated in an ice bath. Aliquots are then lysed and precipitated with 75:25
Acetone: Ethanol for 2 h, then spun down. The resulting supernatant is then collected for metabolomic analysis while the precipitate is used for proteomics analysis. SPOT applied to metabolomics is best utilized for untargeted analysis. 50
microliters of supernatant extracted from SPOT sample preparation were analyzed
through either reverse phase LC or HILIC in a global untargeted analysis with
simultaneous analysis of molecular fragmentation. This approach showed reproducible results comparable to traditional metabolomic methods and is efficient
with the ability to take cells from pellets to desalted samples ready for MS analysis
within 9 h. Additionally, this method led to the extraction of changing metabolites
key for biological information. SPOT was applied to human acute promyelocytic
leukemia (HL-60) cells that were exposed to zinc intoxication. Additionally, data
was collected at various time points throughout the analysis from 6 h to 24 h.
This investigation highlighted three pathways that appeared significantly modified with zinc treatment: tryptophan metabolism, purine metabolism, and eicosanoid signaling. Metabolomics allowed for the discovery of cellular responses not
found with proteomics and transcriptomics on the same sample. These pathways
were previously identified with genomic technologies and are supported by these
metabolomic data sets extracted using SPOT. The continued use of the SPOT protocol will answer many biological questions, through the incorporation of highthroughput, time-resolved, large-scale data sets for untargeted multi-omics analysis.
4.6.2 Metabolomic Analysis with Imaging Mass Spectrometry
IMS has been used with high success for metabolomic analysis. Although LC-MS
is more suitable for absolute quantitation, IMS maintains the spatial information
from a tissue section. A paper from 2018 utilized IMS, in coordination with immunohistochemistry, qPCR, western blotting and enzyme assays, to elucidate the
E. S. Rivera et al.
4.6.1 Combining Metabolomics with Other Omics Technolgies
LC-MS has been successfully utilized for metabolomics, but previous sample preparation methods made metabolomics incompatible with proteomic and lipidomic
analysis. In order to maximize the data extracted from a sample, a new method
known as sample preparation for multi-omics technologies (SPOT) has been developed for high-throughput multi-omics analysis by various collaborations at
Vanderbilt University [25, 91]. This technology allows for proteomic, transcriptomic, and metabolomic analysis from the same sample, with common sample preparation methodology. This common preparation allows for high-throughput sample
analysis, which would be optimal for applications such as rapid threat assessment.
This LC-MS based method allows for temporally resolved data sets in addition to
multi-omics analyses, optimal for addressing complex bio-logical questions.
This novel multi-omics sample preparation method utilizes cells but can be
applied to tissue samples as well. Cells are lysed, undergo a freeze-thaw cycle, and
then are sonicated in an ice bath. Aliquots are then lysed and precipitated with 75:25
Acetone: Ethanol for 2 h, then spun down. The resulting supernatant is then collected for metabolomic analysis while the precipitate is used for proteomics analysis. SPOT applied to metabolomics is best utilized for untargeted analysis. 50
microliters of supernatant extracted from SPOT sample preparation were analyzed
through either reverse phase LC or HILIC in a global untargeted analysis with
simultaneous analysis of molecular fragmentation. This approach showed reproducible results comparable to traditional metabolomic methods and is efficient
with the ability to take cells from pellets to desalted samples ready for MS analysis
within 9 h. Additionally, this method led to the extraction of changing metabolites
key for biological information. SPOT was applied to human acute promyelocytic
leukemia (HL-60) cells that were exposed to zinc intoxication. Additionally, data
was collected at various time points throughout the analysis from 6 h to 24 h.
This investigation highlighted three pathways that appeared significantly modified with zinc treatment: tryptophan metabolism, purine metabolism, and eicosanoid signaling. Metabolomics allowed for the discovery of cellular responses not
found with proteomics and transcriptomics on the same sample. These pathways
were previously identified with genomic technologies and are supported by these
metabolomic data sets extracted using SPOT. The continued use of the SPOT protocol will answer many biological questions, through the incorporation of highthroughput, time-resolved, large-scale data sets for untargeted multi-omics analysis.
4.6.2 Metabolomic Analysis with Imaging Mass Spectrometry
IMS has been used with high success for metabolomic analysis. Although LC-MS
is more suitable for absolute quantitation, IMS maintains the spatial information
from a tissue section. A paper from 2018 utilized IMS, in coordination with immunohistochemistry, qPCR, western blotting and enzyme assays, to elucidate the
E. S. Rivera et al.