The effect of pre-analytical conditions on blood metabolomics in epidemiological studies

AbstractBackgroundSerum and plasma are commonly used biofluids for large-scale metabolomic-epidemiology studies. Their metabolomic profile is susceptible to changes due to variability in pre-analytical conditions and the impact of this is unclear.MethodsParticipant-matched EDTA-plasma and serum samples were collected from 37 non-fasting volunteers and profiled using a targeted nuclear magnetic resonance (NMR) metabolomics platform (N=151 traits). Metabolic concentrations were compared between reference (pre-storage: 4°C, 1.5h; post-storage: no sample preparation or NMR-analysis delays) and four, pre-storage, blood processing conditions, where samples were incubated at (i) 4°C, 24h; (ii) 4°C, 48h; (iii) 21°C, 24h; (iv) 21°C, 48h, before centrifugation; and two, post-storage, sample processing conditions in which samples (i) thawed overnight, then left for 24h before addition of sodium buffer followed by immediate NMR analysis; (ii) thawed overnight, addition of sodium buffer, then left for 24h before profiling. Linear regression models with random-intercepts were used to assess the impact of these six pre-analytical conditions on EDTA-plasma/serum metabolome.ResultsFatty acids, beta-hydroxybutyrate, glycoprotein-acetyls and most lipid-related traits, in serum and plasma, were robust to the tested pre and post-storage conditions. Pre-storage conditions impacted concentrations of glycolysis metabolites, acetate, albumin and amino-acids by levels that could potentially bias research results (up to 1.4SD difference compared with reference). Post-storage conditions affected histidine, phenylalanine and LDL-particle-size, with differences up to 1.4SD.ConclusionsMost metabolic traits are robust to the pre- and post-storage conditions tested here and that may commonly occur in large-scale cohorts. However, concentrations of glycolysis metabolites, and amino-acids may be compromised.Key messagesIn large scale epidemiological studies, blood processing delays, incubation at high temperature prior to long term storage, and NMR profiling delays after long term storage, may occur.Concentrations of fatty acids, beta-hydroxybutyrate, glycoprotein acetyls and most lipid-related traits are robust to variations in pre-storage temperature and duration of incubation (4°C or 21°C for up to 48h prior to centrifugation) and post-storage sample handling (24h delay in sample preparation or NMR profiling).Glycolytic metabolite concentrations are altered by pre-storage conditions and amino-acids, particularly histidine and phenylalanine, by both, pre and post-storage conditions.

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Long-term storage conditions for carriers with denitrifying biomass of the fluidized, methanol-fed denitrification reactor of the Montreal Biodome, and the impact on denitrifying activity and bacterial population

Water Research ◽

10.1016/j.watres.2006.03.002 ◽

2006 ◽

Vol 40 (9) ◽

pp. 1836-1840 ◽

Cited By ~ 25

Author(s):

Véronique Laurin ◽

Normand Labbé ◽

Pierre Juteau ◽

Serge Parent ◽

Richard Villemur

Keyword(s):

Bacterial Population ◽

Storage Conditions ◽

Long Term Storage ◽

Denitrification Reactor ◽

The Impact ◽

Term Storage

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The Impact of Different Long-Term Storage Conditions on the Viability of Lichen-Forming Ascomycetes and their Green Algal Photobiont, Trebouxia spp.

Plant Biology ◽

10.1055/s-2003-40794 ◽

2003 ◽

Vol 5 (3) ◽

pp. 324-330 ◽

Cited By ~ 51

Author(s):

R. Honegger

Keyword(s):

Storage Conditions ◽

Long Term Storage ◽

Green Algal ◽

The Impact ◽

Term Storage

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A Study on the Combination of Enzyme Stabilizers and Low Temperatures in the Long-Term Storage of Glutamate Biosensor

Chemosensors ◽

10.3390/chemosensors9060129 ◽

2021 ◽

Vol 9 (6) ◽

pp. 129

Author(s):

Andrea Bacciu ◽

Paola Arrigo ◽

Rossana Migheli ◽

Alessandra T. Peana ◽

Gaia Rocchitta ◽

...

Keyword(s):

Enzymatic Activity ◽

Low Temperatures ◽

Triethylene Glycol ◽

Storage Conditions ◽

Low Temperature Storage ◽

Long Term Storage ◽

The Impact ◽

Term Storage

The importance of physiological glutamate has been widely demonstrated in cognitive and memory processes, as well as in neurotransmission. The involvement of physiological glutamate in several pathologies has also been established. Therefore, analytical devices for studying variations in physiological glutamate are of fundamental importance, particularly in preclinical studies. The necessary knowledge to develop and characterize biosensors for glutamate detection is often restricted to only a few research groups. However, many more groups have sought to implant such analytical devices to study the glutamatergic system in vivo. On this basis, a series of studies was undertaken to explore the medium-term storage of biosensors, thereby allowing their usage results to be differentiated from their construction and characterization processes to facilitate the wider diffusion and use of such sensors. Therefore, it has become vital to determine the best storage conditions to extend the life and functionality of these biosensors, especially due to the diachronic instability of the enzyme present on the surface. In the present study, we analyzed the impact of glycols, such as glycerol and triethylene glycol, as enzyme stabilizers coupled with long-term storage at low temperatures (−20 and −80 °C) on biosensor performance. The biosensors were observed for 5 months and evaluated for their enzymatic activity by measuring the VMAX(app) and KM(app). The analytical features were also evaluated in terms of the Linear Region Slope, which is one the most important parameters for indicating the efficiency and the sensitivity of biosensors. Interestingly, both glycols proved to be capable of increasing enzymatic activity and maintaining good biosensor efficiency over time. Moreover, the combination with low-temperature storage highlighted the different behaviors of the two glycols. In particular, glycerol was more effective in stabilizing the enzyme and maintaining analytical performance when the biosensors were stored at −20 °C. Instead, triethylene glycol performed the same function as glycerol but when the biosensors were stored at −80 °C.

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Long-Term Storage Considerations for Spacecraft Lubricants

Lubricants ◽

10.3390/lubricants8030032 ◽

2020 ◽

Vol 8 (3) ◽

pp. 32 ◽

Cited By ~ 2

Author(s):

Michael Buttery ◽

Simon Lewis ◽

Anthony Kent ◽

Rachel Bingley ◽

Matthew Cropper

Keyword(s):

Best Practice ◽

Experimental Studies ◽

Storage Conditions ◽

Tribological Performance ◽

Support Design ◽

Best Practice Guidelines ◽

Long Term Storage ◽

The Impact ◽

Term Storage

Spacecraft mechanisms commonly undergo extended periods of storage, either on-ground, or in-flight and there are an increasing number of missions for which some element of long-term storage may be required. Despite the obvious potential for degradation of lubricants during storage which might impact mechanism functionality or life and so even become mission-threatening, today’s understanding of storage phenomena is rather incomplete. This paper provides consolidation and review of recent experimental studies in this area and considers the range of storage conditions and associated degradation phenomena which could impact different lubricants. Whilst some storage best practice guidelines exist, experimental verification of the impact of storage phenomena has rarely been carried out and test data is rather scarce and incomplete. Given the absence of comprehensive data to support design, lubricant selection or the development of storage protocols, it is shown that for all lubricant types careful control of storage and test environments combined with monitoring of the evolving tribological performance during periodic mechanism exercising are presently the most effective storage risk mitigations.

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Co-encapsulation of vegetable oils with phenolic antioxidants and evaluation of their oxidative stability under long-term storage conditions

LWT ◽

10.1016/j.lwt.2021.111033 ◽

2021 ◽

Vol 142 ◽

pp. 111033

Author(s):

Lorine Le Priol ◽

Justine Gmur ◽

Aurélien Dagmey ◽

Sandrine Morandat ◽

Karim El Kirat ◽

...

Keyword(s):

Oxidative Stability ◽

Vegetable Oils ◽

Storage Conditions ◽

Phenolic Antioxidants ◽

Long Term Storage ◽

Term Storage

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Effects of Long-Term Storage on the Detection of Proteins, DNA, and mRNA in Tissue Microarray Slides

Journal of Histochemistry & Cytochemistry ◽

10.1369/0022155411423779 ◽

2011 ◽

Vol 59 (12) ◽

pp. 1113-1121 ◽

Cited By ~ 35

Author(s):

Christina Karlsson ◽

Mats G. Karlsson

Keyword(s):

Gene Copy Number ◽

Tissue Microarrays ◽

Storage Conditions ◽

Gene Copy ◽

Histological Techniques ◽

Long Term Storage ◽

Term Storage ◽

Formalin Fixed

Storage of tissue slides has been claimed to induce dramatically reduced antigen detection particularly for immunohistochemistry (IHC). With tissue microarrays, the necessity to serially cut blocks in order to obtain as much material as possible is obvious. The presumed adverse effect of storage might hamper such an approach. The authors designed an experimental setting consisting of four different storage conditions with storage time of tissue slides of up to 1 year. Detection of proteins, DNA, and mRNA was performed using IHC and in situ hybridization techniques. Slight but significant changes in IHC occurred over time. The most important factor is the primary antibody used: four showed no significant changes, whereas limited decreases in 8 antibodies could be detected by image analysis. Whether the antigen was nuclear or cytoplasmic/membranous did not matter. No major differences between different storage conditions could be shown, but storage at 4C was overall the best procedure. Furthermore, gene copy number aberrations, chromosomal translocations, and the presence of mRNA could be detected on slides stored up to 1 year. In conclusion, in tissues optimally formalin fixed and using modern histological techniques, only minute changes in tissue antigenicity are induced by long-term storage.

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Rapid, Reproducible, Quantifiable NMR Metabolomics: Methanol and Methanol: Chloroform Precipitation for Removal of Macromolecules in Serum and Whole Blood

Metabolites ◽

10.3390/metabo8040093 ◽

2018 ◽

Vol 8 (4) ◽

pp. 93 ◽

Cited By ~ 9

Author(s):

Cora McHugh ◽

Thomas Flott ◽

Casey Schooff ◽

Zyad Smiley ◽

Michael Puskarich ◽

...

Keyword(s):

Human Serum ◽

1H Nmr ◽

Whole Blood ◽

Nmr Spectra ◽

High Quality ◽

Healthy Human ◽

Long Term Storage ◽

The Impact ◽

Term Storage

Background: Though blood is an excellent biofluid for metabolomics, proteins and lipids present in blood can interfere with 1d-1H NMR spectra and disrupt quantification of metabolites. Here, we present effective macromolecule removal strategies for serum and whole blood (WB) samples. Methods: A variety of macromolecule removal strategies were compared in both WB and serum, along with tests of ultrafiltration alone and in combination with precipitation methods. Results: In healthy human serum, methanol:chloroform:water extraction with ultrafiltration was compared to methanol precipitation with and without ultrafiltration. Methods were tested in healthy pooled human serum, and in serum from patients with sepsis. Effects of long-term storage at −80 °C were tested to explore the impact of macromolecule removal strategy on serum from different conditions. In WB a variety of extraction strategies were tested in two types of WB (from pigs and baboons) to examine the impact of macromolecule removal strategies on different samples. Conclusions: In healthy human serum methanol precipitation of serum with ultrafiltration was superior, but was similar in recovery and variance to methanol:chloroform:water extraction with ultrafiltration in pooled serum from patients with sepsis. In WB, high quality, quantifiable spectra were obtained with the use of a methanol: chloroform precipitation.

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Is adipose tissue suitable for detection of (synthetic) cannabinoids? A comparative study analyzing antemortem and postmortem specimens following pulmonary administration of JWH-210, RCS-4, as well as ∆9-tetrahydrocannabinol to pigs

Archives of Toxicology ◽

10.1007/s00204-020-02843-x ◽

2020 ◽

Vol 94 (10) ◽

pp. 3421-3431

Author(s):

Nadine Schaefer ◽

Frederike Nordmeier ◽

Ann-Katrin Kröll ◽

Christina Körbel ◽

Matthias W. Laschke ◽

...

Keyword(s):

Adipose Tissue ◽

Synthetic Cannabinoids ◽

Standard Addition ◽

Storage Conditions ◽

Long Term Storage ◽

Low Concentrations ◽

Concentration Changes ◽

Term Storage ◽

Plateau Level

Abstract Examining fatal poisonings, chronic exposure may be reflected by the concentration in tissues known for long-term storage of drugs. Δ9-tetrahydrocannabinol (THC) persists in adipose tissue (AT), but sparse data on synthetic cannabinoids (SC) are available. Thus, a controlled pig study evaluating antemortem (AM) disposition and postmortem (PM) concentration changes of the SC 4-ethylnaphthalene-1-yl-(1-pentylindole-3-yl)methanone (JWH-210) and 2-(4-methoxyphenyl)-1-(1-pentyl-indole-3-yl)methanone (RCS-4) as well as THC in AT was performed. The drugs were administered pulmonarily (200 µg/kg body weight) to twelve pigs. Subcutaneous (s.c.) AT specimens were collected after 15 and 30 min and then hourly up to 8 h. At the end, pigs were sacrificed and s.c., perirenal, and dorsal AT specimens were collected. The carcasses were stored at room temperature (RT; n = 6) or 4 °C (n = 6) and specimens were collected after 24, 48, and 72 h. After homogenization in acetonitrile and standard addition, LC–MS/MS was performed. Maximum concentrations were reached 0.5–2 h after administration amounting to 21 ± 13 ng/g (JWH-210), 24 ± 13 ng/g (RCS-4), and 22 ± 20 ng/g (THC) and stayed at a plateau level. Regarding the metabolites, very low concentrations of N-hydroxypentyl-RCS-4 (HO-RCS-4) were detected from 0.5 to 8 h. PM concentrations of parent compounds did not change significantly (p > 0.05) over time under both storage conditions. Concentrations of HO-RCS-4 significantly (p < 0.05) increased in perirenal AT during storage at RT. These results suggest a rapid distribution and persistence in s.c. AT. Furthermore, AT might be resistant to PM redistribution of parent compounds. However, significant PM increases of metabolite concentrations might be considered in perirenal AT.

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3′ MACE RNA-sequencing allows for transcriptome profiling in human tissue samples after long-term storage

Laboratory Investigation ◽

10.1038/s41374-020-0446-z ◽

2020 ◽

Vol 100 (10) ◽

pp. 1345-1355 ◽

Cited By ~ 2

Author(s):

Stefaniya Boneva ◽

Anja Schlecht ◽

Daniel Böhringer ◽

Hans Mittelviefhaus ◽

Thomas Reinhard ◽

...

Keyword(s):

Rna Sequencing ◽

Storage Time ◽

Transcriptome Profiling ◽

Rna Seq ◽

Fresh Tissue ◽

Long Term Storage ◽

Ffpe Samples ◽

The Impact ◽

Term Storage

Abstract This study aims to compare the potential of standard RNA-sequencing (RNA-Seq) and 3′ massive analysis of c-DNA ends (MACE) RNA-sequencing for the analysis of fresh tissue and describes transcriptome profiling of formalin-fixed paraffin-embedded (FFPE) archival human samples by MACE. To compare MACE to standard RNA-Seq on fresh tissue, four healthy conjunctiva from four subjects were collected during vitreoretinal surgery, halved and immediately transferred to RNA lysis buffer without prior fixation and then processed for either standard RNA-Seq or MACE RNA-Seq analysis. To assess the impact of FFPE preparation on MACE, a third part was fixed in formalin and processed for paraffin embedding, and its transcriptional profile was compared with the unfixed specimens analyzed by MACE. To investigate the impact of FFPE storage time on MACE results, 24 FFPE-treated conjunctival samples from 24 patients were analyzed as well. Nineteen thousand six hundred fifty-nine transcribed genes were detected by both MACE and standard RNA-Seq on fresh tissue, while 3251 and 2213 transcripts were identified explicitly by MACE or RNA-Seq, respectively. Standard RNA-Seq tended to yield longer detected transcripts more often than MACE technology despite normalization, indicating that the MACE technology is less susceptible to a length bias. FFPE processing revealed negligible effects on MACE sequencing results. Several quality-control measurements showed that long-term storage in paraffin did not decrease the diversity of MACE libraries. We noted a nonlinear relation between storage time and the number of raw reads with an accelerated decrease within the first 1000 days in paraffin, while the numbers remained relatively stable in older samples. Interestingly, the number of transcribed genes detected was independent on FFPE storage time. RNA of sufficient quality and quantity can be extracted from FFPE samples to obtain comprehensive transcriptome profiling using MACE technology. We thus present MACE as a novel opportunity for utilizing FFPE samples stored in histological archives.

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