Impact of Prolonged Blood Incubation and Extended Serum Storage at Room Temperature on the Human Serum Metabolome.
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ABSTRACT:
SUBMITTER: Kamlage B
PROVIDER: S-EPMC5875996 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BackgroundBlood cultures are one of the most important tests performed by microbiology laboratories. Many hospitals, particularly in low and middle-income countries, lack either microbiology services or staff to provide 24 h services resulting in delays to blood culture incubation. There is insufficient guidance on how to transport/store blood cultures if delays before incubation are unavoidable, particularly if ambient temperatures are high. This study set out to address this knowledge gap.MethodsIn three South East Asian countries, four different blood culture systems (two manual and two automated) were used to test blood cultures spiked with five common bacterial pathogens. Prior to incubation the spiked blood culture bottles were stored at different temperatures (25 °C, in a cool-box at ambient temperature, or at 40 °C) for different lengths of time (0 h, 6 h, 12 h or 24 h). The impacts of these different storage conditions on positive blood culture yield and on time to positivity were examined.ResultsThere was no significant loss in yield when blood cultures were stored < 24 h at 25 °C, however, storage for 24 h at 40 °C decreased yields and longer storage times increased times to detection.ConclusionBlood cultures should be incubated with minimal delay to maximize pathogen recovery and timely result reporting, however, this study provides some reassurance that unavoidable delays can be managed to minimize negative impacts. If delays to incubation ≥ 12 h are unavoidable, transportation at a temperature not exceeding 25 °C, and blind sub-cultures prior to incubation should be considered.
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Project description:Cellular metabolomics provides insights into the metabolic processes occurring within cells and can help researchers understand how these processes are regulated and how they relate to cellular function, health, and disease. In this technical note, we investigated the effects of solvent evaporation equipment and storage condition on high-coverage cellular metabolomics. We previously introduced a robust CIL LC-MS-based cellular metabolomics workflow that encompasses various steps, including cell harvest, metabolic quenching, cell lysis, metabolite extraction, differential chemical isotope labeling, and LC-MS analysis. This workflow has consistently served as the cornerstone of our collaborative research and service projects. As a core facility catering to users with diverse research needs and financial resources, we have encountered scenarios requiring short-term sample storage. For example, the need often arises to transport samples at room temperature from user sites to our core facility. Herein, we present a study in which we compared different solvent evaporation methods (specifically, the nitrogen blowdown evaporator, SpeedVac concentrator, and lyophilizer) and diverse storage conditions (including dried samples stored in a freezer, samples stored in a freezer with methanol, dried samples stored at room temperature, and samples stored at room temperature with methanol). Our findings indicate that the choice of solvent evaporation equipment did not significantly impact the cellular metabolome. However, we observed a noteworthy change in the metabolome after 7 days of storage when cells were stored with methanol, regardless of whether they were kept at -80 °C or room temperature, in contrast to cells that were dried and frozen. Importantly, we detected no significant alterations in cells that were dried and stored at room temperature. In conclusion, to ensure the production of high-quality CIL LC-MS metabolomics results, we strongly recommend that, in situations where low-temperature storage is not feasible, cell samples should be thoroughly dried before storage or shipment at room temperature.
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Project description:Plasma-derived microRNAs (miRNAs) are being used as biomarkers, and have been associated with human liver disease and function including fibrosis, inflammation, and drug-induced liver injury. They may also be biomarkers of the drug metabolism function of the liver. In order for plasma miRNA to function as a clinical biomarker, predictable variability is necessary during processing from whole blood to plasma. The current study evaluated the variability of miRNA in whole blood stored for 0.5, 1, 2, 4, 8, and 12 hours following the blood draw under clinical conditions (room temperature) prior to the separation of the plasma. Four healthy volunteers were recruited. Blood from all subjects was collected twice. MicroRNA-16 (miR-16) and miR-223 were evaluated because many studies have shown them to be reliably present in plasma and useful for normalization. miRNA concentrations were measured by real-time polymerase chain reaction. The coefficient of variability of the cycle threshold values for subjects for miR-223 and miR-16 ranged from ?3.6 to 6.8% and ?1.48 to 4.1%, respectively, over the 12-hour incubation. A second blood collection was performed to determine interday variability. The coefficient of variance from the initial blood draw compared with the final blood draw for each subject ranged from 0.42 to 7.9% for miR-16 and 1.7 to 8.3% for miR-223, indicating that these miRNAs have limited interday variability. We conclude that plasma miR-16 or miR-223 concentrations are stable in whole blood at room temperature for up to 12 hours.
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Project description:BackgroundCool temperature egg storage prior to incubation is a common practice in the broiler industry; however, prolonged egg storage causes increased embryonic mortality and decreased hatchability and growth in surviving chicks. Exposing eggs to short periods of incubation during egg storage (SPIDES) reduces the adverse consequences of prolonged storage. SPIDES increases blastodermal cell viability by reducing apoptosis, though the counteracting mechanisms are unclear. To define the impact of prolonged storage and SPIDES, transcriptome analysis compared gene expression from blastoderms isolated from eggs exposed to the following treatments: control (CR, stored at 17 °C for 4 days), prolonged storage (NSR, stored at 17 °C for 21 days), SPIDES (SR, stored at 17 °C for 21 days with SPIDES), and incubated control (C2, stored at 17 °C for 4 days followed by incubation to HH (Hamburger-Hamilton) stage 2, used as the ideal standard development) (n = 3/group). Data analysis was performed using the CLC Genomics Workbench platform. Functional annotation was performed using DAVID and QIAGEN Ingenuity Pathway Analysis.ResultsIn total, 4726 DEGs (differentially expressed genes) were identified across all experimental group comparisons (q < 0.05, FPKM> 20, |fold change| > 1.5). DEGs common across experimental comparisons were involved in cellular homeostasis and cytoskeletal protein binding. The NSR group exhibited activation of ubiquitination, apoptotic, and cell senescence processes. The SR group showed activation of cell viability, division, and metabolic processes. Through comparison analysis, cellular respiration, tRNA charging, cell cycle control, and HMBG1 signaling pathways were significantly impacted by treatment and potential regulatory roles for ribosomal protein L23a (RPL23A) and MYC proto-oncogene, BHLH transcription factor (MYC) were identified.ConclusionsProlonged egg storage (NSR) resulted in enriched cell stress and death pathways; while SPIDES (SR) resulted in enriched basic cell and anti-apoptotic pathways. New insights into DNA repair mechanisms, RNA processing, shifts in metabolism, and chromatin dynamics in relation to egg storage treatment were obtained through this study. Although egg storage protocols have been examined through targeted gene expression approaches, this study provided a global view of the extensive molecular networks affected by prolonged storage and SPIDES and helped to identify potential upstream regulators for future experiments to optimize egg storage parameters.
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Project description:Advances in the "omics" field bring about the need for a high number of good quality samples. Many omics studies take advantage of biobanked samples to meet this need. Most of the laboratory errors occur in the pre-analytical phase. Therefore evidence-based standard operating procedures for the pre-analytical phase as well as markers to distinguish between 'good' and 'bad' quality samples taking into account the desired downstream analysis are urgently needed. We studied concentration changes of metabolites in serum samples due to pre-storage handling conditions as well as due to repeated freeze-thaw cycles. We collected fasting serum samples and subjected aliquots to up to four freeze-thaw cycles and to pre-storage handling delays of 12, 24 and 36 hours at room temperature (RT) and on wet and dry ice. For each treated aliquot, we quantified 127 metabolites through a targeted metabolomics approach. We found a clear signature of degradation in samples kept at RT. Storage on wet ice led to less pronounced concentration changes. 24 metabolites showed significant concentration changes at RT. In 22 of these, changes were already visible after only 12 hours of storage delay. Especially pronounced were increases in lysophosphatidylcholines and decreases in phosphatidylcholines. We showed that the ratio between the concentrations of these molecule classes could serve as a measure to distinguish between 'good' and 'bad' quality samples in our study. In contrast, we found quite stable metabolite concentrations during up to four freeze-thaw cycles. We concluded that pre-analytical RT handling of serum samples should be strictly avoided and serum samples should always be handled on wet ice or in cooling devices after centrifugation. Moreover, serum samples should be frozen at or below -80°C as soon as possible after centrifugation.
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Project description:Complex incubation strategies have evolved to solve the trade-off between parent survival and care for their eggs with often brief departures (recesses) that maximize egg survival, and infrequent extended recesses maximizing adult condition. Here we examined incubation behaviour of sanderlings (Calidris alba), a species that exhibits both biparental and uniparental incubation behaviour. During 11 breeding seasons in Greenland, we have quantified incubation variability with thermologgers placed in nests. We estimated the impact of environmental conditions and individual characteristics on the occurrence and the duration of recesses. We found that extended recesses are a unique feature of uniparentals, and their frequency and duration increased in colder temperatures. The relationship was mediated by body condition, with individuals in poor condition performing longer extended recesses in colder temperatures. This suggests that extended recesses may represent a shift towards self-maintenance at the expense of the egg care, allowing birds to continue incubating under unfavourable conditions. Our study illustrates how extended recesses may be a key breeding strategy to overcome high energetic costs associated with incubation. Quantifying such behavioural flexibility paves the way for tracking future behavioural responses of individuals in the face of changing environments.
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Project description:Red blood cells (RBCs) degrade progressively during the weeks of refrigerated storage. No universally accepted definition of 'fresh' or 'old' RBCs exists. While practices vary from country to country, preservative solutions permitting shelf life as long as 7 weeks have been licenced. Transfusion of stored RBCs, particularly those at the end of the approved shelf life, has been implicated in adverse clinical outcomes. The results of observational analyses, animal models and studies in volunteers have proved provocative, controversial and contradictory. A recently completed randomized controlled trial (RCT) in premature infants exemplifies the difficulties with moderately sized clinical studies. Several other RCTs are in progress. The effect of RBC storage may well vary according to the clinical setting. Resolution of the importance of the storage lesion may require large pragmatic clinical trials. In the meantime, institutions involved in blood collection and transfusion should explore strategies that assure blood availability, while limiting the use of the oldest RBCs currently approved by regulation.
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