Biopreservation and Biobanking最新文献

Aim of the Survey: When it comes to collaboration between academic biobanks and the pharmaceutical/biotechnology industry, the criteria for effective collaborations are still unclear. Researchers in industry and academic biobanks can have different incentives and requirements that the other party is often not familiar with. This survey was conducted in an attempt to increase understanding of these fundamental knowledge gaps that may be obstacles to optimal collaboration between academia and industry. Key Findings from the Survey: There were 53 total respondents. Although this was a global survey, most respondents (n = 29) were from North America, likely reflecting overall investment in research in this region and possibly increased interactions between academia and industry as well. Most respondent academic biobanks collect multiple sample types with most (>90%) collecting both biofluids (including blood) and tissue. Most of the participating academic biobanks were aware that they were not (35%), or only partially (35%), using the full potential of their inventory. One option for increasing utilization rates is by collaborating with industry partners. The main issues when working with industry were perceived to be a combination of challenges including contractual (55%), consent restrictions (45%), timelines (41%), or time pressure (36%). Time taken to put agreements together was also a significant hurdle (54%), together with the industry's administrative requirements (36%). Brief Conclusions from the Survey: To take advantage of opportunities for joint collaboration, it is essential that the parties involved build trust. The first step is to understand the different requirements and needs of the other party and to establish efficient structures for joint cooperation. This survey has highlighted key areas to be addressed as the next steps for strengthening bonds between academic biobanks and industry partners.

Background: The population of blast cells among peripheral blood mononuclear cells (PBMCs) obtained from patients is a desirable specimen for analyzing gene expression in diseases including acute myeloid leukemia. Although the enrichment of blast cells often needs to be performed at a central laboratory, acceptable conditions for sample transport from clinical sites remain to be established. Methods: We evaluated storage temperature, duration, and tube type before initiating sample processing for the analysis of cluster of differentiation (CD)33⁺ myeloid cells among PBMCs as an alternative to CD34⁺/CD33⁺ blast cells. Results: CD33⁺ myeloid cells were successfully purified by MACS. The cell viability and the RNA integrity were sustained during storage up to 48 hours before sample processing. Storage at 4°C had minimal effects on gene expression, whereas storage at room temperature induced the senescence pathway, characterized by the expression of stress-inducible genes. A CPT tube was also better than an ethylenediaminetetraacetic acid tube for minimizing gene expression change. Conclusions: Our study provided important clues for establishing a sample handling approach for gene expression analysis with purified cell fractions from human PBMCs. To keep the variation of gene expression to a minimum, samples should be delivered at 4°C within 48 hours before processing.

Multipotent mesenchymal stromal/stem cell (MSC) spheroids generated in three-dimensional culture are of considerable interest as a novel therapeutic tool for regenerative medicine. However, the lack of reliable methods for storing MSC spheroids represents a significant roadblock to their successful use in the clinic. An ideal storage medium for MSC spheroids should function as both a vehicle for delivery and a cryoprotectant during storage of spheroids for use at a later time. In this study, we compared the outcomes after subjecting MSC spheroids to a freeze/thaw cycle in three Good Manufacturing Practices-grade cryopreservation media, CryoStor10 (CS10), Stem-Cellbanker (SCB), and Recovery Cell Culture Freezing Media (RFM) or conventional freezing medium (CM) (CM, Dulbecco's modified Eagle's medium containing 20% fetal bovine serum and 10% dimethyl sulfoxide) as a control for 2 months. The endpoints tested were viability, morphology, and expression of stem cell markers and other relevant genes. The results of LIVE/DEAD™ assays and annexin V/propidium iodide staining suggested that viability was relatively higher after one freeze/thaw cycle in CS10 or SCB than after freeze/thaw in CM or RFM. Furthermore, the relative "stemness" and expression of MSC markers were similar with or without freeze/thaw in CS10. Scanning electron microscopy also indicated that the surface morphology of MSC spheroids was well preserved after cryopreservation in CS10. Thus, even though it was tested for a short-term period, we suggest that CS10, which has been approved for clinical use by the U.S. Food and Drug Association, is a promising cryopreservation medium that would facilitate the development of MSC spheroids for future clinical use.

Background: Blood transfusion has advanced toward component therapy for specific requirements during trauma and surgery. Oxidative stress is induced in erythrocytes during storage. Hence, antioxidants as additives can be employed to counteract oxidative stress and enhance antioxidant defenses. Therefore, this study investigates the combinatorial effects of vitamin C and l-carnitine on erythrocytes during storage. Methodology: Erythrocyte samples were categorized into control and experimental groups-vitamin C (10 mM) and l-carnitine (10 mM) and stored under blood bank conditions (at 4°C) for 35 days. Hemoglobin (Hb), antioxidant enzymes (superoxide dismutase [SOD], catalase [CAT] and glutathione peroxidase [GPX]), lipid peroxidation products (conjugate dienes and thiobarbituric acid reactive substances [TBARSs]), protein oxidation products, metabolic markers (glucose, lactate dehydrogenase), glutathione (GSH), superoxides, and hemolysis were assessed at weekly intervals. Results: SOD activity increased on day 7 in the controls, whereas it increased on days 7 and 14 in the experimental groups. CAT activity increased on day 35 in both the groups. GPX activity increased on day 7 in the controls. Hb levels decreased on days 14 and 35 in the controls and on day 35 in the experimental groups. Hemolysis increased from day 7 onward in both the groups. Protein oxidation products were maintained throughout the storage. GSH levels increased on day 21 in the controls and on days 14 and 21 in the experimental groups. Superoxides and conjugate dienes decreased from day 14 in both the groups. TBARSs decreased on day 7 in the experimental groups. Conclusion: Vitamin C and l-carnitine have synergistically enhanced the efficacy of stored erythrocytes in terms of Hb, antioxidant enzymes, and lipid peroxidation.

Oxidative stress is one of the main causes of loss of sperm function during chilled storage. The aim of the current study was to evaluate the effects of a fructose-based extender, which was supplemented with catalase or uric acid, on the motility, viability, morphological integrity, and lipid peroxidation (LPO) of Colossoma macropomum spermatozoa. Sperm was diluted in extenders containing catalase (0; 0.1; 0.8; and 1.5 kU/L) or uric acid (0; 0.25; 0.5; and 1.0 mmol/L) and then stored at 4.3 ± 0.6°C for 96 hours. The chilling storage time had more significant and pronounced effects on practically all the measured sperm quality parameters than the different concentrations of both antioxidants added to the extenders. This was true for sperm motility, motility duration, sperm viability, and the percentage of normal spermatozoa. In fact, for all these parameters, values were higher in the extenders supplemented with catalase or uric acid, than those not supplemented with these antioxidants, especially after 96 hours. The LPO process showed an antioxidant-dependent response. In catalase-supplemented extenders thiobarbituric acid reactive substance (TBARS) levels increased gradually and significantly with time, but remained stable during the 96 hours of chilled storage in all samples in which uric acid was added. Despite this, TBARS levels were lower in the extenders supplemented with both catalase and uric acid than in those not having these antioxidants. Inverse correlations were found between sperm motility and the damage in sperm flagella. Our findings suggest that the supplementation of an extender with catalase or uric acid is beneficial and protects fish sperm membranes from damage caused by oxidative stress during low-temperature storage. The extenders containing 0.1 kU/L of catalase and 0.25 mmol/L of uric acid provided effective antioxidant protection for the spermatozoa of this important Amazonian fish.

A systematic review was performed to summarize the scientific evidence and critically evaluate the effects of cryopreservation on sperm morphology in freshwater fish, and to assess the methodologies for sperm morphology classification. The search strategy was applied to four electronic databases (CAB Direct, Pub Med, Scopus, and ISI Web of Science). The main inclusion criteria involved studies on semen from freshwater fish subjected to the cryopreservation process and evaluation of sperm quality through morphology. The risk of bias was assessed with respect to randomization, allocation concealment, blinding, incomplete outcome data, and selective reporting. A total of 6 publications reporting sperm cryopreservation from 4 species with a total 74 fish individuals were included in this review. A high methodological variability among the results of the studies was observed due to the species-specific protocols and diversity of freshwater fish species studied. All included studies reported negative effects of cryopreservation on sperm quality, especially morphology, highlighting the increase in incidence of sperm abnormalities. However, only five studies statistically compared abnormalities between groups (fresh and cryopreserved sperm). Our results suggest the need to elaborate on a new morphological classification of fish spermatozoa, by considering the structure and physiology of fish sperm. This classification should be developed based on the sperm characterization and observing damage caused by different cryopreservation protocols.

Vitrification of reproductive cells is definitely essential and integral in animal breeding, as well as in assisted reproduction. However, issues accompanied with this technology such as decreased oocyte competency and relatively low embryo survival rates appear to be a tough conundrum that has long perplexed us. As significant organelles in cell metabolism, mitochondria play pivotal roles in numerous pathways. Nonetheless, extensive evidence has demonstrated that vitrification can seriously impair mitochondrial function in mammalian oocytes. Thus, in this article, we summarize the current progress in oocyte vitrification and particularly outline the common mitochondrial abnormalities alongside subsequent injury cascades seen in mammalian oocytes following vitrification. Based on existing literature, we tentatively come up with the potential mechanisms related to mitochondrial dysfunction and generalize efficacious ways which have been recommended to restore mitochondrial function.

Objective: Amniotic fluid (AF) plays a crucial role in diagnosing and predicting perinatal diseases, specifically preeclampsia (PE). Adequate preservation of AF samples is essential for advancing the development of PE-related biomarkers and understanding the disease's mechanisms. Materials and Methods: This study presents a method for preserving proteins in AF on a solid medium, specifically a nitrocellulose membrane, which is referred to as an AF membrane. Samples were collected from normotensive subjects and PE patients and treated with direct freezing and the AF membrane methods, respectively. Protein quality was assessed through sodium dodecyl sulfate-page and capillary electrophoresis. Liquid chromatography tandem mass spectrometry (LC-MS/MS) with data-independent acquisition was employed for proteomic analysis. Bioinformatics analysis identified differentially expressed proteins and pathways distinguishing normotensive subjects from PE patients. Results: Comparison of the AF membrane method to the direct freezing method showed no significant impact on the protein content in the AF. The preservation methods employed did not result in evident protein differences or degradation in the AF obtained from both normotensive subjects and PE patients. Analysis based on Gene Ontology and HALLMARK gene sets revealed the upregulation of pathways associated with angiotensin, reactive oxygen species, and coagulation in PE patients. Furthermore, several biomarkers previously reported to be increased in PE serum, namely ENG, ERN1, FLT1, GDF15, HSPA5, LGALS3, PAPPA, PTX3, and SERPINE1, were significantly elevated in the AF. Conclusion: The AF membrane method proved to be highly effective, reliable, and durable for preserving proteins in AF samples. Preserving AF samples in a solid state holds significant value in discovering novel protein biomarkers and investigating the underlying mechanisms of PE.

Many molecular approaches have been employed for the quality control (QC) of biobanked DNA samples. Since 2003, the National Biobank of Korea (NBK) has provided various studies with over half a million quality-controlled genomic DNA samples using conventional agarose gel electrophoresis and spectrophotometry. We assessed the postanalytical genomic data quality of DNA samples (n = 41) with a different range of the DNA quality index such as genomic quality number (GQN) for developing an evidence-based best practice for DNA quality criteria. We examined the quality indices of three different platforms, including single nucleotide polymorphism arrays, methylation arrays, and next-generation sequencing, using the same DNA samples (n = 41) of different quality, ranging from 4.0 to 10.0 values of the GQN. Our data analysis revealed that higher GQN value and/or double-stranded DNA concentration resulted in higher quality genomic data. In addition, all the analyzed DNA samples successfully generated good-quality genomic data. This study provides a guide for the QC of biobanked DNA samples for genomic analysis platforms.