Biopreservation and Biobanking最新文献

Research studies aimed at advancing cancer prevention, diagnosis, and treatment depend on a number of key resources, including a ready supply of high-quality annotated biospecimens from diverse ethnic populations that can be used to test new drugs, assess the validity of prognostic biomarkers, and develop tailor-made therapies. In November 2011, KHCCBIO was established at the King Hussein Cancer Center (KHCC) with the support of Seventh Framework Programme (FP7) funding from the European Union (khccbio.khcc.jo). KHCCBIO was developed for the purpose of achieving an ISO accredited cancer biobank through the collection, processing, and preservation of high-quality, clinically annotated biospecimens from consenting cancer patients, making it the first cancer biobank of its kind in Jordan. The establishment of a state-of-the-art, standardized biospecimen repository of matched normal and lung tumor tissue, in addition to blood components such as serum, plasma, and white blood cells, was achieved through the support and experience of its European partners, Trinity College Dublin, Biostór Ireland, and accelopment AG. To date, KHCCBIO along with its partners, have worked closely in establishing an ISO Quality Management System (QMS) under which the biobank will operate. A Quality Policy Manual, Validation, and Training plan have been developed in addition to the development of standard operating procedures (SOPs) for consenting policies on ethical issues, data privacy, confidentiality, and biobanking bylaws. SOPs have also been drafted according to best international practices and implemented for the donation, procurement, processing, testing, preservation, storage, and distribution of tissues and blood samples from lung cancer patients, which will form the basis for the procurement of other cancer types. KHCCBIO will be the first ISO accredited cancer biobank from a diverse ethnic Middle Eastern and North African population. It will provide a unique and valuable resource of high-quality human biospecimens and anonymized clinicopathological data to the cancer research communities world-wide.

Biobanks of fresh, unfixed human normal and malignant tissues represent a valuable source for gene expression analysis in translational cancer research and molecular pathology. However, the success of molecular and cellular analysis in both clinical and translational research is strongly dependent on the collection, handling, storage, and quality control of fresh human tissue samples. The aim of this study was to evaluate an innovative vacuum-based refrigerated system, as a logistically feasible technology to increase the collection of tissue specimens, preserving the integrity of cellular and molecular components. We tested randomly-selected tissues stored under vacuum at 4°C by using endpoints important for research and diagnosis, including tissue morphology, epitope stability, and RNA integrity. Gene expression was evaluated by qualitative and quantitative RT analysis of selected housekeeping and tissue-specific genes. Tissue morphology and overall protein stability were generally well preserved, being compromised only in gallbladder tissue. By contrast, phosphoprotein and RNA analysis demonstrated a time-dependent degree of degradation, with progressive loss of stability from 24 to 72 hours. However, this reduction in RNA quality did not represent a limitation for successful expression analysis of selected genes. Indeed, a comparative qualitative and quantitative RT-PCR analysis showed that RNA extracted from tissues stored under vacuum is suitable for gene expression profiling, but requires highly sensitive technologies, such as quantitative RT-PCR. These data suggest that the refrigerated vacuum-based system represents a suitable and feasible technology for routine transport of fresh specimens from surgery to biobanks, thus increasing the opportunity to collect biospecimens.

A critical issue in defining protocols for biobanking practices is the preservation of total RNA for assessing the whole transcriptome and ensuring that it can be utilized in clinically oriented studies. Storage conditions, such as temperature and the length of time that tissues and purified RNA stay frozen, may directly impact RNA preservation. In this study, we evaluated a) the quality of RNA (as measured by RNA Integrity Number) purified from head and neck tumor tissues stored at -140°C for distinct time intervals of up to 7 years, and b) the quality of their respective RNAs stored for 4 years at -80°C when diluted at either 250 ng/μL or 25 ng/μL, with repeated freezing and thawing. Additionally, we generated a profile of the RNA collection of human tumors from different body sites stored at the AC Camargo Biobank. Our results showed no significant change in RIN values according to length of storage at -140°C. With respect to RNA aliquots stored at -80°C, RNA integrity at 250 ng/μL was preserved, while statistically significant degradation was observed at 25 ng/μL after only 8 months of storage. The RNA collection from most of the human tumors stored at the AC Camargo Biobank exhibited high quality, with average RIN around seven. However, ovary and stomach samples had the greatest RNA degradation. Taken together, the results show that both the temperature of preservation and the concentration of RNA should be strictly controlled by the biobank staff involved in macromolecule purification. Moreover, the RNAs from our biobank can be useful for the most demanding methods of gene expression analysis by virtue of adherence to optimal standard operating procedures for both tissue and macromolecule laboratories.

Background: The preanalytical phase is considered the most vulnerable phase in biopreservation, biobanking, and laboratory diagnostics. Accurate mixing after blood collection is claimed to be important and recommended by the manufacturers.

Objective: To evaluate whether it is really necessary to mix the primary blood tubes immediately after blood collection by means of evacuated tube systems.

Material and methods: Blood from 300 outpatients was equally and randomly divided into three groups: G1, sodium citrate vacuum tubes; G2, lithium heparin vacuum tubes; and G3, K2EDTA vacuum tubes. All vacuum tubes were processed using three different procedures. Procedure 1: Gold Standard (P1): All specimens mixed gently and carefully by inverting five times as recommended; Procedure 2: Rest time (P2): All specimens remained 5 min in the upright position, followed by gentle careful mixing by inverting five times; Procedure 3: No mix (P3): All specimens were left in upright position without mixing afterwards. The influence of the primary mixing tube procedure was evaluated for clinical chemistry, hematology, and coagulation parameters by paired t-test. The bias from the mixing procedure was also compared with quality specifications derived from biological variation.

Results: Significant differences (p<0.017) were found for: i) red blood cell count and hematocrit when P1 was compared with P2; ii) alanine aminotransferase and erythrocyte sedimentation rate when P1 was compared with P3; iii) red blood cell count, hematocrit, and hemolysis index when P2 was compared with P3. Surprisingly, clinically significant differences were found only for sodium when P1 was compared with P2, and P1 was compared with P3. No fibrin filaments or microclots were observed in any samples.

Conclusion: Primary blood tubes mixing after collection with evacuated tube system appears to be unnecessary.

The primary responsibility of biobanks is to collect biospecimens that are true reflections of the local population, thereby promoting translational research that is applicable to the community. The Swedish Cervical Cytology Biobank (SCCB) was designed as a hospital-integrated biobank in 2011. The SCCB has now been implemented in 10 county councils scattered across the country. It is headquartered at Karolinska University Hospital in Stockholm. The SCCB now processes more than 60% of the liquid-based gynecological cell samples obtained throughout Sweden. To improve the productivity of health care and research that rely on SCCB samples, a high level validation of the biobank system according to the principles of Good Laboratory Practices (GLP) is required. The performance of an entire high-throughput system validated by measuring the cell yield proved unsatisfactory after 1 year of sample collection and aliquoting. However, the results led to a number of high quality technical interventions for workflow enhancement. Subsequently, the improved process was applied to the system and led to a significant increase in cell yield. After the integration of the improved high quality methodology into the SCCB, the biobank services progressed more rapidly to serve the needs of personalized medicine and clinical studies. This enhancement was mainly due to the increased ability of the biobank to provide samples to research groups without any risk of leaving insufficient sample volumes for the care of the donor.

Biological materials collected in harsh environments such as archaeological excavations, at crime scenes, after mass disasters, in museums, or non-invasively in the field constitute a highly valuable source of genetic information. However, poor quality and limited quantity of the DNA extracted from these samples can be extremely challenging during further analyses. Here we have reviewed how degradation, decomposition, and contamination can affect DNA analysis, and how correct sample collection and storage methods will ensure the best possible conditions for further genetic analysis. Furthermore, highly efficient protocols for collection, decontamination, and extraction of DNA from minute amounts of biological material are presented.

Despite marked developments in the field of cryopreservation of cells and tissues for research and therapeutic applications, post-thaw cell death remains a significant drawback faced by cryobiologists. Post cryopreservation apoptosis and necrosis are normally observed within 6 to 24 h after post-thaw culture. As a result, massive loss of cell viability and cellular function occur due to cryopreservation. However, in this new generation of cryopreservation science, scientists in this field are focusing on incorporation of apoptosis and necrosis inhibitors (zVAD-fmk, p38 MAPK inhibitor, ROCK inhibitor, etc.) to cryopreservation and post-thaw culture media. These inhibitors target and inhibit various proteins such as caspases, proteases, and kinases, involved in the cell death cascade, resulting in reduced intensity of apoptosis and necrosis in the cryopreserved cells and tissues, increased cell viability, and maintenance of cellular function; thus improved overall cryopreservation efficiency is achieved. The present article provides an overview of various cell death pathways, molecules mediating cryopreservation-induced apoptosis and the potential of certain molecules in targeting cryopreservation-induced delayed-onset cell death.