what are pre analytical factors

The optimal thickness of sections in FFPE tissue for nucleic acid evaluation depends on the dimensions and cellularity of the tissues. The chorionic tissue should be collected in normal saline with 10 mM EDTA and separated from maternal tissue under a stereo-microscope or dissecting microscopeto evaluate contamination with endometrial decidual tissue of maternal origin (189, 190). Stability of miRNA in human urine supports its biomarker potential. To avoid dilution effect of nucleic acids of undesired cells, marking the area containing the target cells on the hematoxylin and eosin (H&E) slide and corresponding paraffin block is necessary. Comparative stabilities of quantitative human immunodeficiency virus RNA in plasma from samples collected in VACUTAINER CPT, VACUTAINER PPT, and standard VACUTAINER tubes. Urine miRNAs are more stable and nuclease resistant compared to large mRNA and snRNA (small nuclear RNA) (153), and other cellular RNAs (154, 155). Enriched white blood cell (WBC) layer provides a good source of nucleic acids for molecular assays. Knowledge gaps in understanding of pre-analytical factors are identified in the clinical years amongst undergraduate students due to lack of formal teaching modules on the pre-analytical phase. Care should be taken to maintain the hydration of fresh non-fixed tissues and to avoid drying by covering in gauze soaked with normal saline (46). HIV-1 drug resistance genotyping from dried blood spots stored for 1 year at 4 C. Choo JM, Leong LE, Rogers GB. After final centrifugation, the pellet can also be used for DNA isolation (126, 127). The transport media that have been developed for culture may not be suitable for molecular tests and the media developed for molecular tests are not suitable for culture. Carnoy's fixative preserves tissues better than methanol-acetone when examined in more than 3 months post-fixation period (22). Teschendorff AE, Yang Z, Wong A, Pipinikas CP, Jiao Y, Jones A, et al. For RNA analysis, the fixation time in NBF should be limited to 48 hours at 4C or 8 hours at ambient temperature (uncontrolled RT) and the blocks should be analyzed within 1 year (18). Advanced Search Browse by Analyte Browse by Pre-analytical Factor Search SOPs SOP Compendiums. Preanalytical and analytical factors affecting laboratory results Performing reliable procedures in sample processing will increase the accuracy and reproducibility of these diagnostic tests. mtDNA content is affected by the cell separation method used and the time between blood withdrawal and cell separation. official website and that any information you provide is encrypted There are variable paraffin waxes regarding quality and melting points (ranging from 47C to 64C) (31), which may cause confounding effects on nucleic acid extraction (14, 32). Bosschieter J, Bach S, Bijnsdorp IV, Segerink LI, Rurup WF, van Splunter AP, et al. Rainen L, Arbique JC, Asthana D, Earley MC, Geiszler RL, Krieg-Schneider F, et al. A comprehensive meta-analysis by Greytak et al. Effects of heparin on polymerase chain reaction for blood white cells. Godfrey TE, Kim S-H, Chavira M, Ruff DW, Warren RS, Gray JW, et al. While some studies have reported acceptable results in RNA quantity of BM samples taken in EDTA tubes and stored at RT for 48 hours, care should be taken in such instances. Most of the articles about pre-analytical studies in human samples were collected from the past 20-year interval. . These RNAs become resistant to RNase and consequently can contaminate DNA extraction products (29). Frati P, Fineschi V, Di Sanzo M, La Russa R, Scopetti M, Severi FM, et al. Williams DL, Gillis TP, Dupree WG. The alcohol-containing mouthwash specimens are optimal media for preventing bacterial growth on swabs (108). Impact of nasopharyngeal swab types on detection of Bordetella pertussis by PCR and culture. Stanta G, Schneider C. RNA extracted from paraffin-embedded human tissues is amenable to analysis by PCR amplification. The descriptive results of that review are presented as below and the summary is depicted in table 1. The specimen should be shipped on wet ice within one week for CMV evaluation (160). Amniotic fluid can be processed without culture after 15 weeks of gestation (46, 181). Analytical Performance of COVID-19 Detection Methods (RT-PCR Wright D, Manos M. Sample preparation from paraffin-embedded tissues. Preservation Methods Differ in Fecal Microbiome Stability, Affecting Suitability for Field Studies. Long-term preservation and storage of mycobacteria. A key second step is to establish a reliable and specific method to detect viral infection . In: Kottke-Marchant K, Davis B, editors. Ross et al. Subsequently, the samples should be air-dried for 1 hour and finally put in a storage packet and stored at RT (123). Gibellini D, De Crignis E, Re MC. Between 0.5 to 4 ml or 1/10 of the sample is usually taken for quantitative fluorescent PCR (QF-PCR); meanwhile, the remainder is preserved for karyotype analysis (191). Certain practices in specimen collection, transportation, and storage can affect the integrity of nucleic acids before analysis. An article published in a 2012 report by Bonini and his colleague supports these findings. http://creativecommons.org/licenses/by/3.0/, https://www.cdc.gov/cmv/clinical/lab-tests.html, http://cdc.gov/dpdx/diagnosticprocedures/stool/moleculardx.html, www.cdc.gov/mumps/lab/specimen-collect.html, www.cdc.gov/flu/professionals/diagnosis/molecular-assays.htm, up to 72h optimal, but possible up to 6 days, DNA (Neisseria gonorrhea, Chlamydia trachomatis). Schlesinger Y, Halle D, Eidelman A, Reich D, Dayan D, Rudensky B, et al. Cree IA, Deans Z, Ligtenberg MJ, Normanno N, Edsjo A, Rouleau E, et al. Pre-analytical Factors: Classification Pre-analytical Factor Value(s) Storage: Time at room temperature: 0 h 24 h 48 h 72 h 96 h: View more. Watanabe M, Hashida S, Yamamoto H, Matsubara T, Ohtsuka T, Suzawa K, et al. In this article, we have reviewed most of the important issues in sample handling from bed to bench before starting molecular tests, which can be used in diagnostic as well as research laboratories. Influence of pre-analytical procedures on genomic DNA integrity in blood samples: the SPIDIA experience. The common examples are HER2 amplification in breast cancer, EGFR mutations and ALK rearrangements in lung cancer, BRAF mutations in malignant melanoma, RAS mutations in colorectal cancer, and BCR/ABL1 in chronic myelogenous leukemia (4). They are stable for approximately one week (167, 168). An official website of the United States government. Nechvatal JM, Ram JL, Basson MD, Namprachan P, Niec SR, Badsha KZ, et al. The https:// ensures that you are connecting to the Stability study of cervical specimens collected by swab and stored dry followed by human papillomavirus DNA detection using the cobas 4800 test. Molecular Pathology and Cytogenetics Division, Pathology Department, Children's Medical Center, Tehran University of Medical Sciences, Tehran, Iran, 2 Lee SH, Erber WN, Porwit A, Tomonaga M, Peterson LC. There is controversy about the effect of heparin on the molecular assays. Automation through pre-analytical robotic workstations, specimen labelers, specimen management systems, and automated phlebotomy tray preparation can significantly reduce the rate of errors that are due to active human factors. Optimal Fixation Conditions and DNA Extraction Methods for MLPA Analysis on FFPE Tissue-Derived DNA. RNA progressive loss at 37C is remarkable after two months (128). The storage time is reduced to few weeks at -20C (141, 145). Foss RD, Guha-Thakurta N, Conran RM, Gutman P. Effects of fixative and fixation time on the extraction and polymerase chain reaction amplification of RNA from paraffin-embedded tissue Comparison of two housekeeping gene mRNA controls. Diagnosis of mycobacterial infections by nucleic acid amplification: 18-month prospective study. Noncryogenic preservation of mammalian tissues for DNA extraction: an assessment of storage methods. Stability of endogenous and added RNA in blood specimens, serum, and plasma. Describing any variable whose value can affect the outcome of a subsequent analysis. Thavaraj S, Stokes A, Guerra E, Bible J, Halligan E, Long A, et al. Latitude in sample handling and storage for infant faecal microbiota studies: the elephant in the room? They may be unexpected and surprising. Despite all these challenges, genomic and gene expression data generated from FFPE specimens often yield acceptable results in comparison to the original fresh samples. Parekh BS K, Pate S, Nkengasong J, Vercaheren G, Sands A, Ghys P, et al. 3. Vaught JB. Department of Pathology, School of Medicine, Iran University of Medical Sciences, Tehran, Iran, 6 Juan D, Alexe G, Antes T, Liu H, Madabhushi A, Delisi C, et al. In: Kessler HH, editor. Mall C, Rocke DM, Durbin-Johnson B, Weiss RH. Garca-Closas M, Egan KM, Abruzzo J, Newcomb PA, Titus-Ernstoff L, Franklin T, et al. Huijsmans CJ, Damen J, van der Linden JC, Savelkoul PH, Hermans MH. Preanalytics and Precision Pathology: Pathology Practices to Ensure Molecular Integrity of Cancer Patient Biospecimens for Precision Medicine. Sample storage conditions significantly influence faecal microbiome profiles. Paraffin embedding contributes to RNA aggregation, reduced RNA yield, and low RNA quality. on case-matched FFPE and fresh or frozen human sample, using the national cancer institutes Biospecimen Research Database (http://biospecimens.cancer.gov/brd) was done. According to CLSI, for RNA viruses like human immunodeficiency virus (HIV) (and hepatitis C virus (HCV)), the plasma should be separated from whole blood into a second tube within four hours of specimen collection and WHO/UNAIDS guidelines recommend storing serum and plasma at 4C 8C for up to a maximum of one week. Guio H, Okayama H, Ashino Y, Saitoh H, Xiao P, Miki M, et al. Sun F, Reichenberger EJ. Springer J, Morton CO, Perry M, Heinz WJ, Paholcsek M, Alzheimer M, et al. 161 A review article that is in preparation by our office will focus on the effects . Paraffin waxes permeate the tissue in liquid form, solidify rapidly when cooled, and preserve tissue structure in sectioning (27). Fragments of RNA with 300 to 700 bps length could be amplified from methacarn-fixed tissues (24). Nataa KG MM, Milena S, Davidovi S, Dijana T. Direct PCR amplification of the HVSI region in mitochondrial DNA from buccal cell swabs. If no stabilization solution is available and the specimen cannot be frozen, RNA extraction should be done up to four hours after collection (46). It can be either kept at RT for 24 hours or preferably be stored at 4C (ranging from 2C to 8C) for seven days (89). The isolation of nucleic acids from fixed, paraffin-embedded tissues-which methods are useful when? There is a study conducted on Twenty-five lung adenocarcinoma FFPE samples which had been stored at RT for 0.5, 3, 6, 9, and 12 years. Schrader C, Schielke A, Ellerbroek L, Johne R. PCR inhibitors-occurrence, properties and removal. It is mentioned that no significant difference exists between miRNA levels in urines regarding storage at RT up to 72 hours or after seven cycles of freezing and thawing (156). To get reliable results, serum should be used in less than one day or in 2-7 days when preserved at RT or at 4C respectively (80, 81). Ashoor G, Syngelaki A, Poon L, Rezende J, Nicolaides K. Fetal fraction in maternal plasma cellfree DNA at 11-13 weeks' gestation: relation to maternal and fetal characteristics. Colotte M, Coudy D, Tuffet S, Bonnet J. Many pre-analytical factors can alter test results by producing changes that do not reflect the patient's true physiological or clinical condition. Al-Soud WA, Radstrom P. Purification and characterization of PCR-inhibitory components in blood cells. Li S, Tuck-Muller CM, Yan Q, Wertelecki W, Chen H. A rapid method for PCR amplification of DNA directly from cells fixed in Carnoy's fixative. But another study showed that all (100%) shorter fragments of RNA up to 151 bps were amplified by reverse transcription-polymerase chain reaction (RT-PCR) for housekeeping gene (glucose-6-phosphate dehydrogenase) in paraffin-embedded breast cancer samples (39). According to some authors, DNA in CSF is stable at 22C to 25C for up to 24 hours (54), at 2C to 6C for 24 to 72 hours (115), at -20C at least 1 year, and at -70C more than 1 year (54). Although after 6 months, some DNA degradation could be observed in tissues stored in ethanol (20). Resuspension of swabs in transportation fluid can be done according to the manufacturers instructions and stored at -70C or a lower temperature. Comparison of nasopharyngeal flocked swabs and nasopharyngeal wash collection methods for respiratory virus detection in hospitalized children using real-time polymerase chain reaction. Johnson SR, Elkins TE. Ruiz CA, Chaney ME, Tosi AJ. Mouthwash specimen collection is recommended to be done 1 hour after eating or drinking and tooth brushing. (18). da Cunha Santos G. FTA cards for preservation of nucleic acids for molecular assays: a review on the use of cytologic/tissue samples. JAIDS Journal of Acquired Immune Deficiency Syndromes. Correlation of Smoking-Associated DNA Methylation Changes in Buccal Cells With DNA Methylation Changes in Epithelial Cancer. Kopreski MS, Benko FA, Kwak LW, Gocke CD. Quality Assurance and Quality Control in the Molecular Laboratory. Platelet counts and the percentage of platelets retained in the plasma (platelet yield) were higher in plasma obtained by double centrifugation (1000 x g for 5 min followed by 800 x g for 10 min) than by single centrifugation (3500 x g for 10 min).However, WBC counts were higher in single-spin plasma than in double-spin plasma, but there was no difference in RBC counts. For microbiome studies, it is ideal to transport fecal specimens immediately at -20C or -80C. Alfirevic Z, Navaratnam K, Mujezinovic F. Amniocentesis and chorionic villus sampling for prenatal diagnosis. Kim D, Hofstaedter CE, Zhao C, Mattei L, Tanes C, Clarke E, et al. Some specific general considerations should be taken into account when dealing with PND specimens in the clinical laboratory: 1. Beutler E, Gelbart T, Kuhl W. Interference of heparin with the polymerase chain reaction. Bricarelli FD, Hastings RJ, Kristoffersson U, Cavani S. Cytogenetic Guidelines and Quality Assurance A common European framework for quality assessment for constitutional and acquired cytogenetic investigations. Allowable transportation time at 4C is 24-48 hours (134-136). Collection of buccal cells by swab is becoming a widely used procedure for genotyping, detection of early signs of cancer, and regional infection diagnosis (163-166). Human DNA stability is different according to geographic origin due to changes in urine matrix (146). Goelz et al. We hope that the implementation of these recommended key pre-analytical procedures in molecular diagnostic laboratories will improve the baseline level of quality for biospecimens and increase the confidence level while reporting the results. proposed a set of recommendations that are meant to apply to tissue and blood specimens of cancer patients (Compton et al.) The authors deeply appreciate the assistance of Shokoofeh Siroosi in data collection and manuscript preparation.
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