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Introduction: With the global COVID-19 pandemic, nasopharyngeal swabbing has become commonplace and can occasionally cause discomfort, tearing, and anxiety., Objective: To evaluate whether using a neural distraction device during nasopharyngeal swabbing can mitigate patient discomfort., Methods: Participants were randomized into nasal swab with vibration first (treatment-control or Tc) or nasal swab without vibration first (control-treatment or Ct). Then the swab was repeated with the opposite technique. The vibration device was used to stimulate the infraorbital nerve at the midpupillary line, while a nasopharyngeal swab was performed according to CDC protocol. After each swab, participants filled out a survey to assess pain, discomfort, and tearing. This procedure was replicated on the contralateral nostril for a total of 4 swabs and 4 surveys., Results: Thirty-four subjects were included in the analysis. In the Ct group, there was a median 1.0-point decrease in pain with the use of vibration (95% CI [-1.8, -0.3] , P = 0.0117), 2.5-point decrease (95% CI [-3.0, -0.8], P = 00039) in discomfort, and a 1.0-point decrease (95% CI [-2.6, -0.2] P = 0.0303) in tearing when the vibratory device was used. In the Tc group, there was a median 2.0-point decrease in pain with vibration (95% CI [ -2.7, -1.1], P < 0.0001), 2.3-point decrease in discomfort (95% CI [-3.1, -1.5], P < 0.0001), and a 1.5-point decrease in tearing (95% CI [-2.5, -0.5], P = 0.0037) when vibration was used., Conclusion: Vibratory stimulation of the infraorbital nerve is safe and effectively reduced pain, discomfort, and tearing during nasopharyngeal swab. Vibration is a reasonable option to minimize patient discomfort during COVID-19 or other microbiological testing., Competing Interests: Declaration of conflicting interestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Stool specimen collection during a foodborne or enteric illness outbreak investigation is essential for determining the outbreak etiology and for advancing the epidemiologic understanding of the pathogens and food vehicles causing illness. However, public health professionals face multifaceted barriers when trying to collect stool specimens from ill person during an outbreak investigation. The Colorado Integrated Food Safety Center of Excellence (Colorado IFS CoE) and the Arizona Department of Health Services surveyed local public health agencies (LPHAs) to identify barriers to collecting ≥2 clinical specimens in foodborne and enteric illness outbreaks. The most commonly selected patient-related barrier was that the patient did not think it is important to provide a stool sample because they are well by the time the LPHA follows-up (61%). The most frequently selected outbreak-related barrier was the LPHA did not learn about the outbreak until after symptoms had resolved (61%). Time/personnel not being available for stool collection was the most frequently chosen health department-related barrier (51%). Timing of the outbreak (e.g., on a weekend or holiday) was the most frequently selected transportation-related barrier (51%) to collecting ≥2 stool specimens. Many of the frequently cited barriers in this survey were similar to those previously reported, such as workforce capacity and patient privacy concerns, indicating that these barriers are ongoing. Reducing barriers to stool collection during outbreaks will require efforts led at the national and state levels, such as increased enteric illness program funding, educating public health staff on the importance of specimen collection during every enteric illness outbreak, and providing specimen collection resources to LPHA staff., Competing Interests: Declaration of Competing Interest This work was supported by the Centers for Disease Control and Prevention 5NU50CK000522-03-00, otherwise the authors have no relevant financial or non-financial interests to disclose., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

Nasopharyngeal swabs have been widely to prevent the spread of coronavirus disease 2019 (COVID-19). Nasopharyngeal COVID-19 testing is a generally safe and well-tolerated procedure, but numerous complications have been reported in the media. Therefore, the present study aimed to review and document adverse events and suggest procedural references to minimize preventable but often underestimated risks. A total of 27 articles were selected for the review of 842 related documents in PubMed, Embase, and KoreaMed. The complications related to nasopharyngeal COVID-19 testing were reported to be rarely happened, ranging from 0.0012 to 0.026%. Frequently documented adverse events were retained swabs, epistaxis, and cerebrospinal fluid leakage, often associated with high-risk factors, including severe septal deviations, pre-existing skull base defects, and previous sinus or transsphenoidal pituitary surgery. Appropriate techniques based on sufficient anatomical knowledge are mandatory for clinicians to perform nasopharyngeal COVID-19 testing. The nasal floor can be predicted by the line between the nostril and external ear canal. For safe testing, the angle of swab insertion in the nasal passage should remain within 30° of the nasal floor. The swab was gently inserted along the nasal septum just above the nasal floor to the nasopharynx and remained on the nasopharynx for several seconds before removal. Forceful insertion should be attempted, and alternative examinations should be considered, especially in vulnerable patients. In conclusion, patients and clinicians should be aware of rare but possible complications and associated high-risk factors. The suggested procedural pearls enable more comfortable and safe nasopharyngeal COVID-19 testing for both clinicians and patients., Competing Interests: All authors have no potential conflicts of interest to disclose., (© 2022 The Korean Academy of Medical Sciences.)

Transnasal swab testing for the detection of SARS-CoV-2 is well established. The Centers for Disease Control and Prevention advocates swabbing either of the anterior nares, middle turbinate, or nasopharynx for specimen collection depending on available local resources. The purpose of this review is to investigate complications related to transnasal SARS-CoV-2 testing with specific attention to specimen collection site and swab approach. The literature demonstrates that while nasopharyngeal swabbing is associated with an increased risk of complications, it should remain the gold-standard test due to greater diagnostic accuracy relative to anterior nasal and middle turbinate swabs., Competing Interests: Competing interests: JHC, SP and MK declare they have no relevant conflicts of interest. RMN is part of the Medical Advisory Board for GlaxoSmithKline, Lyra, Sanofi, Regeneron, and AstraZeneca., (© American Federation for Medical Research 2021. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Suboptimal human semen handling in vitro may induce sperm damage. However, the effects of semen swim-up, pellet swim-up, density gradient, and density gradient followed by SU on sperm motility, morphology, DNA fragmentation, acrosome reaction, intracellular reactive oxygen species, and mitochondrial activity were not fully understood., Objectives: To study the impact of four sperm preparation techniques on sperm functional parameters., Materials and Methods: This study was conducted on 60 infertile men with a minimum sperm concentration of 20 × 10 6 /ml and total sperm motility of ≥30%. Each raw semen sample was divided into four aliquots. Each aliquot was prepared by one of the tested techniques. Various sperm characteristics were assessed before and after sperm preparation., Results: Density gradient and density gradient followed by SU resulted in significantly higher DNA fragmentation percentages compared with semen swim-up (p < 0.001 and p < 0.001, respectively) and pellet swim-up (p < 0.001 and p < 0.001, respectively). Significantly higher percentages of spermatozoa with intact acrosome were detected in semen swim-up (p < 0.001) and pellet swim-up (p < 0.001) compared with raw semen. The percentage of reactive oxygen species-positive spermatozoa was significantly higher after pellet swim-up (p < 0.001), density gradient (p < 0.001), and density gradient followed by SU (p < 0.001) than raw semen. In addition, the percentages of 100% stained midpiece (active mitochondria) were significantly higher in semen swim-up (p < 0.001) and pellet swim-up (p < 0.001) compared with raw semen., Discussion and Conclusion: To the best of our knowledge, this is the first report comparing the impact of these techniques on various sperm functional parameters. Semen swim-up was more effective than density gradient in selecting better spermatozoa in terms of DNA integrity, reactive oxygen species levels, acrosome status, and mitochondrial activity. Randomized clinical trials comparing these four techniques are required to test their impact on embryo development and pregnancy outcomes., (© 2021 American Society of Andrology and European Academy of Andrology.)

Research Question: During the embryo transfer procedure, to what degree of temperature drop are embryos exposed to between loading the transfer catheter and placing it into the uterus?, Design: Twenty-nine simulated embryo transfer procedures were carried out across five clinics. A thermocouple probe was used for standardized measurements inside the embryo transfer catheter to investigate the change in temperature that occurred in the time period between loading and placing the catheter in the uterus., Results: In all cases, the temperature at the loaded catheter tip fell rapidly to ambient temperature during transit from the embryo transfer workstation in the laboratory to the procedure room, even though embryo transfer procedures, ambient temperatures and embryo transfer catheter temperatures at loading varied between clinics., Conclusions: Given the sensitivity of the pre-implantation embryo to its immediate environment, the rapid and profound drop in temperature observed at the catheter tip that houses the embryo during transit from laboratory to the uterus may affect embryo viability and health. This issue should be addressed to ensure that the tight temperature control aimed for by IVF laboratories continues throughout the embryo transfer procedure, and could improve clinical outcomes., (Copyright © 2021. Published by Elsevier Ltd.)

Background: Transient global amnesia (TGA) is a clinical syndrome characterized by a temporary short-term memory loss with inability to retain new memories, usually lasting 2 to 8 h. TGA may be related to several medical procedures, including angiography, general anesthesia, gastroscopy., Case Presentation: We report a 58-year-old woman who experiencing TGA one hour after the execution of her first-time nasopharyngeal swab for COVID-19. Brain MRI showed a typical punctate Diffusion Weight Image (DWI) hippocampal lesion., Conclusions: This is the first report of TGA after the execution of nasopharyngeal swab for COVID-19. This association lengthen the list of medical procedures associated with TGA, and we discuss the possible plausible mechanisms by which a nasopharyngeal swab could trigger TGA.

Background: Procedural pain is underestimated in hospitalized preterm infants. The aim of this study was to assess the reliability, validity, and clinical utility of the Neonatal Facial Coding System (NFCS), Douleur Aiguë du Nouveau-né (DAN) scale, Neonatal Infant Pain Scale (NIPS), and Premature Infant Pain Profile (PIPP) in premature infants undergoing heel blood collection. We assume that the four scales were similar in reliablility and validity (but different in clinical utility)., Methods: The pain assessments were performed on 111 premature infants using the four scales. Internal consistency was determined by Cronbach's α, and the reliability was determined by the intraclass correlation coefficients. Concurrent validity was evaluated by Spearman's rank correlations. Bland-Altman plots were used to investigate the convergent validity., Results: The internal consistency and their reliability of the scales were high (p < 0.001). Scores were significantly higher at the time of blood collection (p < 0.001). Mean scores of clinical utility of PIPP were significantly higher than NFCS and DAN (p < 0.05) but not higher than the NIPS (p > 0.05)., Conclusions: The four scales were reliable and valid. This study suggests that the PIPP and NIPS has good clinical utility and are better choice for evaluating procedural pain in premature infants., Impact: The aim of this study was to assess the reliability, validity, and clinical utility of NFCS, DAN, NIPS, and PIPP in premature infants undergoing heel blood collection. The results showed that the four scales have high reliability and internal consistency; the PIPP and NIPS have good clinical utility and are better choice for evaluating procedural pain in premature infants. Our study results provided a reference for clinical workers in choosing pain assessment scales and conduction intervention.

Background/aim: To evaluate complications and risk factors associated with transumbilical incision as an organ removal site in laparoscopic pancreatectomy (LP)., Patients and Methods: In total, 52 patients who underwent LP between 2009 and 2017 were included in this study. The development of superficial surgical site infection (SSI) and transumbilical port-site incisional hernia was recorded., Results: None of the patients had SSI. However, three (5.77%) presented with transumbilical incisional hernia. No variables were significantly associated with the risk of transumbilical incisional hernia., Conclusion: No evident risk factors correlated with hernia formation. Hence, incisional hernia might have occurred at a certain probability. In some cases, it was caused by technical problems. However, the use of transumbilical incision as an organ removal site was feasible, and a new incision for organ removal alone was not required., (Copyright © 2021 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Introduction: The pneumatic tube system (PTS) is widely used for sample delivery. We aimed to investigate the impacts of PTS on hemostasis assays., Methods: Triplicate samples from 30 healthy volunteers were delivered to the core laboratory manually by human courier or via the 500 m long-distance PTS or via the 1000 m long-distance PTS. Comparisons of 19 hemostasis tests were conducted., Results: Although PT, INR, APTT, FII, FV, FVII FIX, FX, FXII, DD, α2-PI, and PC had statistical significance (all P < .05), all had low average bias remaining within clinical acceptable limits. PTS transportation only resulted in a statistically significant and clinically relevant decrease in FVIII activity. In the 500 m-PTS group, 66.7% (20/30) of samples for FVIII testing had a bias greater than 8.3%. Moreover, in the 1000 m-PTS group, 96.7% (29/30) of samples had a bias of over 8.3%, and the maximal bias achieved 42.1%., Conclusions: Pneumatic tube system in our institution could be used to deliver blood samples for hemostasis tests evaluated in this study except FVIII activity assay., (© 2020 John Wiley & Sons Ltd.)

Spread Through Air Spaces (STAS) is a form of invasion characterized by neoplastic cell dissemination in the lung parenchyma surrounding the outer edge of the tumor. Its possible artifactual origin is widely debated in the literature. The aim of this study is to investigate the potential impact of gross sampling procedures in causing STAS. A prospective series of 51 surgical lung specimens was collected (35 adenocarcinomas, 68.6%; 13 squamous cell carcinomas, 25.5%; 2 large-cell neuroendocrine carcinomas, 3.9%; 1 atypical carcinoid, 2%). The fresh tissue was sectioned with a new and clean blade for each cut, to obtain a tissue slice comprising the upper lung parenchyma, the tumor, and the lower parenchyma. This slice was cut in half and separately processed. The same procedure was repeated in the residual (specular) specimen after formalin fixation. STAS was identified in 33/51 (64.7%) cases, the predominant pattern being cluster formation (29 cases, 87.9%), the remaining 4 cases having single-cell invasion. Comparing STAS detection in upper and lower lung parenchyma areas (ie, before and after the blade crossed the tumor), no significant preferential STAS distribution was observed, indeed being almost overlapping (60.6% and 63.6% for fresh and 61.3% and 65.6% for fixed tissues, respectively). There was no difference between STAS occurrence in freshly cut and fixed corresponding samples. These findings indicate that STAS is not a pathologist-related artifactual event because of knife transportation of tumor cells during gross specimen handling and support the notion that it is a phenomenon preexisting to surgical tissue processing., Competing Interests: Conflicts of Interest and Source of Funding: The authors have disclosed that they have no significant relationships with, or financial interest in, any commercial companies pertaining to this article., (Copyright © 2020 Wolters Kluwer Health, Inc. All rights reserved.)

Long and sharp objects can be foreign body intentionally or accidentally ingested. Timing of endoscopy relies on foreign body shape and size, localization in gastrointestinal tract, patient's clinical conditions, occurrence of symptoms, or onset of complications. We present a case of a 47-year-old male with no known comorbidity, who accidentally swallowed a portion of a naso-pharyngeal swab half-broken during the second diagnostic test for SARS-CoV-2. The intact swab had a total length of 15 cm and was made of wood. The patient was asymptomatic, laboratory tests were normal, and neck-chest-abdominal X-ray and CT scan were negative for major complications. Upper gastrointestinal endoscopy was promptly performed to prevent the long sharp swab from crossing the pylorus leading to serious complications and, therefore, risk surgical intervention. The patient was intubated and the procedure was carried out under general anesthesia. In the gastric body, broken naso-pharyngeal swab was detected among the food debris, and using a latex rubber hood, the 7.5 cm foreign body was removed with a retrieval alligator-tooth forceps. Our hospital is located in a high-risk area of COVID-19 outbreak where many naso-pharyngeal swabs are performed, and to our knowledge, this is the first report of swab ingestion during SARS-CoV-2 test.

We report the case of a bronchial foreign body, following a tracheostomy site swab for SARS-CoV-2, aiming to raise awareness and vigilance. A qualified nurse was performing a routine SARS-CoV-2 swab on a 51-year-old woman, fitted with a tracheostomy in the recent past following a craniotomy. This was part of the discharging protocol to a nursing home. During the sampling, part of the swab stylet snapped and was inadvertently dropped through the tracheostomy site. Initial CT imaging was reported as showing no signs of a foreign body but some inflammatory changes. Bedside flexible endoscopy through the tracheostomy site revealed the swab in a right lobar bronchus. This was subsequently removed by flexible bronchoscopy. This case highlights the need for clear guidance on how samples for SARS-CoV-2 are taken from patients with front of neck airways (laryngectomy/tracheοstomy) and the potential pitfalls involved., Competing Interests: Competing interests: None declared., (© BMJ Publishing Group Limited 2020. No commercial re-use. See rights and permissions. Published by BMJ.)

Introduction: Measurements from frozen sample collections are important key indicators in clinical studies. It is a prime concern of biobanks and laboratories to minimize preanalytical bias and variance through standardization. In this study, we aimed at assessing the effects of different freezing and thawing conditions on the reproducibility of medical routine parameters from frozen samples. Materials and Methods: In total, 12 pooled samples were generated from leftover lithium heparinized plasma samples from clinical routine testing. Aliquots of the pools were frozen using three freezing methods (in carton box at -80°C, flash freezing in liquid nitrogen, and controlled-rate freezing [CRF]) and stored at -80°C. After 3 days, samples were thawed using two methods (30 minutes at room temperature or water bath at 25°C for 3 minutes). Ten clinical chemistry laboratory parameters were measured before (baseline) and after freeze-thaw treatment: total calcium, potassium, sodium, alanine aminotransferase, lactate dehydrogenase (LDH), lipase, uric acid, albumin, c-reactive protein (CRP), and total protein. We evaluated the influence of the different preanalytical treatments on the test results and compared each condition with nonfrozen baseline measurements. Results: We found no significant differences between freezing methods for all tested parameters. Only LDH was significantly affected by thawing with fast-rate thawing being closer to baseline than slow-rate thawing. Potassium, LDH, lipase, uric acid, albumin, and CRP values were significantly changed after freezing and thawing compared with unfrozen samples. The least prominent changes compared with unfrozen baseline measurements were obtained when a CRF protocol of the local biobank and fast thawing was applied. However, the observed changes between baseline and frozen samples were smaller than the measurement uncertainty for 9 of the 10 parameters. Discussion: Changes introduced through freezing-thawing were small and not of clinical importance. A slight statistically based preference toward results from slow CRF and fast thawing of plasma being closest to unfrozen samples could be supported.

Background: Objective methods for distinguishing melanoma in situ (MIS) from photodamaged skin (PS) are needed to guide treatment in patients with melanocytic proliferations. Melanocyte density (MD) could serve as an objective histopathological criterion in difficult cases. Calculating MD via manual cell counts (MCC) with immunohistochemical (IHC)-stained slides has been previously published. However, the clinical application of this method is questionable, as quantification of MD via MCC on difficult cases is time consuming, especially in high volume practices., Methods: ImageJ is an image processing software that uses scanned slide images to determine cell count. In this study, we compared MCC to ImageJ calculated MD in microphthalmia transcription factor-IHC stained MIS biopsies and control PS acquired from the same patients., Results: We found a statistically significant difference in MD between PS and MIS as measured by both MCC and ImageJ software (P < 0.01). Additionally, no statistically significant difference was found when comparing MD measurements recorded by ImageJ vs those determined by the MCC method., Conclusion: MD as determined by ImageJ strongly correlates with the MD calculated by MCC. We propose the use of ImageJ as a time-efficient, objective, and reproducible tool to assess MD., (© 2020 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Human umbilical vein endothelial cells (HUVECs) have great potential in tissue engineering, regenerative medicine, and clinical applications. There is an ever-increasing demand to provide living HUVECs and HUVECs-hydrogel constructs to end users when needed in cell-based therapy and clinical applications. However, current methods to provide living cells and their constructs are mainly continuous culture and cryopreservation, which are high-cost, labor-intensive, time-consuming and inflexible. The research about hypothermic storage of HUVECs and their hydrogel constructs is still limited. Here, we studied the cell survival of HUVECs without encapsulation (W/O Encap) or with encapsulation (alginate, alginate with carboxymethyl chitosan [CMCH]) at 4°C and 25°C during 7 days, respectively. Also, we explored the optimal CMCH concentration for hypothermic storage, which is 0.5% (w/v) at 4°C and 25°C. Moreover, we evaluated the cell attachment after hypothermic storage. Our results enable the hypothermic storage of HUVECs and HUVEC-hydrogel constructs, and facilitate their application in tissue engineering and clinical medicine.

Background: Coronavirus disease 2019, a highly transmissible respiratory infection, has created a public health crisis of global magnitude. The mainstay of diagnostic testing for coronavirus disease 2019 is molecular polymerase chain reaction testing of a respiratory specimen, obtained with a viral swab. As the incidence of new cases of coronavirus disease 2019 increases exponentially, the use of viral swabs to collect nasopharyngeal specimens is anticipated to increase drastically., Case Report: This paper draws attention to a complication of viral swab testing in the nasopharynx and describes the premature engagement of a viral swab breakpoint, resulting in impaction in the nasal cavity., Conclusion: This case highlights a possible design flaw of the viral swab when used to collect nasopharyngeal specimens, which then requires an aerosol-generating procedure in a high-risk patient to be performed. The paper outlines a safe technique of nasal foreign body removal in a suspected coronavirus disease 2019 patient and suggests alternative testing materials.

Background and Aim: Avoiding abdominal incisions is one of the unique features of natural orifice specimen extraction (NOSE) surgery. There has been no consensus on whether the avoidance of abdominal incisions can reduce the systemic inflammatory response after NOSE surgery. This study was designed to evaluate the safety and feasibility of transanal NOSE, especially the inflammatory response after transanal NOSE versus mini-laparotomy (LAP)., Methods: A total of 172 colorectal cancer patients who underwent transanal NOSE were matched with 172 patients who underwent LAP for colorectal cancer. Clinical characteristics, pathological features, perioperative parameters, and indicators of the inflammatory response were collected and compared. The inflammatory response was assessed by measuring body temperature, neutrophil count, C-reactive protein levels, and procalcitonin levels., Results: Patients in the NOSE group had better short-term outcomes, such as lower incidence of wound infection, less postoperative pain, less need for anesthetic drugs, and faster recovery of intestinal function. Regarding the inflammatory response, the average body temperature of patients in the NOSE group was higher on postoperative day (POD) 2 than that in the LAP group. A higher median neutrophil count and C-reactive protein levels were observed in the NOSE group on POD3 and POD5 than was observed in the LAP group., Conclusions: Transanal NOSE is safe and feasible for colorectal cancer, with better short-term outcomes. Although transanal NOSE produced a larger systemic inflammatory response than LAP in the early postoperative stages, this response did not appear to translate into infectious morbidity., (© 2019 Journal of Gastroenterology and Hepatology Foundation and John Wiley & Sons Australia, Ltd.)

Objectives: To monitor blood culture contamination (BCC) rates in a tertiary care hospital in Saudi Arabia. Methods: Blood cultures submitted to the Microbiology Laboratory of King Fahad Hospital, Madina, Saudi Arabia between January and December 2017 were analyzed prospectively. Positive blood cultures were either designated as true bacteremia with confirmed bloodstream infection or BCC., Results: Among 5,536 blood cultures from 2201 patients, 364 (6.6%) mirrored BCC. There was an upward trend in contamination rates in specific months. With respect to total blood cultures from respective units over a one-year period, medical ward contributed to the highest contamination rate (10.3%). Blood culture contamination rate in the wards ranged from 4.5-10.3%, with a higher contamination rate in elderly, aged 60-80 years. Staphylococcus epidermidis (S. epidermidis) was the most frequent contaminant (44.5%). Conclusion: The escalated contamination rates in September to October may be attributed to difficulty in sampling blood by the less competent nurses during annual pilgrimage season. High influx of patients and shortage of trained nurses may have resulted in increased incidence in December-January and March-April. The prevalence of skin-resident S. epidermidis may be due to improper aseptic conditions. Ours is the first report on evaluation of BCC rates in Madina and call for renewed efforts in this direction.

The handling-induced dark neuron is a histological artifact observed in brain samples handled before fixation with aldehydes. To explore associations between dark neurons and immunohistochemical alterations in mouse brains, we examined protein products encoded by Cav3 (neuronal perikarya/neurites), Rbbp4 (neuronal nuclei), Gfap (astroglia), and Aif1 (microglia) genes in adjacent tissue sections. Here, dark neurons were incidental findings from our prior project, studying the effects of age and high-fat diet on metabolic homeostasis in male C57BL/6N mice. Available were brains from 4 study groups: middle-aged/control diet, middle-aged/high-fat diet, old/control diet, and old/high-fat diet. Young/control diet mice were used as baseline. The hemibrains were immersion-fixed with paraformaldehyde and paraffin-embedded. In the hippocampal formation, we found negative correlations between dark neuron hyperbasophilia and immunoreactivity for CAV3, RBBP4, and glial fibrillary acidic protein (GFAP) using quantitative image analysis. There was no significant difference in dark neuron hyperbasophilia or immunoreactivity for any protein examined among all groups. In contrast, in the hippocampal fimbria, old age seemed to be associated with higher immunoreactivity for GFAP and allograft inflammatory factor-1. Our findings suggest that loss of immunohistochemical reactivity for CAV3, RBBP4, and GFAP in the hippocampal formation is an artifact associated with the occurrence of dark neurons. The unawareness of dark neurons may lead to misinterpretation of immunohistochemical reactivity alterations.

Background: Next generation sequencing (NGS) can recover DNA data from valuable extant and extinct museum specimens. However, archived or preserved DNA is difficult to sequence because of its fragmented, damaged nature, such that the most successful NGS methods for preserved specimens remain sub-optimal. Improving wet-lab protocols and comprehensively determining the effects of sample age on NGS library quality are therefore of vital importance. Here, I examine the relationship between sample age and several indicators of library quality following targeted NGS sequencing of ~ 1300 loci using 271 samples of pinned moth specimens (Helicoverpa armigera) ranging in age from 5 to 117 years., Results: I find that older samples have lower DNA concentrations following extraction and thus require a higher number of indexing PCR cycles during library preparation. When sequenced reads are aligned to a reference genome or to only the targeted region, older samples have a lower number of sequenced and mapped reads, lower mean coverage, and lower estimated library sizes, while the percentage of adapters in sequenced reads increases significantly as samples become older. Older samples also show the poorest capture success, with lower enrichment and a higher improved coverage anticipated from further sequencing., Conclusions: Sample age has significant, measurable impacts on the quality of NGS data following targeted enrichment. However, incorporating a uracil-removing enzyme into the blunt end-repair step during library preparation could help to repair DNA damage, and using a method that prevents adapter-dimer formation may result in improved data yields.

This work presents the development of a methodology for the accurate and precise quantification of the renal biomarker Cystatin C in human urine by Isotope Dilution Mass Spectrometry (IDMS). The procedure is based on the addition of a known quantity of the proteotypic peptide ALDFAVG*EYNK labelled with 13 C 2 -glycine to the urine sample followed by protein hydrolysis using trypsin. Then, preconcentration and purification of the isotope diluted peptide was carried out by a selective monoclonal antibody bound to magnetic beads and final measurement was done after injection of the sample in a HPLC-MS/MS triple quadrupole instrument. The isotopic distribution of the isotope diluted proteotypic peptide was measured by low resolution selected reaction monitoring. Using this aquisition mode, the bandpass of the first quadrupole was widened (FWHM =13 u) so the whole isotopic clusters for both the natural abundance and the labelled peptides entered the collision cell. The proposed acquisition mode provided similar accuracy and precision than the regular SRM mode (FWHM =0.7 u) but a higher sensitivity was observed. The purification of the sample by antibody based enrichment of the target peptide was shown to remove interfering compounds more efficiently in comparison with a sample purification based on semipreparative liquid chromatography. Using 5 ng of the labelled peptide it was possible to quantify Cystatin C in human urine in patients with normal and impaired renal function. Recoveries from 100 to 104% were obtained in samples containing from 90 to 700 μg L -1 of Cystatin C with relative standard deviations from 0.5 to 6%. The stability of Cystatin C in urine samples was evaluated under different storage conditions showing that only when the urine samples were stored at room temperature during more than 10 days, a significant degradation of Cystatin C was observed., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Despite the proven benefits of laparoscopic surgery, it is indeed very costly. The aim of our study was to show an effective way to reduce one of the costs. Between January 2012 and December 2018, we used sterile unpowdered latex-free surgical gloves for specimen retrieval in 243 selected cases of laparoscopic cholecystectomy and appendectomy. The mean retrieval time was 6.7 ± 3.6 min. All procedures were performed safely. Minor wound infection was noted in three patients but there was no case of port site hernia in our series. We conclude that specimen retrieval using sterile, unpowdered, latex-free surgical gloves is safe, effective and cheap. No special additional preparation is required.

Clinical trial faecal collections present challenges through geographical spread and inexperienced participants. Collection techniques have been developed and tested to overcome these challenges, but previous studies investigating these techniques have demonstrated a highly variable capacity for sample preservation. Furthermore, these studies typically only examine either preservation of genetic content or metabolites, not both. This study investigated the Stool Nucleic Acid Collection and Preservation Tube (Norgen BioTek Corp) for the preservation of both microbial DNA and microbial organic acid metabolites in human faecal samples when compared to frozen samples. Twenty six healthy adult participants were instructed to collect a bowel movement, subsample into collection tubes and immediately transfer the remaining bulk to -20 °C storage. Resulting organic acid concentrations remained comparable across methods when the preservation tubes were used correctly. The 16S rRNA gene sequencing data revealed twenty significantly different bacterial genera between the two collection methods. Ten Gram-negative genera were more abundant in the collection tubes, and ten Gram-positive genera were more abundant in the fresh frozen samples. This study has illustrated that faecal collection methods bias the microbial community profile according to Gram status and this should be considered when designing studies that collect and store human faecal samples.

To evaluate the characteristics of microbiological contamination in donor corneas preserved for medium-term. A total of 82 donated corneas from June 1, 2014 to November 30, 2014 were retrospectively analyzed. The corneas were preserved in cornea chambers medium-term solution at 4-8 °C for keratoplasty. After removal of the central corneas for transplantation, the corneoscleral rims were put back into the medium for 1 month at room temperature (20-25 °C). The suspicious contaminated storage solutions indicated with transparency or color change were examined with bacteria and fungi cultivation for strain identification. The data collected included gender, age, procurement site and causes of death of donors, and follow-up of recipients. Statistical analysis was performed using Microsoft Excel and SPSS 24.0. Significance level was set at a P value < 0.05. The overall pathogen positive rate was 9.8% (n = 8), including 7 (87.5%) fungi and 1 (12.5%) bacteria. They were 2 (2.44%) Fusarium, 2 (2.44%) Chromomycosis, 1 (1.22%) Candida albicans, 1 (1.22%) Aspergillus versicolor, 1 (1.22%) Acremonium species, and 1 (1.22%) Enterococcus. 5 contaminated corneas were used for penetrating keratoplasty; although four out of five (80%) had not been given antifungal drugs during more than 6 months following-up period, none of the recipients was infected with a graft. Donor age (P = 0.839), gender (P = 0.062), procurement sites (P = 0.713) and cause of death (P = 0.711) had no statistically significant influence on the contamination rate. All donor corneas have a possibility of microbiological contamination. Strict tissue preservation protocol but not antifungal drugs following keratoplasty seems necessary to prevent graft infection.

The aim is to examine whether brain tissue samples obtained through needle biopsy are better for histopathological evaluation when obtained with defined vacuum pressure, a novel needle rotation method, and using different needle type - Laitinen or Nashold. Moreover the paper aims to answer the question: Does vacuum and mechanical injury resulting from different sampling methods damage the tissue specimen challenging the diagnosis?. Eight hundred biopsy specimens from fresh swine brains were obtained using Nashold and Laitinen brain biopsy needles through inner cannula cutting or needle rotation sampling at vacuum pressure, from 0 to 0.06 MPa. The specimen weight and tissue quality for microscopic assessment were evaluated using the Mair score. Rising aspiration pressure increased the biopsy sample weight. Needle rotation delivered larger biopsy samples than the standard method. Laitinen provided larger samples than the Nashold needle, with the same sampling method or vacuum pressure. A higher histopathological diagnostic quality of tissue was obtained with the Laitinen needle than with Nashold, with higher vacuum pressure than lower pressure and finally with needle rotation than the standard method. No tissue damage caused by higher suction pressure or method of tissue separation was documented. Brain tissue samples obtained through needle biopsy are better for histopathological evaluation when obtained with higher vacuum pressure, a novel needle rotation method and with Laitinen needle. Higher suction pressure and sampling methods did not cause tissue damage.

Although in vivo measurements of larval insects are often performed during developmental studies of necrophilous beetles, their impact on development has not been studied. During measurements insects are taken out of the incubator for a few minutes at room temperature, which may affect the development and eventually the quality of the developmental reference data. Additionally, while being measured larvae are under stress which may have an effect on their development. We conducted an experiment using predatory beetle species Creophilus maxillosus L. (Coleoptera: Staphylinidae) which often occurs and breeds on large vertebrate carcasses. We tested the hypothesis that multiple, in vivo measurements affect the development of C. maxillosus by increasing its duration and changing adult insect size at emergence. As a consequence, we predicted that the multiple insect measurement protocol will affect the accuracy of age estimates using the resultant reference developmental data. Development of C. maxillosus was studied at 7 constant temperatures. All individuals were inspected for developmental landmarks; half of them were also repeatedly measured and weighed. Measured beetles developed longer than non-measured beetles (e.g. 1.59 days longer at 22.5 °C) and at emergence were distinctly smaller (e.g. 1.5 mm shorter and 22 mg lighter at 22.5 °C). The accuracy of age estimates was greater while using the model for non-measured beetles. These results support the claim that multiple in vivo measurements of insects reduce the quality of resultant developmental data. The measurements were particularly detrimental for the adult insect size. Consequently, particular attention should be paid to isomegalen diagrams which are based on insect length. Our findings indicate that these diagrams, when based on multiple, in vivo measurements of larval beetles, will systematically overestimate their age., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Background: There are little data regarding error within the multistep process of Mohs micrographic surgery (MMS)., Objective: A survey of Mohs surgeons was performed to evaluate variation in tissue handling and processing within the MMS process., Methods and Materials: A 9-question electronic survey was distributed to members of the American College of Mohs Micrographic Surgery. A total of 97 responses were analyzed., Results: Most surgeons personally transport the tissue (71.1%), most frequently in a specimen container (54.6%). Method of tissue identification during transportation varied significantly. Most surgeons personally hand-draw the map (77.1%). A diagram was most commonly used to convey tissue orientation to the histotechnician (65.4%). Only 31.3% of histotechnicians used labeling of blocks to identify tissue within the cryostat. Most respondents (72.2%) are the only surgeon processing tissue in the laboratory at one time; however, 28.9% reported performing 10 or more cases per day., Conclusion: Most respondents are responsible for tissue transport and mapping, which likely reduces potential error. However, the method of transport and labeling, as well as identification by the histotechnician, was more variable. Ultimately, having a consistent process and clear communication with personnel will help reduce potential error during Mohs surgery.

Context.—: Phosphatase and tensin homolog (PTEN) is a promising prognostic and potentially predictive biomarker in prostate cancer., Objective.—: To assess the effects of preanalytic variables on an analytically validated and fully automated PTEN immunohistochemistry assay., Design.—: PTEN immunohistochemistry was performed on Ventana immunostaining systems. In benign prostate tissues, immunostaining intensity across variable conditions was assessed by digital image analysis. In prostate tumor tissues, immunostaining was scored visually., Results.—: Delay of fixation for 4 hours or longer at room temperature or 48 hours or longer at 4°C and duration of formalin fixation did not significantly alter immunostaining intensity. Intensity of staining was highest in 10% formalin compared with other fixatives. Tumor tissues with PTEN loss processed using protocols from 11 academic institutions were all evaluable and scored identically. PTEN immunostaining of needle biopsies where tissue blocks had been stored for less than 10 years was more frequently scored as nonevaluable compared with blocks that had been stored for 10 years or longer. This effect was less evident for radical prostatectomy specimens, where low rates of nonevaluable staining were seen for 23 years or more of storage. Storage of unstained slides for 5 years at room temperature prior to immunostaining resulted in equivalent scoring compared with freshly cut slides. Machine-to-machine variability assessed across 3 Ventana platforms and 2 institutions was negligible in 12 tumors, and platform-to-platform variability was also minor comparing Ventana and Leica instruments across 77 tumors (κ = 0.926)., Conclusions.—: Automated PTEN immunostaining is robust to most preanalytic variables in the prostate and may be performed on prostate tumor tissues subjected to a wide range of preanalytic conditions. These data may help guide assay development if PTEN becomes a key predictive biomarker.

Background & Aims: Diagnosis and surveillance of Barrett's esophagus (BE) and eosinophilic esophagitis (EoE) have become emerging public health issues. Cytosponge is a novel, minimally invasive esophageal cell collection device. We aimed to assess the data on safety and acceptability of this device., Methods: We performed a patient-level review of 5 prospective trials assessing Cytosponge performance in patients with reflux disease, BE and EoE in primary and secondary care. Acceptability of Cytosponge and subsequent endoscopy were recorded with visual analogue scale (VAS), wherein 0 and 10 denoted lowest and highest acceptability. Median VAS scores were compared using a Mann-Whitney test. The number of attempts, failures in swallowing the device and occurrence of adverse events were analyzed. Risk factors for failure in swallowing were analyzed using a multivariate regression model., Results: In total, 2672 Cytosponge procedures were performed, in 2418 individuals from 2008 through 2017. There were 2 adverse events related to the device: a minor pharyngeal bleed and a case of detachment (<1:2000). The median acceptability score for the Cytosponge was 6.0 (interquartile range [IQR], 5.0-8.0), which was higher than the score for endoscopy without sedation (median 5.0; IQR, 3.0-7.0; P < .001) and lower than the score for endoscopy with sedation (median 8.0; IQR, 5.0-9.0; P < .001). Nearly all patients (91.1%) successfully swallowed the Cytosponge, most on the first attempt (90.1%). Failure to swallow the device was more likely to occur in secondary care (odds ratio, 5.13; 95% CI, 1.48-17.79; P < .01)., Conclusions: The Cytosponge test is a safe procedure with good acceptability ratings in a variety of health care settings., (Copyright © 2019 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Milenkovic T, Vukovic R, Radojicic B, Mitrovic K, Todorovic S, Zatezalo L. Thirty years of the newborn screening program in Central Serbia: the missed cases of congenital hypothyroidism. Turk J Pediatr 2019; 61: 319- 324. Newborn Screening (NS) program for congenital hypothyroidism (CH) has completely changed the natural history of this devastating disorder for the vast majority of children with CH. However, cases of missed CH do happen, and it is important to keep this possibility in mind during every day clinical practice. The objective of this study was to analyze the recognized cases of undiagnosed CH by the NS program in Central Serbia in order to evaluate the reasons for such omissions. Data regarding the recognized missed cases of CH between the years of 1983 and 2013 were collected and analyzed. During 30 years of the NS program for CH on the territory of Central Serbia, 1,547,122 newborns were screened, and during this period, 14 children with CH were missed by the NS. Five missed cases were children with athyreosis (35.8%), 8 had ectopic thyroid gland (57.1%) and one child had dyshormonogenesis (7.1%). The median age at diagnosis of CH in these missed cases was 1.12 years (range 0.08-13.5 years). In the group of children with missed diagnosis of CH, 71.4% were missed due to errors during sample collection, and 28.6% of patients were missed due to false negative screening results. Continuous education of neonatologists as well as nurses in neonatal units of maternity hospital are necessary to avoid errors in sample collection. It is of high importance to notice in a discharge list if a child is referred to the hospital before the sample is taken in a maternity hospital. On the other hand, the maternity hospital has to inform the screening laboratory about any child who was referred to another hospital if the blood samples for screening are not taken. Parents should be better informed that each child has to be included in the NS which is obligatory in Serbia. In addition, awareness amongst pediatricians in primary care should be raised regarding the possibility of missed cases of CH.

Background: A new device with metallic wires for scrape cytology was developed., Aims: To compare the diagnostic performance of scrape cytology and conventional cytology during endoscopic retrograde cholangiopancreatography for biliary strictures., Methods: A total of 420 cases with biliary stricture underwent transpapillary bile cytology. Among them, there are 79 cases with scrape cytology using the new device (scrape group) and 341 cases with conventional cytology (control group). Seventy-two and 174 cases underwent biliary biopsy at the same time as bile cytology in the scrape and control group, respectively., Results: The sensitivity for malignancy of bile cytology in the scrape and control group was 41.2% [pancreatic cancer (PC): 23.1%, biliary cancer (BC): 52.5%] and 27.1% (PC: 16.3%, BC: 38.0%), respectively (P = 0.023). When analyzed PC and BC, respectively, there was no significant difference between the two groups. In the both groups, the sensitivity was significantly higher for BC than PC. In the scrape group, there was no difference in the sensitivity between cytology and biopsy [39.7% (PC: 17.4%, BC: 55.3%)], but in the control group, a significantly lower sensitivity was observed with cytology than biopsy (36.4% (PC: 19.7%, BC: 50.0%)) (P = 0.046). When analyzed PC and BC, respectively, there was no significant difference between cytology and biopsy. The sensitivity of combined cytology and biopsy was 55.6% (PC: 30.4%, BC: 71.1%) in the scrape group and 47.0% (PC: 24.6%, BC: 64.3%) in the control group., Conclusion: Scrape bile cytology for biliary strictures may be superior to conventional cytology.

Optimal transportation of bacteria is important for accurate clinical interpretation, quantitative assays, and microbiome studies. A transport medium should ideally keep the bacteria alive without supporting growth or altering the relative proportions of the constituent species. We investigated the effect of nasal mucus and mucin on the growth and survival of two Staphylococcus aureus strains in liquid Amies transport medium at room temperature and 4 °C for 14 days. The study showed that the presence of nasal mucus in microbiological samples stimulated undesired S. aureus growth at room temperature in a dose-dependent manner. These findings underscore that microbiological samples from humans and animals should be stored at 4 °C until analysis to avoid undesired S. aureus growth., (Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Molecular research is increasingly dependent on high-quality biobanked biospecimens. Preanalytical variables in tissue processing and preservation may influence the RNA quality and research results. Hence, the effect of long-term storage and clinicopathological parameters on RNA quality needs to be elucidated., Methods: Ninety gastric cancer tissue samples were collected and fresh-frozen in a -80°C freezer for 12 years (2006-2017). The histology was assessed and RNA integrity number (RIN) was detected by an Agilent 2100 Bioanalyzer. The impact of storage duration on RNA integrity and histomorphology was analyzed. The difference between RIN values and clinical variables was analyzed. Correlations between pathological parameters such as tumor cell percentage, stroma percentage, necrosis extent, cellularity, and RIN were assessed, respectively., Results: Long-term storage at -80°C for 12 years did not adversely affect RNA integrity and histomorphology. RNA integrity was also not influenced by tumor location, estimated blood loss, cold/warm ischemia time, and surgical approach. However, RIN values were significantly correlated with the tumor cell percentage and stroma percentage. Gastric cancer tissues with higher tumor cell percentage or lower stroma percentage had higher RIN values., Conclusions: RNA quality of fresh-frozen gastric cancer tissues is influenced by clinical and histological parameters. Standard tissue collection procedure and histological quality control remain essential for tissue biobanking.

Introduction Surgical debridement of orthopaedic infections allows biopsy for microbiology and facilitates successful treatment. It is recommended that biopsy instruments are changed when taking multiple samples. This study compared assessed cross-contamination between biopsy sites when using same instruments to take tissue samples from multiple sites. Materials and methods During the surgical debridement, we defined five sampling sites and marked them with diathermy. Two sampling techniques were performed on same patient to minimise any potential bias arising from the type of host and the severity of infection. First, fresh instruments were used for each biopsy site. Titleond, the instruments used in the first sampling site were reused to take samples from the remaining sites. By comparing the microbiology results of the samples taken by each technique for each site we determined cross-contamination with microorganisms. Results Fifteen patients with foot and ankle infections (mean age 56 years) were included. Ten patients were diabetic and five had neuropathies. Cross-contamination between sampling sites occurred in eight cases when the same instruments were used to take biopsies (P = 0.002, Fisher's exact test). One or more microorganisms were involved in cross-contamination and the latter always occurred between two consecutive sites rather than sites that were further apart. Conclusion It is important to use fresh instruments for each biopsy site when taking multiple samples in musculoskeletal infection as cross-contamination might occur otherwise and affect microbiological studies.

Despite advancements in symptom management for heart failure (HF), this devastating clinical syndrome remains the leading cause of death in the developed world. Studies using animal models have greatly advanced our understanding of the molecular mechanisms underlying HF; however, differences in cardiac physiology and the manifestation of HF between animals, particularly rodents, and humans necessitates the direct interrogation of human heart tissue samples. Nevertheless, an ever-present concern when examining human heart tissue samples is the potential for artefactual changes related to temperature changes during tissue shipment or sample processing. Herein, we examined the effects of temperature on the post-translational modifications (PTMs) of sarcomeric proteins, the proteins responsible for muscle contraction, under conditions mimicking those that might occur during tissue shipment or sample processing. Using a powerful top-down proteomics method, we found that sarcomeric protein PTMs were differentially affected by temperature. Specifically, cardiac troponin I and enigma homolog isoform 2 showed robust increases in phosphorylation when tissue was incubated at either 4 °C or 22 °C. The observed increase is likely due to increased cyclic AMP levels and activation of protein kinase A in the tissue. On the contrary, cardiac troponin T and myosin regulatory light chain phosphorylation decreased when tissue was incubated at 4 °C or 22 °C. Furthermore, significant protein degradation was also observed after incubation at 4 °C or 22 °C. Overall, these results indicate that temperature exerts various effects on sarcomeric protein PTMs and careful tissue handling is critical for studies involving human heart samples. Moreover, these findings highlight the power of top-down proteomics for examining the integrity of cardiac tissue samples., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Salivary polyamines are potential non-invasive tools for screening various types of cancers. For clinical use, the reproducibility of these metabolites should be evaluated under various storage conditions, including duration and temperature, to establish standard operating protocols. Polyamines and amino acids in unstimulated whole saliva were quantified via liquid chromatography-mass spectrometry. Concentrations of time course samples were analysed after short-term storage for up to 240 min and long-term storage for up to 8 days under various storage conditions. As expected, storage at the lowest temperature (-18 °C) exerted the least pronounced effects on the quantified values in both tests. At a higher temperature, polyamines were more stable than amino acids, as evident from polyamine profiling. Addition of ethanol significantly stabilized polyamine profiles even at a higher temperature. Comparative processing of saliva revealed a minor effect of the solvent, whereas drying had a more prominent effect on polyamine profiles. Computational analyses evaluated the ability of polyamines to discriminate pancreatic cancer from controls. Repeated noise added tests were designed on the basis of the results of the storage tests; these analyses confirmed that the discriminative abilities were robust. These data contribute to the standardization of salivary storage conditions, thereby highlighting the clinical utility of saliva.

When sawing during autopsies on human remains, fine dust is produced, which consists of particles of sizes that may fall within the human respirable range, and can act as vectors for pathogens. The goal of this study was to explore the potential effects of saw blade frequency and saw blade contact load on the number and size of airborne bone particles produced. The methodology involved the use of an oscillating saw with variable saw blade frequencies and different saw blade contact loads on dry human femora. Released airborne particles were counted per diameter by a particle counter inside a closed and controlled environment. Results corroborated with the hypotheses: higher frequencies or lower contact loads resulted in higher numbers of aerosol particles produced. However, it was found that even in the best-case scenario tested on dry bone, the number of aerosol particles produced was still high enough to provide a potential health risk to the forensic practitioners. Protective breathing gear such as respirators and biosafety protocols are recommended to be put into practice to protect forensic practitioners from acquiring pathologies, or from other biological hazards when performing autopsies., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Objective: This report presents a summary of the outcomes achieved at ART center FERTILAB in Buenos Aires, Argentina, with transport IVF-ICSI for 25 years (1990-2014)., Methods: This report included all patients submitted to oocyte retrieval for IVF-ICSI whose oocytes were transported from sites of aspiration located 0.5-58 kilometers away from the central laboratory. The numeric data herein reported were taken from annual reports submitted by our institution (Fertilab) and, for purposes of comparison, by all Argentinian centers to the Latin American Registry of Assisted Reproduction (RLA) within the same time period., Results: From 1990 to 2014, 5091 aspirations followed by oocyte transport were performed in our center, resulting in 1258 pregnancies after fresh embryo transfers. The mean pregnancy/aspiration rate for the 25-year period was 24.71%. To validate the efficacy of our transport system, our results were compared to the outcomes of Argentinian centers reporting to RLA Argentina in the period ranging from 1990 to 2014. A total of 79,062 aspirations were performed, yielding 20,047 pregnancies and a pregnancy/aspiration rate of 25.36%. Delivery/aspirations rates were 15.34% for Fertilab patients and 14.79% for RLA Argentina centers., Conclusion: The results showed that the differences in clinical outcomes between our center and the bulk of Argentinian centers were not statistically significant, indicating that oocyte transport does not decrease the effectiveness of IVF-ICSI and might be advantageous under certain circumstances.

Recently, the use of metabolomic analysis of human serum and plasma for biomarker discovery and disease diagnosis in clinical studies has been increasing. The feasibility of using a metabolite biomarker for disease diagnosis is strongly dependent on the metabolite's stability during pre-analytical blood processing procedures, such as serum or plasma sampling and sample storage prior to centrifugation. However, the influence of blood processing procedures on the stability of metabolites has not been fully characterized. In the present study, we compared the levels of metabolites in matched human serum and plasma samples using gas chromatography coupled with mass spectrometry and liquid chromatography coupled with mass spectrometry. In addition, we evaluated the changes in plasma metabolite levels induced by storage at room temperature or at a cold temperature prior to centrifugation. As a result, it was found that 76 metabolites exhibited significant differences between their serum and plasma levels. Furthermore, the pre-centrifugation storage conditions significantly affected the plasma levels of 45 metabolites. These results highlight the importance of blood processing procedures during metabolome analysis, which should be considered during biomarker discovery and the subsequent use of biomarkers for disease diagnosis., (Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.)