Clinical challenges of tissue preparation for spatial transcriptome.

Spatial transcriptomics is considered as an important part of spatiotemporal molecular images to bridge molecular information with clinical images. Of those potentials and opportunities, the excellent quality of human sample preparation and handling will ensure the precise and reliable information generated from clinical spatial transcriptome. The present study aims at defining potential factors that might influence the quality of spatial transcriptomics in lung cancer, para-cancer, or normal tissues, pathological images of sections and the RNA integrity before spatial transcriptome sequencing. We categorised potential influencing factors from clinical aspects, including patient selection, pathological definition, surgical types, sample harvest, temporary preservation conditions and solutions, frozen approaches, transport and storage conditions and duration. We emphasis on the relationship between the combination of histological scores with RNA integrity number (RIN) and the unique molecular identifier (UMI), which is determines the quality of of spatial transcriptomics; however, we did not find significantly relevance between them. Our results showed that isolated times and dry conditions of sample are critical for the UMI and the quality of spatial transcriptomic samples. Thus, clinical procedures of sample preparation should be furthermore optimised and standardised as new standards of operation performance for clinical spatial transcriptome. Our data suggested that the temporary preservation time and condition of samples at operation room should be within 30 min and in 'dry' status. The direct cryo-preservation within OCT media for human lung sample is recommended. Thus, we believe that clinical spatial transcriptome will be a decisive approach and bridge in the development of spatiotemporal molecular images and provide new insights for understanding molecular mechanisms of diseases at multi-orientations.

Predictive Value of Swab Cultures for Cryopreserved Flaps During Delayed Cranioplasties.

OBJECTIVE: To assess the predictive value of swab cultures of cryopreserved skull flaps during cranioplasties for surgical site infections (SSIs). METHODS: A retrospective review was conducted of consecutive patients who underwent delayed cranioplasties with cryopreserved autografts between 2009 and 2017. The results of cultures obtained from swabs and infected surgical sites were assessed. The accuracy, sensitivity, and specificity of swab cultures for SSIs were evaluated. RESULTS: The study included 422 patients categorized into two groups, swab and nonswab, depending on whether swab cultures were implemented during cranioplasties. The overall infection rate was 7.58%. No difference was seen in infection rates between groups. There were 18 false-positive and no true-positive swab culture results. All bacteria between swab cultures and SSI cultures were discordant. Meanwhile, there were 19 false-negative swab cultures. The results showed high specificity but low sensitivity for swab cultures to predict SSI occurrence and the pathogens. CONCLUSIONS: Owing to low accuracy and sensitivity, swab cultures of cryopreserved autografts should not be routinely performed during delayed cranioplasties.

The appropriate number of preoperative core needle biopsy specimens for analysis in breast cancer.

ABSTRACT: Ultrasound (US)-guided core needle biopsy (CNB) has been recognized as a crucial diagnostic tool for breast cancer. However, there is a lack of guidance for hospitals that are not equipped with adjunctive US. The aim of this study was to assess the sensitivity, specificity, and experience of freehanded CNB in the outpatient department, and to determine the minimum number of tissue strips required to obtain concordance for estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor-2 (HER2), and tumor grade with the excised specimen.A prospective study was performed on 95 patients undergoing CNB and subsequent surgical procedures. The reliability of immunohistochemical assessments of the pathological type, tumor grade, ER, PR, and HER2 status in CNBs was compared with that of surgical specimens. Concordance between the CNBs and surgical samples was estimated as a percentage agreement, and analyzed using the chi-square test. A P < .05 was considered significant.The concordance rates of ER, PR, and HER2 status and tumor grade status between CNBs and surgically excised specimens were 97.9%, 91.6%, 82.1%, and 84.2%, respectively. The reliability of taking 2 tissue strips was similar to that of taking six tissue strips in distinguishing malignancy from benignancy, and determining the pathological type without the aid of US. Four tissue strips obtained by CNB showed good accuracy comparable to those obtained by surgical specimens in assessing ER, PR, and HER2 status and tumor grade.Two tissue strips obtained by CNB showed good accuracy in differentiating malignancy from benignancy, while at least 4 strips are recommended to obtain overall conformity of pathological biomarkers.

Rising incidence of appendiceal neoplasms over time: Does pathological handling of appendectomy specimens play a role?

BACKGROUND: Appendectomy is the most common emergent surgical procedure. Primary appendiceal neoplasms are rare entities that are usually detected incidentally in less than 2% of all appendectomies. The increase in the incidence rates of appendiceal neoplasms over time raises the question whether there is an actual change in the disease occurrence or is it a matter of increased recognition and reporting of what would have been previously missed and undiagnosed. OBJECTIVES: In our study, we aimed to review the archived tissue specimens of patients who were diagnosed with appendiceal neoplasms during the past decade at our institution and compare our clinical experience with published data to identify possible reasons that contribute to the increase in incidence rates of such neoplasms over the past few years. METHODS: Using a pathological database of surgical specimens from patients who underwent appendectomies between January 01, 2010 and September 30, 2020 at a large academic medical center, a single-center retrospective cohort analysis was performed, and medical charts of patients were reviewed. RESULTS: Of the total 1568 patients included, 102 (6.5%) had appendiceal neoplasms divided between primary (79.4%) and secondary/metastatic (20.6%) neoplasms. Annual incidence of appendiceal neoplasms over the past 10 years in our institution demonstrated an increasing trend from 5.6% in 2010 to 12.7% in 2020, which we hypothesize might be attributed to submitting more representative sections of the appendix for pathological examination than we had previously. Our results also showed that 2.8% of patients initially presenting with a typical clinical picture of acute appendicitis had appendiceal neoplasms as a truly incidental finding, while 20.3% of patients who underwent elective appendectomies for a suspicious appendiceal mass were found to be neoplastic. Interestingly, among the 80 cases of epithelial neoplasms, more non-carcinoid neoplasms were detected than carcinoid tumors. CONCLUSION: Based on our results and what has been published recently, we confirm an additional increase in incidental appendiceal neoplasms found in appendectomies performed for a clinical picture of acute appendicitis, which may be related to more thorough specimen assessment. Whether this is clinically impactful remains to be determined. However, these data support a modification in the way appendectomy specimens are handled in pathology labs post-operatively.

A review of recent advances in microsampling techniques of biological fluids for therapeutic drug monitoring.

Conventional sampling of biological fluids often involves a bulk quantity of samples that are tedious to collect, deliver and process. Miniaturized sampling approaches have emerged as promising tools for sample collection due to numerous advantages such as minute sample size, patient friendliness and ease of shipment. This article reviews the applications and advances of microsampling techniques in therapeutic drug monitoring (TDM), covering the period January 2015 - August 2020. As whole blood is the gold standard sampling matrix for TDM, this article comprehensively highlights the most historical microsampling technique, the dried blood spot (DBS), and its development. Advanced developments of DBS, ranging from various automation DBS, paper spray mass spectrometry (PS-MS), 3D dried blood spheroids and volumetric absorptive paper disc (VAPD) and mini-disc (VAPDmini) are discussed. The volumetric absorptive microsampling (VAMS) approach, which overcomes the hematocrit effect associated with the DBS sample, has been employed in recent TDM. The sample collection and sample preparation details in DBS and VAMS are outlined and summarized. This review also delineates the involvement of other biological fluids (plasma, urine, breast milk and saliva) and their miniaturized dried matrix forms in TDM. Specific features and challenges of each microsampling technique are identified and comparison studies are reviewed.

Pre-Analytical Sampling and Storage Conditions of Amyloid-ß Peptides in Venous and Capillary Blood.

Previous studies on blood-based biomarkers for Alzheimer's disease suggest a less invasive blood test might be a valuable screening tool for Alzheimer-specific pathology. Pre-analytical sample storage conditions seem to play an important role on amyloid-ß (Aß) stability, impacting reliability and reproducibility. This study shows that Aß40, Aß42, and Aß42/40 levels significantly and early decrease during storage at room temperature in whole blood or plasma. Storing blood samples at 4°C leads to stable Aß peptide concentrations up to 72 h. In addition, Aß peptides can be measured in capillary blood with a stable Aß42/40 ratio up to 72 h at 4°C.

[Veterinary pathological collections: yesterday - today - tomorrow]. / Veterinärpathologische Sammlungen: Gestern ­ Heute ­ Morgen.

INTRODUCTION: History, relevance and development of veterinary pathological collections are presented by analyzing and comparing the collections from Berlin, Munich, Vienna and Zurich of the 19th and 20th century. The indices of the collections are analyzed according to the frequency of animal species, body parts, organs and disease processes or etiologies respectively. Collection differences allow to draw conclusions on the founder of the collection and historical significance. Each collection was part of a university and thus involved in teaching and research. This often ensured the continuous existence of the collections. Nevertheless, changing teaching methods made pathological collections increasingly redundant. A comparison with other university collections, such as those of the University of Zurich, show new application aspects for existing collections and required measurement are discussed.

INTRODUCTION: En analysant et en comparant les collections pathologiques vétérinaires du 19e et 20e siècle de Berlin, Munich, Vienne et Zurich, on illustre l'histoire, la signification et le développement de ces collections. Les catalogues des collections sont analysés par rapport à la fréquence des espèces animales, des parties du corps ou d'organes et des maladies respectivement des étiologies. Les différences permettent des conclusions quant au créateur de la collection et aux circonstances temporelles de la création de la collection. Chacune des collections examinées faisait partie d'une université et étaient donc liée à l'enseignement et à la recherche. Cela a souvent assuré la pérennité des collections. Les changements dans l'enseignement universitaire ont rendu les collections de plus en plus superflues. Une comparaison avec d'autres collections universitaires telles que celles de l'Université de Zurich montre de nouveaux aspects d'utilisation des collections existantes. Les mesures nécessaires pour cela sont discutées.

The COVID-19 pandemic: implications for the cytology laboratory.

The coronavirus disease 2019 (COVID-19) is a pandemic caused by the SARS-CoV-2 virus. The infection has predominantly respiratory transmission and is transmitted through large droplets or aerosols, and less commonly by contact with infected surfaces or fomites. The alarming spread of the infection and the severe clinical disease that it may cause have led to the widespread institution of social distancing measures. Because of repeated exposure to potentially infectious patients and specimens, health care and laboratory personnel are particularly susceptible to contract COVID-19. This review paper provides an assessment of the current state of knowledge about the disease and its pathology, and the potential presence of the virus in cytology specimens. It also discusses the measures that cytology laboratories can take to function during the pandemic, and minimize the risk to their personnel, trainees, and pathologists. In addition, it explores potential means to continue to educate trainees during the COVID-19 pandemic.

Fast prostate retrieval in robot-assisted laparoscopic prostatectomy for next-generation biobanking.

Robotic-assisted laparoscopic radical prostatectomy (RALP) has become the most widespread treatment for organ-confined prostate cancer. Here, we describe a fast specimen retrieval technique for RALP to obtain high-quality tissue specimen with minimal warm ischemia time for next-generation biobanking. Here, we show that using fast retrieval technique, short warm ischemia times can be achieved while not increasing the surgical time. Patients undergoing RALP with written informed consent participated in Helsinki Urological Bank study. Previously operated RALP patients and those, who were not willing to participate in the study, served as a control group. The study consisted of 1685 patients, 684 in fast retrieval and 1001 in control group. We developed a novel fast retrieval technique in which fascia is opened for camera port according to the prostate size and a running suture is placed and tightened against the camera port in the beginning of the operation. Immediately after prostate is freed from attachments, suture is loosened and the prostate is extirpated inside the endoscopic bag through the camera port fascial opening, then the fascial suture is again tightened against the camera port and the RALP procedure is completed. The mean warm ischemia times in fast retrieval group were 20 min 18 s and 22 min 30 s, respectively, in patients without and with lymphadenectomy. The mean console and surgery times with and without lymphadenectomy were similar in both groups. There were no technique-related complications associated with Fast Retrieval procedure. Tissue integrity test results for the RNA and DNA quality showed good quality for the specimen. Fast retrieval technique can easily and safely be utilized to maximize usefulness of RALP tissue specimen in downstream biobank applications.

Procurement and Storage of Surgical Biospecimens.

A biobank is an important nexus between clinical and research aspects of pathology. The collection and storage of high quality surgical samples is essential for diagnosis post-surgery, and can also be used to create vaccines, identify therapeutic targets or establish eligibility of cancer patients in a clinical trial. Therefore, personnel handling surgical tissues should follow standard operating procedures (SOP) to maximize efficiency and preserve tissue quality. This chapter is intended to familiarize novice biobank personnel with the issues associated with different steps of surgical tissue collection including patient consent, sample collection, tissue storage, quality control, and distribution.

Autopsy Biobanking: Biospecimen Procurement, Integrity, Storage, and Utilization.

An autopsy is a specialized surgical procedure consisting of external and internal examination of a deceased individual for the purposes of documenting abnormalities and determining or confirming medical diagnoses that may have contributed to their death. One of the benefits of an autopsy is the opportunity to collect and store biospecimens for the purposes of biobanking. This chapter outlines the procedures necessary to procure, store, and utilize biospecimens obtained during an autopsy. With the emergence of molecular diagnostics, this chapter also discusses factors that influence the integrity of autopsy biospecimens prior to procurement. These include the postmortem interval, as well as premortem factors such as the patient's agonal state, biospecimen temperature, and pH.

Evidence of seasonal variation of ethyl glucuronide in hair: Modeling a seven-year data series.

The assessment of chronic excessive alcohol consumption by ethyl glucuronide (EtG) determination in hair is generally based on a cut-off value of 30 pg/mg recognized by regulatory authorities and scientific societies that guide the decision process. The ongoing debate about the risks connected with the straightforward application of this cut-off refers to the factors that may influence the detected EtG concentration. The present contribution to this debate evaluates the seasonal variation of the averaged EtG values along a seven-year period. Over 65 000 data points have been statistically analyzed to provide a mathematical model that interprets the data, gives insight into several influencing factors, and forecasts progressive data-points of the time series. This model shows that there is an annual pattern in the data exhibiting lower EtG concentrations during warm seasons and higher values in cold seasons. The estimated EtG cycles are characterized by the seasonal variation of ±2.78 pg/mg above and below the overall mean (with 5.56 pg/mg absolute difference overall). This seasonal factor associated with EtG quantification might result in a potential source of bias, at least in the regional/climatic conditions observed in the samples' collection area. Moreover, the EtG time series reveals that the change in the sample pre-treatment procedure has an effect on the modeled pattern as an abrupt increment (+38%) in the mean value of the EtG concentration. This change corresponds to the time when the former protocol of cutting hair into small segments before extraction was substituted by pulverization with a ball mill.

Recent trends in sorption-based sample preparation and liquid chromatography techniques for food analysis.

The accelerated rising of the world's population increased the consumption of food, thus demanding more rigors in the control of residue and contaminants in food-based products marketed for human consumption. In view of the complexity of most food matrices, including fruits, vegetables, different types of meat, beverages, among others, a sample preparation step is important to provide more reliable results when combined with HPLC separations. An adequate sample preparation step before the chromatographic analysis is mandatory in obtaining higher precision and accuracy in order to improve the extraction of the target analytes, one of the priorities in analytical chemistry. The recent discovery of new materials such as ionic liquids, graphene-derived materials, molecularly imprinted polymers, restricted access media, magnetic nanoparticles, and carbonaceous nanomaterials, provided high sensitivity and selectivity results in an extensive variety of applications. These materials, as well as their several possible combinations, have been demonstrated to be highly appropriate for the extraction of different analytes in complex samples such as food products. The main characteristics and application of these new materials in food analysis will be presented and discussed in this paper. Another topic discussed in this review covers the main advantages and limitations of sample preparation microtechniques, as well as their off-line and on-line combination with HPLC for food analysis.

Cryopreservation of Fish Spermatogonial Cells: The Future of Natural History Collections.

As global biodiversity declines, the value of biological collections increases. Cryopreserved diploid spermatogonial cells meet two goals: to yield high-quality molecular sequence data; and to regenerate new individuals, hence potentially countering species extinction. Cryopreserved spermatogonial cells that allow for such mitigative measures are not currently in natural history museum collections because there are no standard protocols to collect them. Vertebrate specimens, especially fishes, are traditionally formalin-fixed and alcohol-preserved which makes them ideal for morphological studies and as museum vouchers, but inadequate for molecular sequence data. Molecular studies of fishes routinely use tissues preserved in ethanol; yet tissues preserved in this way may yield degraded sequences over time. As an alternative to tissue fixation methods, we assessed and compared previously published cryopreservation methods by gating and counting fish testicular cells with flow cytometry to identify presumptive spermatogonia A-type cells. Here we describe a protocol to cryopreserve tissues that yields a high percentage of viable spermatogonial cells from the testes of Asterropteryx semipunctata, a marine goby. Material cryopreserved using this protocol represents the first frozen and post-thaw viable spermatogonial cells of fishes archived in a natural history museum to provide better quality material for re-derivation of species and DNA preservation and analysis.

Diabetes-Associated Biobanking: More Topical Than Ever?

The increasing prevalence of diabetes and its accompanying long-term complications, as well as the associated economic burden, calls for a rapid clinical translation of biomedical research to better valid the physiological relevance of the findings from basic research. To meet this condition, the Collaborative Research Center (CRC) 1118 has established the first nationwide diabetes-specific Biomaterialbank (BMB) that permanently preserves solid and liquid specimen retro- and prospectively at the Institute of Pathology and Department of Endocrinology of the University Hospital Heidelberg. The main purpose of this BMB is to collect, preserve, characterize and provide human diabetic specimen to researchers investigating the role of reactive metabolites (RM) as cause of diabetic late complications. In this review we discuss the urgent need to support translational and clinical research projects by making use of diabetic solid and liquid specimen and provide an insight into the organization and general conditions of biobanking procedures which are pivotal to guaranteeing high-quality human biomaterial. In light of diabetes-tailored biobanking, we describe our newly initiated activities and introduce the diverse technology platforms that can be used for the investigation of promising molecular targets pertinent for diabetes. With this article we demonstrate that the preservation of rare specimen is also particularly relevant in the non-neoplastic field and contributes to basic investigation, promotes comprehensive scientific data and fortifies the sustainability for diabetes research. In addition, the increased understanding of how metabolic imbalance triggers diabetes onset and progression and favors diabetic late symptoms might hold some promise for future innovative diagnostic and/ or therapeutic applications, eventually adding to the improvement of patient care.