Critical Factors in Sample Collection and Preparation for Clinical Metabolomics of Underexplored Biological Specimens.

This review article compiles critical pre-analytical factors for sample collection and extraction of eight uncommon or underexplored biological specimens (human breast milk, ocular fluids, sebum, seminal plasma, sweat, hair, saliva, and cerebrospinal fluid) under the perspective of clinical metabolomics. These samples are interesting for metabolomics studies as they reflect the status of living organisms and can be applied for diagnostic purposes and biomarker discovery. Pre-collection and collection procedures are critical, requiring protocols to be standardized to avoid contamination and bias. Such procedures must consider cleaning the collection area, sample stimulation, diet, and food and drug intake, among other factors that impact the lack of homogeneity of the sample group. Precipitation of proteins and removal of salts and cell debris are the most used sample preparation procedures. This review intends to provide a global view of the practical aspects that most impact results, serving as a starting point for the designing of metabolomic experiments.

Evaluation of four quality indicators of the Pre-Analytical Phase External Quality Assessment Subprogram of the Fundación Bioquímica Argentina.

Pre-analytical phase external quality assessment programs contribute - through the interlaboratory comparison of quality indicators (QIs) - to the continuous improvement of the clinical laboratory total testing process. The purpose of the present work is to document the results derived from measuring four QIs within the framework of a pre-analytical phase external quality assessment subprogram in Argentina. The laboratories participating in this subprogram measured the following QIs: i) patients recalled for a new blood sample collection due to pre-analytical causes; ii) clotted samples from hemogram and coagulation tests; iii) clinical chemistry hemolyzed samples; and iv) requests with transcription errors entered into the laboratory information system. Results were expressed in percentage value and Sigma value. Databases were anonymized. A minimum acceptable quality level for the four QIs measured was recorded in the majority (75%) of the participating laboratories (Sigma > 3.0). It was nonetheless observed that the QIs of hemolyzed samples and requests with transcription errors entered into the laboratory information system deserve more attention. Through this pioneering experience in Argentina, the participating laboratories - some for the first time - could learn about their performance via interlaboratory comparison of results. This experience also proved to be motivating not only to improve the external assessment subprogram but also to continue working on the measurement of pre-analytical QIs for the continuous improvement of the clinical laboratory total testing process in Argentina.

Coffee intake one hour prior to phlebotomy produces no clinically significant changes in routine biochemical test results.

Introduction: Although current guidelines recommend not drinking coffee prior to phlebotomy, our hypothesis is that drinking coffee does not affect the clinical interpretation of biochemical and haematological test results. Materials and methods: Twenty-seven volunteers were studied in basal state (T0) and 1h after (T1) drinking coffee. Routine haematological (Sysmex-XN1000 analyser) and biochemistry parameters (Vitros 4600 analyser) were studied. Results were compared using the Wilcoxon test (P < 0.05). A clinical change was considered when mean percent difference (MD%) was higher than the reference change value (RCV). Results: Coffee intake produced statistically, but not clinically, significant: i) increases in haemoglobin (P = 0.009), mean cell haemoglobin concentration (P = 0.044), neutrophils (P = 0.001), albumin (P = 0.001), total protein (P = 0.000), cholesterol (P = 0.025), high density lipoprotein cholesterol (P = 0.007), uric acid (P = 0.011), calcium (P = 0.001), potassium (P = 0.010), aspartate aminotransferase (P = 0.001), amylase (P = 0.026), and lactate dehydrogenase (P = 0.001), and ii) decreases in mean cell volume (P = 0.002), red cell distribution width (P = 0.001), eosinophils (P = 0.002), and lymphocytes (P = 0.001), creatinine (P = 0.001), total bilirubin (P = 0.012), phosphorus (P = 0.001), magnesium (P = 0.007), and chloride (P = 0.001). Conclusion: Drinking a cup of coffee 1 hour prior to phlebotomy produces no clinically significant changes in routine biochemical and haematological test results.

Effects of pre-analytical sample care and analysis methodology on measures of metabolic acidosis in hemodialysis patients.

INTRODUCTION: We evaluated the effects of pre-analytical care on total carbon dioxide (tCO2 ) in hemodialysis patients, as calculated by blood gas analysis (ctCO2 ) or measured by an enzymatic assay (mtCO2 ). METHODS: Blood samples were collected via vascular access before dialysis sessions. For blood gas analysis, eight aliquots were collected, refrigerated or non-refrigerated, and analyzed at 0, 4, 8, and 24 h after collection. A blood sample was then collected for the enzymatic method and distributed into 14 aliquots. Half of the aliquots were refrigerated. The samples analyzed at time point 0 were centrifuged immediately. The remaining aliquots of both the refrigerated and non-refrigerated clusters were centrifuged before storage. Samples were analyzed at 4, 8, and 24 h post-collection. FINDINGS: By blood gas analysis, no significant change was found in bicarbonate values over time, either in the non-refrigerated or refrigerated samples. ctCO2 values during the experiment showed a minor but statistically significant increase of questionable clinical relevance in both non-refrigerated and refrigerated aliquots. In the enzymatic assay, the reduction in mtCO2 levels during the experiment was negligible. The median absolute reductions at the end of the experiment were 1.77, 1.21, 1.04, and 1.12 mmol/L for the non-centrifuged/non-refrigerated, centrifuged/non-refrigerated, non-centrifuged/refrigerated, and centrifuged/refrigerated aliquots, respectively. DISCUSSION: Our results suggest that measured or calculated tCO2 levels of capped and cooled samples are adequate for analyzing the acid-base status of hemodialysis patients, even when such determination is not performed immediately after collection.

Conocimiento sobre la valoración de hematíes dismórficos en sedimento urinario en profesionales de laboratorio clínico de Lima metropolitana / Knowledge about the assessment of dysmorphic red blood cells in urinary sediment in clinical laboratory professionals in metropolitan Lima

Introducción: A pesar de la existencia de directrices internacionales para la correcta valoración de la hematuria dismórfica, es frecuente que su categorización presente mucha variabilidad, inclusive entre los profesionales del mismo laboratorio. Objetivo: Determinar el nivel de conocimiento de profesionales tecnólogos médicos para la valoración de hematíes dismórficos en sedimento urinario. Material y Métodos: Estudio transversal observacional. Se aplicó un instrumento de medición para evaluar el conocimiento sobre la fase preanalítica y analítica de la valoración de hematíes dismórficos en sedimento urinario a 113 profesionales con más de tres años de experiencia en el área de uroanálisis y que laboran en centros hospitalarios estatales y privados. Resultados. La media del puntaje obtenido fue de 5,4±1,9 de 10 puntos posibles; El 85,0% de los encuestados presento un conocimiento medio e insuficiente y solo el 15,0% un conocimiento avanzado; Se observo asociación entre los niveles de conocimiento y años de experiencia (p=0,006), no se evidenció asociación significativa con la variable sexo (p=0,791) y sedes hospitalarias (p=0,888). Discusión: Se evidenció un bajo conocimiento en los dominios preanalíticos y analíticos para la valoración de hematíes dismórficos, además de una asociación significativa entre el nivel de conocimiento y los años de experiencia del profesional.

Background: Despite the existence of international guidelines for the correct assessment of dysmorphic hematuria, its categorization frequently presents a lot of variability, even among professionals in the same laboratory. Objective: To determine the level of knowledge of professional medical technologists for the assessment of dysmorphic red blood cells in urinary sediment. Material and Methods: Observational cross-sectional study. Ameasurement instrument was applied to assess knowledge about the preanalytical and analytical phase of the assessment of dysmorphic red blood cells in urinary sediment to 113 professionals with more than three years of experience in the area of urinalysis and who work in state and private hospitals. Results.The mean score obtained was 5.4±1.9 out of 10 possible points; 85.0% of the respondents presented medium and insufficient knowledge and only 15.0% advanced knowledge; An association was observed between levels of knowledge and years of experience (p=0.006), no significant association was found with the variable gender (p=0.791) and hospital locations (p=0.888). Discussion: Low knowledge was evidenced in the preanalytical and analytical domains for the assessment of dysmorphic red blood cells, in addition to a significant association between the level of knowledge and the years of experience of the professional.

A Simple Yet Effective Preanalytical Strategy Enabling the Application of Aptamer-Conjugated Gold Nanoparticles for the Colorimetric Detection of Antibiotic Residues in Raw Milk.

The misuse of antibiotics in the cattle sector can lead to milk contamination, with concomitant effects on the dairy industry and human health. Biosensors can be applied in this field; however, the influence of the milk matrix on their activity has been poorly studied in light of the preanalytical process. Herein, aptamer-conjugated gold nanoparticles (nanoaptasensors) were investigated for the colorimetric detection in raw milk of four antibiotics used in cattle. The effect of milk components on the colorimetric response of the nanoaptasensors was analyzed by following the selective aggregation of the nanoparticles, using the absorption ratio A520/A720. A preanalytical strategy was developed to apply the nanoaptasensors to antibiotic-contaminated raw milk samples, which involves a clarification step with Carrez reagents followed by the removal of cations through dilution, chelation (EDTA) or precipitation (NaHCO3). The colorimetric signals were detected in spiked samples at concentrations of antibiotics as low as 0.25-fold the maximum residue limits (MRLs) for kanamycin (37.5 µg/L), oxytetracycline (25 µg/L), sulfadimethoxine (6.25 µg/L) and ampicillin (1 µg/L), according to European and Chilean legislation. Overall, we conclude that this methodology holds potential for the semiquantitative analysis of antibiotic residues in raw milk obtained directly from dairy farms.

Storage Duration Affects the Quantification of Oxidative Stress Markers in the Gastrocnemius, Heart, and Brain of Mice Submitted to a Maximum Exercise.

This study investigated the effect of sample storage duration on the quantification of oxidative stress markers in the gastrocnemius, heart, and brain of mice submitted to a maximum swimming exercise. Thiobarbituric acid reactive substances (TBARSs), protein carbonyl derivatives, total antioxidant capacity (TAC), and the activity of superoxide dismutase (SOD) and catalase (CAT) were quantified in fresh tissues and in samples stored at -80°C for 1, 3, or 6 months, from exercised (n = 13) and nonexercised mice (n = 13). Except for protein carbonyl derivatives in the heart, the exercise resulted in the modification of all markers in all fresh-evaluated samples (p < 0.001). The storage duration did not modify the effect of exercise on protein carbonyl derivatives and TAC. TBARS was stable for 3 months in the gastrocnemius and for 1 month in frozen heart and brain. Accordingly, the exercise effect on TBARS levels observed in fresh samples was absent in the gastrocnemius frozen for 6 months (p = 0.98) and in the heart and brain frozen for 3 months (p = 0.07 and 0.28, respectively) or more (p = 0.21 for heart and p > 0.99 for brain). In addition, CAT and SOD activities were reduced by storage duration in all tissues evaluated (p < 0.05). Our findings show that sample storage duration alters the quantification of oxidative stress markers in mice submitted to maximum exercise, and its effect is tissue and marker dependent. Some recommendations to achieve more accurate and reproducible data in the exercise physiology and oxidative stress markers field are presented.

Errores preanalíticos en la recolección de orina de 24 horas / Preanalytical errors in 24-hour urine collection

RESUMEN Introducción. La recolección de orina en el lapso de 24 horas es necesaria para la medición de constituyentes bioquímicos que presentan una excreción urinaria variable; sin embargo, dicho proceso no está exento de errores preanalíticos. Objetivos. Evaluar el nivel de cumplimiento de las instrucciones para recoger la muestra, identificar la recolección de orinas incompletas y conocer los procesos preanalíticos que aplica un grupo de profesionales. Métodos. Estudio de diseño observacional, en el que se realizó encuestas anónimas y voluntarias a 257 pacientes ambulatorios y 59 profesionales tecnólogos médicos, además de estimar la excreción urinaria de creatinina a 416 muestras. Resultados. El estudio evidenció recolecciones incorrectas (39,7%), uso de recipientes inadecuados (58,14%), mala conservación de las muestras (98,8%), cambio en los hábitos de ingesta de líquidos (23,7%), escasa información y retroalimentación a los pacientes sobre la preparación de la prueba, el 76,92% de muestras presentaron pérdidas o excesos en el volumen recolectado y una alta variabilidad de los criterios que utilizan los profesionales para rechazar las muestras mal colectadas. Conclusión. Se observó un bajo cumplimiento de los pacientes a las indicaciones para la recolección de la muestra, una alta proporción de recolecciones incompletas y discordancia en los procesos preanalíticos para el análisis de orina de 24 horas.

ABSTRACT Introduction. Urine collection within 24 hours is necessary for the measurement of biochemical constituents with variable urinary excretion, however, this process is not free of preanalytical errors. Objectives. To evaluate the level of compliance with the instructions for sample collection, identify the collection of incomplete urine and know the pre-analytic processes applied by a group of professionals. Methods. Observational design study, in which anonymous and voluntary surveys were conducted with 257 outpatients and 59 Medical Technologist professionals, in addition to estimating the urinary excretion of creatinine in 416 samples. Results. The study evidenced incorrect collections (39.7%), use of inappropriate containers (58.14%), poor preservation of samples (98.8%), change in fluid intake habits (23.7%), little information and feedback to patients on the preparation of the test, 76.92% of samples presented losses or excesses in the volume collected and a high variability of the criteria used by professionals to reject poorly collected samples. Conclusion. Low compliance of the patients to the indications for sample collection, a high proportion of incomplete collections, and discordance in the preanalytical processes for the 24-hour urinalysis were observed.

Postprandial lipemia causes oxidative stress in dogs.

Oxidative stress (OS) has been strongly associated with postprandial lipemia (PPL) in humans, and still requires further investigation in dogs. However, since lipemia interferes with spectrophotometric determinations such as those used to assess OS, the present study investigated the effect of PPL on OS parameters of healthy dogs. Twenty dogs had lipemic postprandial samples compared to the average of two non-lipemic moments. Subsequently, PPL was simulated in vitro using a commercial lipid emulsion and twelve pools of non-lipemic serum of these dogs were used to simulate the minimum, median and maximum concentrations of triglycerides obtained during the lipemic state. Serum OS parameters were assessed using the antioxidants uric acid, albumin and total bilirubin; total antioxidant capacity (TAC); total oxidant capacity (TOC); and lipid peroxidation. In vivo PPL caused an increase in albumin, TAC-CUPRAC, TAC-FRAP, uric acid (p < 0.0001), TOC (p = 0.0012) and total bilirubin (p = 0.0245); reduction of TAC-ABTS (p = 0.0008); and did not alter the lipid peroxidation (p = 0.8983). In vitro, levels of albumin increased at the three lipemic concentrations (p < 0.0001), uric acid increased in the median and maximum levels (p < 0.0001), and total bilirubin concentration increased only at the maximum lipemic level (p = 0.0012). All lipemic levels tested increased TAC-ABTS (p = 0.0011) and TAC-FRAP (p < 0.0001). TAC-CUPRAC (p = 0.5002), TOC (p = 0.5938) and lipid peroxidation (p = 0.4235) were not affected by in vitro lipemia. In conclusion, both the in vivo postprandial state and in vitro simulated lipemia affect oxidative stress markers in dogs depending on the oxidative stress marker, and thus the postprandial state and/or lipemic samples should be avoided.

Workflow optimization in a clinical laboratory using Lean management principles in the pre-analytical phase.

BACKGROUND: The application of the Lean methodology in clinical laboratories can improve workflow and user satisfaction through the efficient delivery of analytical results. The purpose of this study was to optimise delivery times of the test results at a clinical laboratory, using Lean management principles in the pre-analytical phase. METHODS: A prospective study with a quasi-experimental design was implemented. Staff functions were restructured and sample flows were modified. Delivery times of clinical results (glucose and haematocrit; 6648 data) from the Medicine and Adult Emergency services for years 2017 and 2018 were compared. RESULTS: A reduction (p < 0.05) in turnaround times in the delivery of glucose test results at the adult emergency service was observed (84 to 73 min, 13%, pre and post). In addition, there was a non-significant reduction in the turnaround times for glucose (Medicine) and haematocrit in both services. In the analytical and post-analytical phase (not intervened), an increase in turnaround times was observed in some cases. CONCLUSIONS: Other studies have indicated that the application of the Lean methodology in clinical laboratories improves workflow, increasing effectiveness and efficiency. This study showed an improvement in the delivery time of test results (glucose - Emergency), giving rise to a culture of cooperation and continuous improvement. It would, however, be essential to address the management model integrating the analytical and post-analytical phases.

Monitoramento da qualidade da amostra para análise de RT-PCR da Covid-19 no LACEN-BA / Sample quality monitoring for Covid-19 RT-PCR analysis at LACEN-BA / Monitoreo de la calidad de la muestra para el análisis Covid-19 RT-PCR en LACEN-BA

A pandemia da Covid-19 tem se apresentado como um dos maiores desafios sanitários desse século. Em dezembro de 2019, na China, o agente etiológico foi identificado como um novo coronavírus, nomeado SARS-CoV-2. No Brasil, o primeiro caso confirmado da Covid-19 ocorreu em fevereiro de 2020 e, no mês seguinte, a Secretaria da Saúde do Estado da Bahia (Sesab) confirmou o primeiro caso na Bahia.O Laboratório Central de Saúde Pública Prof. Gonçalo Moniz (Lacen-BA) centralizou o diagnóstico laboratorial para confirmação dos casos suspeitos de Covid-19 dos 417 municípios baianos, utilizando a técnica de RT-PCR. Este estudo tem como objetivo identificar e analisar as não conformidades das amostras suspeitas de Covid-19 encaminhadas ao Lacen-BA. Trata-se de um estudo descritivo, cujos dados foram obtidos por meio de consulta aos relatórios de amostras e exames em desacordo, disponíveis no sistema Gerenciador de Ambiente Laboratorial (GAL), gerados mensalmente, no período de abril a outubro de 2020. Para garantir a qualidade das amostras recebidas, foram definidos critérios de aceitação/rejeição de amostras e criado o formulário de notificação de não conformidades, assegurando a rastreabilidade das amostras de Covid-19. Através de relatórios diários do sistema GAL, selecionou-se os nove principais motivos de não conformidades, sendo o mais frequente "requisição cancelada pela gerência do GAL devido à expiração do prazo de triagem", com 72,8% dos registros. A inserção da padronização de processos na etapa pré-analítica permite trabalhar com segurança, garantindo a qualidade da amostra a ser processada e, consequentemente, um resultado fidedigno, dentro do prazo acordado.

The Covid-19 pandemic is one of the greatest health challenges of this century. In December 2019, in China, the etiologic agent was identified as a new coronavirus, named SARS-CoV-2. In Brazil, the first case of Covid-19 was confirmed in February 2020 and, in the following month, the Department of Health of the State of Bahia (Sesab) confirms the first case in the state. The Central Public Health Laboratory Prof. Gonçalo Moniz (Lacen/BA) centralized the laboratory diagnosis to confirm the suspected cases of Covid-19 of the 417 municipalities of the state, using the RT-PCR technique. This study aims at identifying and analyzing the non-conformities of the suspected samples of Covid-19 sent to Lacen-BA. This is a descriptive study whose data were obtained by consulting there reports of samples and exams in disagreement, available in the Laboratory Environment Manager (GAL) system, generated monthly, from April to October,2020. To guarantee the quality of the samples received, acceptance / rejection criteria for the samples were defined and a form for the notification of non-conformities was created, ensuring the traceability of the Covid-19 samples. Daily reports from the Laboratory Environment Manager system based the selection of nine main reasons for non-conformities, among which "requisition canceled by the management of the GAL due to the expiration of the screening period" was present in 72.8% of the records. Process standardization, in the pre-analytical stage, allows working with security, guaranteeing the quality of the sample to be processed and a reliable result within the established period.

La pandemia del Covid-19 se ha presentado como uno de los desafíos de salud más grandes de este siglo. En diciembre de 2019, China identificó el agente etiológico del nuevo coronavirus llamado SARS-CoV-2. En Brasil, se notificó el primer caso del Covid-19 en febrero de 2020 y, al mes siguiente, la Secretaría de Salud del Estado de Bahía (Sesab) confirmaba el primer caso en Bahía. El Laboratorio Central de Salud Pública Prof. Gonçalo Moniz (Lacen/BA) centralizó el diagnóstico de laboratorio para confirmar los casos sospechosos del coronavirus de los 417 municipios de Bahía, mediante la técnica de RT-PCR. Este estudio tiene como objetivo identificar y analizar las no conformidades de las muestras sospechosas del Covid-19 enviadas al Lacen/BA. Este es un estudio descriptivo cuyos datos se obtuvieron consultando los informes de muestras y pruebas en desacuerdo disponibles en el sistema Laboratory Environment Manager (GAL), generados mensualmente, de abril a octubre/2020. Con el fin de garantizar la calidad de las muestras recibidas, se definieron criterios de aceptación/rechazo de las muestras y se elaboró un formulario para la notificación de no conformidades, asegurando la trazabilidad de las muestras. Por medio de informes diarios del sistema Laboratory Environment Manager, se seleccionaron nueve principales causas de no conformidades, de las cuales la más frecuente fue "requisición cancelada por la gerencia del GAL por vencimiento del período de cribado" con el 72,8% de los registros. La inserción de la estandarización de procesos en la etapa preanalítica permite trabajar con seguridad, garantizando la calidad de la muestra que procesar y, en consecuencia, un resultado confiable dentro del plazo acordado.

Brazilian laboratory indicators benchmarking program: three-year experience on pre-analytical quality indicators.

OBJECTIVES: In the laboratory medicine segment, benchmarking is the process in which institutions seek to compare with the macro environment (performance comparison and best practices with different laboratories) and improve their results based on quality indicators. The literature has highlighted the vulnerability of the pre-analytical phase in terms of risks and failures and the use of interlaboratory comparison as an opportunity to define a strategic performance benchmark aligned with the laboratory medicine sector, which has been a promising strategy to ensure continuous improvement, identifying within the pre-analytical process the critical activities to guarantee patient safety. In this context, this paper aims to present the three-year experience (2016-2018) of the Benchmarking Program and Laboratory Indicators - in Portuguese, Programa de Benchmarking e Indicadores Laboratoriais (PBIL) - with emphasis on pre-analytical indicators and their comparison against literature references and other programs of benchmarking in the area of laboratory medicine. PBIL is organized by the Brazilian Society of Clinical Pathology/Laboratory Medicine (SBPC/ML) in conjunction with Controllab and coordinated by a Brazilian group with representatives from different countries. METHODS: The data presented in this paper involving the performance results of 180 laboratories with active participation. Results are presented in percentage (%, boxplot graphical in quartiles) and Sigma metric, recognized as the metric that best indicates the magnitude of failures in a process. The Pareto Chart was used to facilitate ordering and to identify the main errors in the pre-analytical phase. The Radar Chart was made available in this work for the purpose of comparing the results obtained in Sigma by the PBIL and IFCC Working Group Laboratory Errors and Patient Safety (WG LEPS). RESULTS: In the study period, just over 80% of the pre-analytical failures are related to Blood culture contamination (hospital-based and non-hospital-based laboratories), Recollect and Non-registered exams, with failure rates of 2.70, 1.05 and 0.63%, respectively. The performance of the PBIL program participants was in line with the literature references, and allowed to identify benchmarks in the laboratory medicine market, target of PBIL, with best practices were observed for some indicators. CONCLUSIONS: The results of the program demonstrate the importance of an ongoing program comparative performance-monitoring program for setting more robust goals and consequently reducing laboratory process failures. Even with these promising premises and results, the contextualized analysis of the program indicators, point to a still significant number of failures in our market, with possibilities for improvement in order aiming to ensure more robust and effective processes.

Evaluation of the influence of pre-analytical factors on total testosterone levels in healthy young men / Avaliação da influência de fatores pré-analíticos nas dosagens de testosterona total em homens jovens saudáveis

ABSTRACT Introduction: Testosterone is the main hormone that regulates male reproductive functions, directly participating in spermatogenesis and increasing sexual activity. The blood measurement of this hormone is essential for the diagnosis of neuroendocrine disorders, such as hypogonadism. However, there is lack of standardization regarding patient preparation for the hormone collection in clinical laboratories. Objective: Evaluate the effect of pre-analytical variables, including fasting, circadian and seasonal variation on testosterone levels in healthy young males. Material and methods: Forty-two volunteers were selected for the study, in the city of Barbacena, Minas Gerais. Four blood samples were collected from each of the participants, three in the winter: the first one in the morning after fasting; the second in the afternoon, without fasting; the third, in the next day morning, without fasting; and the last one taken in the summer, in the morning, after fasting. Results: The analyses showed that there was a significant decrease in total testosterone levels when there was no fasting for eight hours prior to collection and in the afternoon compared to the morning, both with p < 0.001. There was no significant difference in the results obtained in winter and summer. Conclusion: It is recommended that clinical laboratories standardize the collection of total testosterone by performing the test in the morning and after an eight-hour fast, in order to reduce variability and ensure reliability in the results.

RESUMEN Introducción: La testosterona es la principal hormona reguladora de funciones reproductoras masculinas, participando directamente en la espermatogénesis y en el aumento de la actividad sexual. La medición sanguínea de esa hormona es fundamental en el diagnóstico de trastornos neuroendocrinos, como el hipogonadismo. Sin embargo, hay poca estandarización en la preparación adecuada del paciente para la recolección de la hormona en laboratorios clínicos. Objetivo: Evaluar el efecto de variables preanalíticas, incluyendo ayuno, variación circadiana y estacional en las mediciones de testosterona en hombres jóvenes sanos. Material y métodos: Se eligieron 42 voluntarios para el estudio, en la ciudad de Barbacena, Minas Gerais. Se tomaron cuatro muestras de sangre de cada uno de los participantes, de las cuales tres en invierno: la primera, matutina, en ayunas; la segunda, vespertina, sin ayunas; la tercera, el dia siguiente, matutina, sin ayunas; el última se recolectó en verano, por la mañana, en ayunas. Resultados: Los análisis demostraron que hubo reducción significativa en los niveles de testosterona total cuando no se realizó el ayuno de ocho horas antes de la recolección y en el período vespertino en comparación al matutino, ambos con valor de p < 0,001. No hubo diferencia significativa en los resultados obtenidos en invierno y en verano. Conclusión: Se recomienda que los laboratórios clínicos estandarizen la recolección de testosterona total con la realización de la prueba en el período matutino y en ayuno de ocho horas, para reducir la variación y garantizar la confiabilidad de los resultados.

RESUMO Introdução: A testosterona é o principal hormônio regulador das funções reprodutivas masculinas, participando diretamente da espermatogênese e do aumento da atividade sexual. A dosagem sanguínea desse hormônio é fundamental no diagnóstico de distúrbios neuroendócrinos, como o hipogonadismo. Entretanto, há pouca padronização no preparo adequado do paciente para a coleta do hormônio em laboratórios clínicos. Objetivo: Avaliar o efeito de variáveis pré-analíticas, incluindo realização de jejum, variação circadiana e sazonal nas dosagens de testosterona em jovens saudáveis do sexo masculino. Material e métodos: Foram selecionados 42 voluntários para o estudo, na cidade de Barbacena, Minas Gerais. Quatro amostras de sangue de cada um dos participantes foram coletadas, sendo três no inverno: a primeira de manhã, em jejum; a segunda à tarde, sem jejum; a terceira no dia seguinte, de manhã, sem jejum. A última foi coletada no verão, na parte da manhã, em jejum. Resultados: As análises demonstraram que houve diminuição significativa dos níveis de testosterona total quando não foi realizado jejum de 8 horas antes da coleta e no período da tarde em comparação ao período da manhã, ambos com valor de p < 0,001. Não houve diferença significativa nos resultados obtidos no inverno e no verão. Conclusão: Recomendamos que os laboratórios clínicos padronizem a coleta de testosterona total com a realização do exame no período da manhã e em jejum de 8 horas, a fim de reduzir a variabilidade e garantir a confiabilidade nos resultados.

Evaluation of urinalysis pre-analytical phase in a private laboratory of Maringá city, Paraná, Brazil / Avaliação da fase pré-analítica do exame de urina de rotina em laboratório privado da cidade de Maringá, Paraná, Brasil

ABSTRACT Introduction: Routine urinalysis is among the most requested exams in the clinical laboratory, assisting in the diagnosis of various diseases and treatment follow-up. In this case, the pre-analytical phase is extremely important because the quality of the sample directly influences the analysis and interpretation of the result. Objective: The aim of this study was to identify the main errors in the pre-analytical phase of routine urine examination in a private laboratory and their frequency of occurrence. Material and method: Data were collected from 2014 to 2018. In all, 107,277 urine samples were registered and 98 (0.09%) were sent for recollection. Results: Recollect requests were higher among females (81.6%), and the most affected age groups were 51 to 60 years old and 21 to 30 years old. The most common justification for recollection was insufficient material (48.0%), followed by confirmation of results in 24.5% of cases. The predominance of urine recollection in women was due to their having urine tests more often since they are more prone to urinary tract infections, especially in the sexually active and postmenopausal life stages. Conclusion: In general, the urine recollection rate obtained in the research was lower than the goal set by the laboratory; however, the main reasons that led to recollection request could be avoided or minimized if patients had been well educated on the correct collection procedures, indicating the need for constant training and training programs of the work team.

RESUMEN Introdución: El análisis de orina de rutina es una de las pruebas más solicitadas en el laboratorio clínico, pues ayuda en el diagnóstico de diversas enfermedades y en el seguimiento del tratamiento de los pacientes. En este caso, la fase preanalítica es fundamental, puesto que la calidad de la muestra influye directamente en el análisis y en la interpretación del resultado. Objetivo: El objetivo de este estudio fue identificar los principales errores en la fase preanalítica en la prueba de orina rutinaria de un laboratorio privado y su frecuencia de ocurrencia. Material y método: Se recopilaron datos entre 2014 y 2018. En total, se registraron 107.277 muestras de orina; 98 (0,09%) fueron enviadas para nueva extracción. Resultados: La solicitud de nueva extracción fue mayor entre las mujeres (81,6%); los grupos de edad más afectados fueron de 51 a 60 años y de 21 a 30 años. La justificación más común para la toma repetida fue cantidad insuficiente (48%), seguida de confirmación de resultado, en el 24,5% de los casos. El predominio de las muestras de orina en las mujeres ocurrió debido a la frecuencia de los análisis de orina en las mujeres, porque ellas son más propensas a las infecciones urinarias, especialmente en las etapas de la vida sexualmente activa y posmenopáusica. Conclusión: En general, la tasa de toma repetida de orina obtenida en la investigación fue menor que el objetivo estipulado por el laboratorio, pero las principales razones que llevaron a la solicitud de nueva extracción podrían evitarse o minimizarse si los pacientes hubieran sido bien instruidos sobre los procedimientos correctos de recolección, lo que indica la necesidad de programas de capacitación y entrenamiento constante para el equipo de trabajo.

RESUMO Introdução: O exame de urina de rotina está entre os exames mais solicitados no laboratório clínico, pois auxilia no diagnóstico de várias doenças e no acompanhamento do tratamento dos pacientes. Nesse caso, a fase pré-analítica é fundamental, uma vez que a qualidade da amostra influencia diretamente a análise e a interpretação do resultado. Objetivo: O objetivo deste estudo foi identificar os principais erros na fase pré-analítica no exame de urina de rotina de um laboratório privado e sua frequência de ocorrência. Material e método: Dados entre 2014 e 2018 foram coletados. Ao todo, 107.277 amostras de urinas foram cadastradas; 98 (0,09%) foram encaminhadas para recoleta. Resultados: A solicitação de recoletas foi maior no sexo feminino (81,6%); as faixas etárias mais acometidas foram de 51 a 60 anos e de 21 a 30 anos. A justificativa mais comum para recoleta foi material insuficiente (48%), seguida por confirmação de resultado, em 24,5% dos casos. O predomínio de recoletas de urina no sexo feminino ocorreu devido à frequência da realização dos exames de urina em mulheres, pois elas estão mais propensas a infecções urinárias, principalmente na fase de vida sexualmente ativa e na pós-menopausa. Conclusão: De forma geral, o índice de recoletas de urinas obtido na pesquisa foi menor que a meta estipulada pelo laboratório, mas os principais motivos que levaram à solicitação de recoleta poderiam ser evitados ou minimizados se os pacientes tivessem sido bem instruídos quanto aos corretos procedimentos de coleta, o que indica a necessidade de programas de capacitação e treinamento constantes da equipe de trabalho.

El cumplimiento del procedimiento pre-analítico contribuye a la seguridad del paciente. Laboratorio clínico primer nivel de atención. Distrito 09d17-milagro-salud. 2018-2019 / Compliance with the pre-analytical procedure contributes to the safety of the patient. Clinical laboratory first level of care. District 09d17-Milagro-Salud. 2018-2019

El procedimiento pre-analítico en el laboratorio clínico es un conjunto de pasos que inician a partir de la orden de examen realizada por el médico, verificación de la identificación del paciente, criterios de aceptación y rechazo de la muestra, recolección de la muestra, identificación de la muestra, transporte seguro, en temperatura y tiempo apropiado. Luego ingresan para su preparación, y termina en el momento que se inicie con el análisis de las pruebas. Objetivos: Determinar el cumplimiento del procedimiento pre-analítico que contribuye a la seguridad del paciente en el laboratorio clínico. Método: El presente, es un estudio observacional, de tipo cuantitativo, de alcance descriptivo no experimental, de corte transversal. Resultados: de acuerdo a los resultados obtenidos de 112 muestras, con base a la ficha de observación, se logró evidenciar que ha habido muestras con calidad inadecuada como son las hemolizadas, coagulas y contaminadas. Y por estar en esas condiciones, son rechazadas por el laboratorio clínico, lo que conlleva a que se realice otra toma en el paciente, causando malestar, demora en la entrega de sus resultados e insatisfacción. También se evidencia que las muestras hemolizadas son mayores a las coaguladas y contaminadas. Conclusión: Este estudio ha ayudado a conocer que en los puestos de toma de muestra no se está implementando las normas de calidad para evitar errores que han estado causando malestar en los pacientes. Así como también, los puestos de toma de muestra no disponen de equipos para ayudar a conservar las muestras y trazabilidad de sus análisis(AU)

The pre-analytical procedure in the clinical laboratory is a set of steps that start from the order of examination carried out by the doctor, verification of the patient's identification, criteria for acceptance and rejection of the sample, collection of the sample, identification of the sample, safe transport, at appropriate temperature and time. Then they enter for their preparation, and it ends when the analysis of the tests begins. Objectives: To determine compliance with the pre-analytical procedure that contributes to patient safety in the clinical laboratory. Method: This is an observational, quantitative, descriptive, non-experimental, cross-sectional study. Results: according to the results obtained from 112 samples, based on the observation file, it was possible to show that there have been samples with inadequate quality such as hemolizada, coagulated and contaminated ones. In addition, because they are in these conditions, the clinical laboratory the clinical laboratory rejected them, which leads to another taking in the patient, causing discomfort, delay in the delivery of results and dissatisfaction. It is also evident that hemolizada samples are greater than those that are coagulated and contaminated. Conclusion: This study has helped to know that in the sampling stations not implemented quality standards to avoid errors that have been causing discomfort in patients. As well as, the sampling stations do not have equipment to help conserve the samples their traceability and analyzes(AU)

Stability of refrigerated whole blood samples for osmotic fragility test

ABSTRACT Background: The osmotic fragility test (OFT), conventionally used for assisting the diagnosis of many erythrocyte disorders, is a manual and time-consuming analysis not daily performed in many medical laboratories. This study was aimed at defining the stability of whole blood samples used for assessing erythrocyte osmotic resistance. Methods: Twenty-one consecutive routine whole blood samples collected into 5.4 mg K2EDTA were tested immediately after collection (day 0) and at different time intervals afterward (day 1, 2, 3, 4, 7, 10 and 14) after storage at 4 °C. The OFT was performed with the Osmored Monotest (1.3% glycerol; Eurospital, Trieste, Italy). Results at the different time points were compared with those obtained at day 0 and with the reference change value (i.e., 33%). Results: The median value of both hyperosmolar and hyposmolar resistance increased from baseline, reaching statistical significance at day 7 for hyperosmolar resistance and at day 1 for hyposmolar resistance, respectively. The median relative increase of hemolysis percentage values become greater than the reference change value at day 3 for hyposmolar resistance, while this limit was never overcome for hyperosmolar resistance. A significant inverse association was found between the mean increase in hyperosmolar resistance and the baseline value of hyperosmolar resistance (r = −0.92), mean corpuscular volume (MCV; r = −0.46) or mean corpuscular hemoglobin (MCH; r = −0.44), as well as between the mean increase in hyposmolar resistance and the baseline value of hyposmolar resistance (r = −0.86), or patient age (r = −0.56). Conclusions: The sample stability seems critical for the OFT. Whole blood specimens should not be stored refrigerated at 4 °C for >2 days before testing.

Comparación de dos tipos de recipientes para analizar gases en sangre y otras mediciones relacionadas contra jeringas con heparinato de litio liofilizado / Comparison of two types of containers to analyze blood gases and other related measurements against syringes with lyophilized lithium heparinate / Comparação de dois tipos de recipientes para analisar gases em sangue e outras medições relacionadas contra seringas com heparinato de lítio liofilizado

El estudio de gases en sangre involucra pruebas relacionadas con el equilibrio ácido-base y estado de oxigenación (pH, pO2, SO2, pCO2, HCO3 -). Además, en los equipos multiparamétricos se realizan otras determinaciones (mediciones relacionadas) como: Na+, K+, Cl-, Ca2+, glucosa y lactato. El objetivo de este trabajo fue comparar los resultados de medición de dos tipos de recipientes (tipo 2: jeringa preparada en el laboratorio con heparinato de Na+ líquido diluido y tipo 3: microtubo con heparinato de Li líquido) contra el recipiente recomendado por el CLSI en su guía 46-A2 (tipo 1: jeringa con heparinato de Li liofilizado balanceado con zinc). El análisis se hizo desde un punto de vista estadístico y clínico para establecer la posibilidad de usar indistintamente estos tres tipos de recipientes. Se analizaron un total de 254 muestras. Para evaluar la aceptación clínica de los resultados se tomó como estándar de calidad la variabilidad biológica. No se encontraron diferencias clínicamente significativas en los analitos del recipiente tipo 2 respecto del tipo 1, excepto para Ca2+. Se rechazaron desde el punto de vista clínico varios analitos del recipiente tipo 3. En conclusión, el uso del recipiente tipo 3 fue inapropiado. Sin embargo, el recipiente tipo 2 sería apto para el análisis de este tipo de muestras.

Blood gas analysis involves tests related to the acid-base balance and oxygenation state (pH, pO2, SO2, pCO2, HCO3 -). In multiparametric equipment, some ion and metabolite (related measurements) are performed: Na+, K+, Cl-, Ca²+, glucose and lactate. The objective of this study was to compare two types of containers (type 2: syringe prepared in the laboratory with diluted liquid sodium heparinate and type 3: microtube with liquid lithium heparinate) against the container recommended by CLSI in its guide 46-A2 (type 1: syringe with lyophilized lithium heparinate balanced with inc). The analysis was made from a statistical and clinical point of view to establish the possibility of indiscriminately using these three types of containers. A total of 254 samples were analyzed. To establish the clinical acceptance of the results, the biological variability quality standard was used. No clinically significant differences were found in the analytes of the type 2 container compared to type 1, except for Ca+. Several analytes of the type 3 container were rejected from the clinical point of view. In conclusion, the use of the type 3 container is inappropriate; however, the type 2 container would be suitable for the analysis of this type of samples.

O estudo de gases em sangue envolve testes relacionados com o equilíbrio ácido-base e estado de oxigenação (pH, pO2, SO2, pCO2, HCO3 -). Além disso, nos equipamentos multiparâmetros, outras determinações (medições relacionadas) como: Na+ , K+, Cl-, Ca2+, glicose e lactato são realizadas. O objetivo deste trabalho foi comparar os resultados de medição de dois tipos de recipientes (tipo 2: seringa preparada no laboratório com heparinato de Na+ líquido diluído e tipo 3: microtubo com heparinato de Li líquido) contra o recipiente recomendado pelo CLSI em seu guia 46-A2 (tipo 1: seringa com heparinato de Li liofilizado equilibrado com zinco). A análise foi feita do ponto de vista estatístico e clínico, para estabelecer a possibilidade de utilização indiscriminada desses três tipos de recipientes. Um total de 254 amostras foram analisadas. Para avaliar a aceitação clínica dos resultados, a variabilidade biológica foi tomada como padrão de qualidade. Não foram encontradas diferenças clinicamente significativas nos analitos do recipiente tipo 2 em relação ao tipo 1, exceto para Ca²+. Vários analitos do recipiente tipo 3 foram rejeitados do ponto de vista clínico. Em conclusão, o uso do contêiner tipo 3 foi inadequado. No entanto, o recipiente tipo 2 seria apto para a análise deste tipo de amostras.

Normalization in Human Glioma Tissue.

For tissues obtained from glioma samples with/without nonneoplastic brain there is no consensus for universal reference gene but there are some potential genes that might have good stability, under certain conditions. Considering all points described in this work, the care with tissue collection, until gene amplification, directly impacts on the reliable characterization of its mRNA levels. Moreover, it is clear the importance of selecting the most appropriate reference genes for each experimental situation, to allow the accurate normalization of target genes, especially for genes that are subtly regulated.

Stability of refrigerated whole blood samples for osmotic fragility test.

BACKGROUND: The osmotic fragility test (OFT), conventionally used for assisting the diagnosis of many erythrocyte disorders, is a manual and time-consuming analysis not daily performed in many medical laboratories. This study was aimed at defining the stability of whole blood samples used for assessing erythrocyte osmotic resistance. METHODS: Twenty-one consecutive routine whole blood samples collected into 5.4 mg K2EDTA were tested immediately after collection (day 0) and at different time intervals afterward (day 1, 2, 3, 4, 7, 10 and 14) after storage at 4 °C. The OFT was performed with the Osmored Monotest (1.3% glycerol; Eurospital, Trieste, Italy). Results at the different time points were compared with those obtained at day 0 and with the reference change value (i.e., 33%). RESULTS: The median value of both hyperosmolar and hyposmolar resistance increased from baseline, reaching statistical significance at day 7 for hyperosmolar resistance and at day 1 for hyposmolar resistance, respectively. The median relative increase of hemolysis percentage values become greater than the reference change value at day 3 for hyposmolar resistance, while this limit was never overcome for hyperosmolar resistance. A significant inverse association was found between the mean increase in hyperosmolar resistance and the baseline value of hyperosmolar resistance (r = -0.92), mean corpuscular volume (MCV; r = -0.46) or mean corpuscular hemoglobin (MCH; r = -0.44), as well as between the mean increase in hyposmolar resistance and the baseline value of hyposmolar resistance (r = -0.86), or patient age (r = -0.56). CONCLUSIONS: The sample stability seems critical for the OFT. Whole blood specimens should not be stored refrigerated at 4 °C for >2 days before testing.