Especificaciones de calidad en base a error total: ¿Cuál es la mejor elección? / Quality specifications based on total error: What's the best choice? / Especificações de qualidade com base em erro total: Qual é a melhor escolha?

El objetivo del trabajo fue evaluar el desempeño anual de los métodos, en términos de error total (ET), las distintas especificaciones de calidad disponibles y el modelo Seis Sigma para calificar desempeño. Se evaluaron analitos con variabilidad biológica (VB), muy baja, baja, media y alta. Se calculó el ET (ETc) y el sigma (s) mensual a dos niveles de control; el ET permitido (ETp) para cada analito se obtuvo de 8 fuentes (metas biológicas y regulatorias). Se consideró desempeño aceptable cuando ETc

The aim of this work was to evaluate the annual performance of the methods in terms of total error (TE), the different quality specifications available, and the Six Sigma model to qualify performance. Analytes with very low, low, medium and high biological variability (BV) were evaluated. TE (TEc) and sigma (s) were calculated monthly at two levels of control; allowable TE (TEa) for each analyte was obtained from 8 sources (biological and regulatory goals). Acceptable performance was considered when TEc

O objetivo do trabalho foi avaliar o desempenho anual dos métodos em termos de erro total (ET), as diferentes especificações de qualidade disponíveis e o modelo Seis Sigma para qualificar desempenho. Foram avaliados analitos com variabilidade biológica (VB) muito baixa, baixa, média e alta. Calculou-se ET (ETc) e o sigma (s) mensal a dois níveis de controle; o ET permitido (ETp) para cada analito foi obtido de 8 fontes (metas biológicas e regulatórias). Levou-se em consideração o desempenho aceitável quando ETc < ETp e s≥3. Foi observada a estabilidade analítica durante o período de avaliação. Não foram alcançadas as metas biológicas para analitos com VB muito baixa, algo semelhante aconteceu para analitos com VB baixa e média; com VB alta alcançaram todas as especificações. O desempenho s e a regra de controle de Westgard dependeram do ETp escolhido; para magnésio, com CLIA (ETp=25%) se obteve s >10 (World Class) e simples regra (13s), com VB mínima s <3 e multirregra. Conclui-se que a aceitação do desemperno do método e as regras de controle dependeram do ETp escolhido, sem disporem nesse meio de metas mínimas a alcançar. O monitoramento mensal do ETc mostrou a estabilidade analítica com variabilidade típica de cada método.

A novel 96-microwell-based high-throughput spectrophotometric assay for pharmaceutical quality control of crizotinib, a novel potent drug for the treatment of non-small cell lung cancer / Um ensaio espectrofotométrico de alto rendimento à base de novo de 96 micropoços para o controlo de qualidade farmacêutica crizotinibe, um novo potente fármaco para o tratamento de cancro do pulmão de células não-pequenas

This study describes the development and validation of a novel 96-microwell-based high throughput spectrophotometric assay for pharmaceutical quality control of crizotinib (CZT), a novel drug for the treatment of non-small cell lung cancer. We examined the reaction between CZT and 1,2-naphthoquinone-4-sulphonate, a chromogenic reagent. A red-colored product showing a maximum absorption peak (λ_max) at 490 nm was produced in an alkaline medium (pH 9). We examined stoichiometry of the reaction and postulated the reaction mechanism. To our knowledge, this is the first study to describe a color-developing reaction for the proposed assay. The reaction was performed in a 96-microwell plate, and the absorbance of the colored product was measured using an absorbance reader at 490 nm. Under optimized reaction conditions, Beer's law, which shows a correlation between absorbance and CZT concentration, was obeyed in the range of 4-50 µg/well with an appropriate correlation coefficient (0.999). The limits of detection and quantification were 1.73 and 5.23 µg/well, respectively. The assay showed high precision and accuracy. The proposed assay was applied successfully for the determination of CZT in capsules. Thus, the assay proposed in this study is practical and valuable for routine application in pharmaceutical quality control laboratories...

Este estudo descreve o desenvolvimento e a validação de um novo ensaio espectrofotométrico em larga escala em 96 micropoços para o controle farmacêutico de crizotinibe (CZT), novo fármaco para o tratamento de câncer de pulmão de células não pequenas. Examinamos a reação entre o CZT e o 4-sulfonato de 1,2-naftoquinona, um reagente cromogênico. Obteve-se, em meio alcalino (pH 9), produto vermelho, com absorção máxima (λ_max) em 490 nm. Examinamos a estequiometria da reação e propusemos mecanismo de reação. Este, segundo nosso conhecimento, é o primeiro estudo para descrever reação de desenvolvimento de cor para o ensaio proposto. A reação foi realizada em placas de 96 micropoços e mediu-se a absorbância do produto colorido utilizando-se leitor de absorbância a 490 nm. Sob condições otimizadas de reação, a lei de Beer, que mostra a correlação entre a absorbância e a concentração de CZT, foi obedecida na faixa de 4-50 µg/poço, com coeficiente de correlação apropriado (0,999). Os limites de detecção e de quantificação foram, respectivamente, 1,73 e 5,23 µg/poço. O ensaio mostrou alta precisão e exatidão. O ensaio proposto foi aplicado com sucesso para a determinação de CZT em cápsulas e é prático e válido para a aplicação de rotina em laboratórios de controle farmacêutico...

Evaluación del desempeño analítico de tres métodos de cuantificación de hemoglobina A1c / Analytical performance evaluation of three methods for hemoglobin A1c quantification / Avaliação do desempenho analítico do três métodos de quantificação de hemoglobina A1c

El laboratorio debe garantizar la exactitud de los resultados de HbA1c cumpliendo con los requisitos analíticos internacionales de calidad, cada vez más estrictos y asegurar que una variación de HbA1c de 0,5 puntos porcentuales (%-NGSP) o más entre dos controles consecutivos de un paciente diabético se deba a una variación clínica y no a una variación analítica. En este trabajo se evaluó el desempeño analítico de tres métodos comerciales para HbA1c: inmunoturbidimétrico, enzimático y cromatográfico de intercambio catiónico. Para tal fin, se procesaron por cada método distintos controles comerciales de HbA1c, con trazabilidad al método de referencia IFCC, determinándose en cada caso Coeficiente de Variación Total, Bias, Error Total, Valor de Referencia del Cambio y cambio clínico significativo de HbA1c en el punto crítico 7,0%-NGSP. En las condiciones analíticas de este trabajo, solamente el método inmunoturbidimétrico tuvo un desempeño analítico aceptable, permitiendo atribuir un cambio de 0,5%-NGSP a una variación clínica significativa del paciente. Frente a las recomendaciones internacionales sobre el uso de HbA1c en el control y diagnóstico de diabetes, es indiscutible la importancia de elegir un método que satisfaga los requerimientos analíticos mínimos de calidad para asegurar la utilidad clínica del resultado de HbA1c.

The laboratory must guarantee the accuracy of HbA1c results meeting the increasingly strict international analytical quality standards and assuring that an HbA1c variation of 0.5 percentage points (%-NGSP) or more between two consecutive controls of a diabetic patient is due to a clinical variation and not to an analytical variation. In this paper, the analytical performance of three commercial methods for HbA1c: Immunoturbidimetric, Chromatographic and Enzymatic Cation Exchange, were evaluated. For this purpose, commercial controls with assigned values traceable to the IFCC reference method for HbA1c were processed. For each methodology, total Coefficient of Variation (CV%), Bias%, Total Error (TE%), Change Reference Value and Clinically Significant Change (CSC) at the critical point of HbA1c 7.0%-NGSP were determined. Within the analytical conditions of this study, only the immunoturbidimetric method had an acceptable analytical performance, allowing attribute a change in 0.5%-NGSP to a significant clinical variation. Faced with international recommendations on the use of HbA1c on control and diagnosis of diabetes, the importance of choosing a method that meets the minimum analytical quality requirements to ensure the clinical utility of HbA1c result is undeniable.

O laboratório deve garantir a precisão dos resultados da HbA1c cumprindo com os requisitos analíticos internacionais de qualidade cada vez mais exigentes e garantir que uma variação de HbA1c de 0,5 pontos percentuais (% - NGSP) ou mais entre duas verificações consecutivas de um doente diabético seja devido a uma variação clínica e não a uma variação analítica. Neste trabalho foi avaliado o desempenho analítico de três métodos comerciais para HbA1c: imunoturbidimétrico, enzimático e cromatográfico de intercâmbio catiônico. Para esse fim, foram processados diversos controles comerciais de HbA1c por cada método, com rastreabilidade ao método de referência IFCC, determinando em cada caso Quociente de Variação Total, Bias, Erro Total, Valor de Referência da Alteração e Alteração Clinicamente Significativa de HbA1c no ponto crítico 7,0%-NGSP. Nas condições de análise deste estudo, apenas o método imunoturbidimétrico teve um desempenho analítico aceitável, permitindo atribuir uma alteração de 0,5%-NGSP a uma variação clínica significativa do paciente. Perante as recomendações internacionais sobre o uso da HbA1c no controle e diagnóstico da diabetes, é inegável a importância de escolher um método que atenda os requisitos analíticos mínimos de qualidade de análise para garantir a utilidade clínica do resultado HbA1c.

Técnica de cuantificación de la agregación eritrocitaria por análisis digital de imágenes / Erythrocyte aggregation quantification technique using digital image analysis / Técnica de quantificagáo do agregado eritrocitário por análise digital de imagens

La caracterización de agregados eritrocitarios es importante para analizar las posibles alteraciones en la microcirculación observadas en ciertas patologías vasculares como la hipertensión y la diabetes. El objetivo de este trabajo fue estandarizar una técnica que pueda ser utilizada por cualquier operador con un equipamiento similar. Para ello se prepararon distintas suspensiones de glóbulos rojos de dadores sanos en plasma autólogo, que fueron observadas con un microscopio óptico invertido. Se registraron para su análisis las imágenes de los agregados con una cámara digital. Se realizaron los recuentos de células individuales, agregados de 2 a 4 células, agregados de 5 ó más células y amas (redes de agregados de gran tamaño). Se midió el perímetro y el área, obteniéndose un parámetro de forma (ASP) de cada agregado de 5 ó más células. Los resultados obtenidos permitieron estandarizar el protocolo de trabajo concluyendo que la dilución óptima de glóbulos rojos en plasma autólogo para esta técnica es 0,5%.

Characterization of erythrocyte aggregates is important in the analysis of the possible alterations observed in the microcirculation of certain vascular pathologies such as hypertension and diabetes. The objective of this work was to standardize a technique that can be used by any operator having similar equipment. For that purpose, different suspensions of red blood cells from healthy donors were prepared in autologous plasma and then observed with an inverted light microscope. The images of the aggregates were recorded with a digital camera in order to be later analyzed. Individual cell count was carried out, as well as 2 to 4 cell- aggregates, 5 or more cell- aggregate and amas (big aggregate networks). Measurement of perimeter and area of each of the aggregates made up of 5 or more cells was performed, getting a shape parameter (ASP). Due to the results obtained, this working protocol has been standardized and it can be concluded that the optimal dilution of red blood cells in autologous plasma is 0.5% for this particular technique.

A caracterizagáo de agregados eritrocitários é importante para analisar as possíveis alteragóes na microcirculagáo observadas em certas patologias vasculares como a hipertensáo e a diabetes. O objetivo deste trabalho foi padronizar uma técnica que possa ser utilizada por qualquer operador com um equipamento similar. Para isso foram preparadas diversas suspensóes de glóbulos vermelhos de doadores saudáveis em plasma autólogo, que foram observadas com um microscópio óptico invertido. Foram registradas para a sua análise as imagens dos agregados com uma camera digital. Realizaramse as recontagens de células individuais, agregados de 2 a 4 células, agregados de 5 ou mais células e amas (redes de agregados de grande tamanho). Foi medido o perímetro e a área, obtendo um parametro de forma (ASP) de cada agregado de 5 ou mais células. Os resultados obtidos permitiram padronizar o protocolo de trabalho concluindo que a diluigáo ótima de glóbulos vermelhos em plasma autólogo para esta técnica é de 0,5%.

Impacto del Programa de Evaluación Externa de la Calidad en Química Clínica en México de 2004 a 2008 / Impact of the Program of External Quality Assessment in Clinical Chemistry in Mexico from 2004 to 2008 / Impacto do Programa de Avaliação Qualidade Externa em Química Clínica no México de 2004 a 2008

Se valoró el impacto del Programa de Evaluación Externa de la Calidad, aplicable a laboratorios clínicos en el área de Química Clínica, en México, con base en resultados obtenidos por los laboratorios durante el ciclo marzo 2008-febrero 2009 y el periodo 2004-2008, mediante un estudio analítico, longitudinal y retrospectivo de los resultados obtenidos por los laboratorios que participaron en el Programa de Evaluación Externa de la Calidad de la Facultad de Ciencias Químicas, Universidad Autónoma de San Luis Potosí. El análisis estadístico se realizó con los programas Microsoft® Office Excel y Epi Info T. El porcentaje de laboratorios clínicos con desempeño aceptable (excelente y bueno) por analito, durante el ciclo de evaluación marzo 2008-febrero 2009, fue del 75% al 82%, que aumentó cuando se utilizaron métodos automatizados y semiautomatizados. Para el periodo 2004-2008, los laboratorios en 2004 tuvieron 3,02 veces mayor riesgo de no calificar con desempeño aceptable (p<0,05) que en 2008. En conclusión, el Programa de Evaluación Externa de la Calidad de la Facultad de Ciencias Químicas, Universidad Autónoma de San Luis Potosí, ha tenido un impacto favorable en el desempeño global de los laboratorios clínicos, que permite asegurar su calidad analítica.

The impact of the External Quality Assessment Program, applicable to clinical laboratories in the area of Clinical Chemistry in Mexico, was studied, based on laboratory results during the March 2008-February 2009 cycle and the 2004-2008 period, through analytical, longitudinal, retrospective analyses of the results obtained by the laboratories that participated in the External Quality Assessment Program of the School of Chemical Sciences of Universidad Autónoma de San Luis Potosí. Statistical analysis was performed with Microsoft® Office Excel and Epi Info T programs. The percentage of clínical laboratories with acceptable performance (excellent and good) by analyte during the evaluation cycle in March 2008-February 2009 was 75% to 82%, whích íncreased when automated and semíautomated methods were used. For the 2004-2008 period, the laboratories, in 2004, had 3.02 times greatet risk of not qualifying with acceptable performance (p<0.05) than in 2008. In conclusión, the External Qualíty Assessment of the School of Chemícal Sciences of Universidad Autónoma de San Luis Potosí has had a strong impact on the overall performance of clinical laboratories, whích ensures the latter's qualíty.

Evaluación de espectrofotómetros para la elaboración de material de referencia empleado en la calibración y el control de la determinación de hemoglobina / Evaluation of spectrophotometers for the preparation of reference material used in the calibration and control of the determination of hemoglobin

Debido a la variabilidad de resultados entre laboratorios en las determinaciones de hemoglobina, se debe promover la implantación de sistemas de garantía de la calidad para generar confianza en los resultados. A fin de aportar elementos para su establecimiento, se realizó una evaluación instrumental de espectrofotómetros y se preparó un lote de controles y calibradores de hemoglobina siguiendo protocolos internacionales. Después de elaborar el material, se monitoreó homogeneidad (CV calibrador: 0,33 por ciento, control: 0,38 por ciento) y estabilidad (CV calibrador: 0,13 por ciento, control: 0,90 por ciento). La evaluación instrumental demostró la condición óptima de los espectrofotómetros (exactitud de la longitud de onda 100 por ciento, inexactitud fotométrica 0,9 por ciento y 1,2 por ciento, CV precisión fotométrica 0,25 por ciento, respuesta lineal con K2Cr2O7) lo que permitió su empleo para asignar valores al material preparado, que fue útil para el control de la determinación de hemoglobina. Esto constituye un aporte al desempeño del laboratorio y contribuye a la emisión de resultados confiables

On account of the variability of results observed between laboratories in hemoglobin determinations, the installation of quality assurance systems should be promoted to generate confidence on laboratory results. In order to contribute with elements for the establishment of these systems we realized an instrumental evaluation of photometric equipment and prepared a regular lot of hemoglobin standards and controls following international protocols. After the elaboration, we monitored the homogeneity (VC standard: 0.33 percent, control: 0.38 percent) and stability (VC standard: 0.13 percent, control: 0.90 percent). The instrumental evaluation showed the optimal condition of the photometric equipment (wave length accuracy 100 percent, photometric inaccuracy 0.9 percent y 1.2 percent, VC photometric precision 0.25 percent, lineal response with K2Cr2O7), this allowed its employment for the value assignation of the material, that was useful for the daily practice of hemoglobin determination. This constitutes a contribution to the performance of clinical laboratory and contributes with the emission of reliable results

Influencia de las variables preanalíticas en la determinación de proteína S / Influence of preanalytical variables in S protein determination

La proteína S (PS) regula el sistema de coagulación mostrando actividad de cofactor de la Proteína C activada (PCa) con la cual forma un complejo equimolecular. En presencia de iones calcio este complejo inactiva por proteólisis los factores V y VIII activados por trombina. La proteína S plasmática circula 40% libre (fracción que presenta actividad de cofactor de la PCa) y 60% unida al C4-BP (proteína ligante de la fracción C4 del complemento). El objetivo fue comparar el dosaje de PS realizado por método coagulable e inmunoturbidimétrico e investigar cómo las variables preanalíticas afectan los niveles de PS determinados. Se obtuvieron los siguientes resultados: método coagulable: CV intra ensayo: (n=20): 4%, CV interensayo (n=20, 3 días): 3,4%. Método inmunoturbidimétrico: CV intraensayo (n=20): 3,7%.CV Inter. ensayo (n=20,3 días): 4,5%. Existe buena correlación (R2=0,94) entre ambos métodos, cuando la calibración por el método coagulable se realiza en la misma corrida analítica que las muestras. Cuando se realizó el estudio de Bland y Altman los dos métodos mostraron ser comparables en todos los niveles de PS estudiados. No se observaron diferencias significativas entre las muestras determinadas frescas y conservadas a -20 y -80 °C descongeladas solo una vez.

Protein S has an essential anticoagulant function acting as activated Protein C cofactor and forming an equimolecular complex with it. In the presence of calcium this complex regulates the coagulation process inactivating thrombin activated factors V and VIII by proteolysis. In plasma there are two different forms: a) free Protein S which acts as the cofactor of activated protein C (representing about 40% of total Protein S) and b) C4-BP(C4 binding protein) bound protein S which exhibits no activity as cofactor of activated Protein C (representing about 60% of total PS). The objetive was to compare the PS dosage determination by two methods: immunoturbidimetric and clotting, and to investigate how pre-analytical variables affect the results. The following results were obtained: Clotting method: CV intra assay: (n=20): 4%, CV interassay (n=20, 3 days): 3.4%; immunoturbidimetric method CV intra assay (n=20): 3.7%: CV inter assay (n=20.3 days): 4.5%. There is a good correlation (R2 = 0.94) between both methods; when the clotting method is calibrated in batch with the samples. There is significant difference between fresh and frozen ( -20 °C and -80 °C) samples when the latter have been desfrozen only once.

[Feasibility of using Statistical tests in evaluation of non-uniformity]

Non-uniformity test is essentially the only required daily QC procedure in nuclear medicine practice. Noise creates statistical variation or random error in a flood image. Non-uniformity on the other hand does not have statistical nature and may be regarded as systemic error. The present methods of non-uniformity calculation do not distinguish between these two types of error. The Jarque-Bera and Kolmogorov-Smirnov tests were examined as alternative methods in calculation of non-uniformity in flood test images. Using the Monte carol method, uniform and non-uniform flood images of different matrix sizes and count density were generated. The uniformity of the images was calculated using the present and proposed methods. The results were also tested using 1300 planar images of 128x128 matrix size. The proposed methods were more accurate and sensitive to non-uniformity at low count density. However, their precisions were less than the conventional methods. There were no significant differences between these procedures at high count density. The integral and differential uniformity do not distinguish between noise always present in the data or in occasional non-uniformity. In a uniform intact flood image, the difference between maximum and minimum pixel count [the value of integral uniformity] is much more than recommended values for non-uniformity. After filtration of image this difference decreases but still remains high. The proposed methods are more sensitive to non-uniformity at low count density and may be used as alternative methods in daily uniformity test

[Comparison of acceptance tests for SPECT systems in Tehran]

Acceptance test is a necessary procedure after SPECT system installation. The goal of this test is to reveal system's present condition, to compare it with manufacturer's specifications and also as a base for later tests. This study investigated four SPECT systems in Tehran. All of the quality control tests are performed on the basis of NEMA and IAEA recommendations and by a same group. These tests include intrinsic spatial resolution, intrinsic energy resolution, temporal resolution, intrinsic linearity, maximum count rate, pixel size, intrinsic and extrinsic uniformity, sensitivity, reconstruction spatial resolution with and without scatter and centre of rotation. Results of this investigation show that three systems have minimum acceptance conditions, but the fourth one due to it's suboptimal energy resolution and spatial resolution lacked the required specifications for acceptance. It was shown during the next six months after installation that this system showed frequent impairments and even had been out of service for a while. However, other systems did not show any considerable problems. The acceptance test is an essential step after installation of any SPECT system. If there is no considerable deficits at the initial acceptance test of a SPECT system, it won't become troublesome for a long time

Alternative methods for evaluation of non-uniformity in nuclear medicine images

Non-uniformity test is the most essential in daily quality control procedures of nuclear medicine equipments. However, the calculation of non-uniformity is hindered due to high level of noise in nuclear medicine data. Non-uniformity may be considered as a type of systematic error while noise is certainly a random error. The present methods of uniformity evaluation are not able to distinguish between systematic and random error and therefore produce incorrect results when noise is significant. In the present study, two hypothetical methods have been tested for evaluation of non-uniformity in nuclear medicine images. Using the Monte Carol method, uniform and non-uniform flood images of different matrix sizes and different counts were generated. The uniformity of the images was calculated using the conventional method and proposed methods. The results were compared with the known non-uniformity data of simulated images. It was observed that the value of integral uniformity never went below the recommended values except in small matrix size of high counts [more than 80 millions counts]. The differential uniformity was quite insensitive to the degree of non-uniformity in large matrix size. Matrix size of 64'64 was only found to be suitable for the calculation of differential uniformity. It was observed that in uniform images, a small amount of non-uniformity changes the p-value of Kolmogorov-Smirnov test and noise amplitude of fast fouries transformation [FFT] test significantly while the conventional methods failed to detect the non-uniformity. The conventional methods do not distinguish noise, which is always present in the data and occasional non-uniformity at low count density. In a uniform intact flood image, the difference between maximum and minimum pixel count [the value of integral uniformity] is much more than the recommended values for non-uniformity. After filtration of image, this difference decreases, but remains high. Both proposed methods were more sensitive to the non-uniformity at a much lower count density

El problema de las partículas en suspensión en preparados inyectables de pequeño volumen: estudio de la uniformidad visual en los inspectores / The problem of suspension particles in parenteral preparations of small volume: visual uniformity study in the inspectors

Se estudió la uniformidad en la inspección visual de productos inyectables de pequeño volumen en un grupo de inspectores voluntarios adultos del sexo masculino. La muestra estuvo constituida por 600 ampolletas de vidrio que contenían 2 ml de agua bidestilada. El número de ampolletas inspeccionadas simultáneamente por cada inspector fue de tres, durante 15 segundos, en un lapso de 90 minutos. El diseño experimental se realizó en tres etapas. Los resultados de las primeras dos etapas demostraron, mediante la prueba de Kruskal-Wallis, que no hubo diferencias significativas en el número de ampolletas detectadas defectuosas por un mismo inspector, ni en los horarios ni en los días estudiados; tampoco hubo diferencias significativas entre los inspectores. Los resultados de la tercera etapa demostraron, por un modelo de regresión lineal, que el número de ampolletas detectadas defectuosas se conservó constante a lo largo de seis meses. El método de inspección visual aquí descrito para la idenficación de partículas en productos inyectables de pequeño volumen demostró ser reproducible, confiable y aplicable en el control de calidad del producto terminado