MÉTODOS PARA LA IDENTIFICACIÓN DEL ANTÍGENO DE HISTOCOMPATIBILIDAD HLA-B27: COMPARACIÓN DE CUATRO PROTOCOLOS TÉCNICOS / METHODS FOR IDENTIFICATION OF ANTIGEN HLA-B27 HISTOCOMPATIBILITY: FOUR TECHNICAL COMPARISONS OF PROTOCOLS / MÉTODOS PARA A IDENTIFICAÇÃO DO ANTIGÉNIO DE HISTOCOMPATIBILIDADE HLA-B27: CAMPARAÇAO DE QUATRO PROTOCOLOS TÉCNICOS

Introducción: La asociación del HLA-B27 y las Espondiloartritis, ha hecho evidente que la tipificación del HLA-B27 sea considerada como un apoyo en el diagnóstico de estas enfermedades. Los métodos más empleados para la determinación del antígeno HLA-B27 en los laboratorios clínicos y en investigación son: la microlinfocitotoxicidad (MCTX), la citometría de flujo digital (CMFd), la citometría de flujo análoga (CMFa) y la reacción en cadena de la polimerasa con primers de secuencia específicos (PCR-SSP). Objetivo: Comparar MCTX con la CMFd, la CMFa con la CMFd, y la técnica de CMFd frente a PCR-SSP. Métodos: Se analizaron 4109 solicitudes de HLA-B27 en población con manifestaciones sugestivas de EAS remitidas entre 2009 y 2012 al Hospital Militar Central y al Instituto de Referencia Andino. Se evaluaron las frecuencias obtenidas por Chi cuadrado (X2); para estimar la concordancia metodológica se utilizó el Coeficiente de Correlación Intraclase (CCI). Los análisis se realizaron con el paquete estadístico SPSS V18. Resultados: Al evaluar 467 datos por la técnica de CMFa frente a PCR-SSP, la CMFa mostró 239 resultados entre positivos y en rango indeterminado, de los cuales, luego de ser confirmados PCRSSP, solo 213 demostraron la expresión de HLA-B27 (p<0.05). Se obtuvieron 208 resultados realizados por CMFd y PCR-SSP simultáneamente, observándose una alta correspondencia entre estas técnicas (p<0.05). Para evaluar la concordancia entre la MCTX y CMFd se analizaron 34 datos, revelando un 100% de correspondencia entre esta dos metodologías (CCI=1,p<0.05). Conclusión: La citometría de flujo digital es un método rápido que presenta un desempeño altamente confiable para la identificación de HLA-B27, resultados que se recomiendan confirmar por PCR SSP.

Introduction: The association between HLA-B27 and spondyloarthritis has made clear the fact that identification of HLA-B27 antigen is considered as a support in the diagnosis of these diseases. The most commonly used methods for determination of the HLA-B27 antigen in clinical laboratories as well as in their research, are microlymphocytotoxicity (MCTX), digital flow cytometry (CMFd), analogous flow cytometry (CMFa) and the Single Specific Primer-Polymerase Chain Reaction (PCRSSP). Objective: compare the CMFd against MCTX, CMFa against CMFd and CMFd against PCR-SSP. Methods: 4109 requests for HLA-B27 were analyzed with manifestations suggestive of SpA submitted between 2009 and 2012 at Hospital Militar Central and Instituto de Referencia Andino. To analyze the frequencies Chi square (X2) was evaluated; to estimate the methodological concordance the intraclass correlation coefficient (ICC) was used. All proposed analyzes were performed with SPSS V18. Results: 467 data obtained by CMFa versus PCR-SSP evaluated the CMFA showed 239 results between positive and indeterminate range, which, after being confirmed by molecular biology (PCRSSP), only 213 showed the expression of HLA-B27 (p <0.05). PCR-SSP and CMFd performed 208 results simultaneously, showing a high correlation between these techniques (p <0.05). To evaluate the correlation between CMFd and MCTX, 34 data were analyzed, revealing a 100% match on the positive results from these two methodologies (ICC = 1, p <0.05). Conclusion: The digital flow cytometry is a rapid method that presents a highly reliable for the initial identification of HLA-B27; results confirmed by PCR SSP recommend performance.

Introdução: a associação do HLA-B27 e as Espondilartrite, evidenciou que a tipificação do HLAB27 seja considerada como um suporte no diagnóstico dessas doenças. Os métodos mais usados para a determinação do antígeno HLA-B27 nos laboratórios clínicos e no investigação são: a microlinphocitotoxicity (MCTX), a citometria de fluxo digital (CMFd), a citometria de fluxo análoga (CMFa) e a reação em cadeia de a polimerasa com primers de sequência específicos (PCR-SSP). Objetivo: Comparar MCTX com a CMFd, a CMFa com a CMFd, e a técnica de CMFd com PCRSSP. Métodos: 4109 solicitudes de HLA-B27 em população com manifestações sugestivas de EAS remitidas entre 2009 e 2012 ao Hospital Militar e ao Instituto de Referencia Andino, foram analisadas. Avaliaram-se as frequências obtidas por Chi quadrado (X2); para estimar a concordância metodológica foi utilizado o Coeficiente de Correlação Intraclasse (CCI). Os análises estão feitos com o paquete estadístico SPSS V18. Resultados: A CMFa mostrou 239 resultados entre positivos e em rango indeterminado quando avaliou-se 467 dados com a técnica de CMFa com PCR-SSP. Só 213 deles demostraram a expressão de HLA-27 (p<0.05), depois de ser confirmados PCR-SSP. Foram obtidos 208 resultados por CMFd y PCR-SSP em simultâneo, com uma alta correspondência entre estas técnicas (p<0.05). Para avaliar a concordância entre MCTX y CMFd analisaram-se 34 dados, revelando um 100% de correspondência entre as duas metodologias (CCI=, p<0.05). Conclusão: A citometria de fluxo é um método rápido que tem um desempeno muito confiável para a identificação de HLA-B27, resultados recomendados para confirmar por PCR SSP.

Role of hla class-I antigens in delusional disorder.

The investigation was conducted to find out whether there is any association between delusional disorder and HLA antigens. The sample comprised 50 patients with delusional disorder and 282 control samples collected from normal controls. Statistical analysis revealed that the frequency of A3 antigen of the locus A are significantly higher. In case of HLA - B locus significantly higher frequency of B5 and B21 antigens have also been observed. The present study shows that there may be some association of HLA class-1 antigens with delusional disorder.

Comparison of HLA-B27 typing methods -PCR-SSP, microlymphocytotoxicity, and flow cytometry / 대한임상병리학회지

BACKGROUND: HLA-B27 typing has long been performed by the microlymphocytotoxicity method(MCT) but the flow cytometry method(FCM) was introduced several years ago. False positive results due to the HLA-B7 cross reactive groups(CREG) were the main drawback of the serologic method. The authors performed polymerase chain reaction-sequence specific primer(PCR-SSP) test for HLA-B27 to compare the results with serologic methods. METHODS: PCR-SSP test for HLA-B27 was performed on four hundred forty one samples. Three hundred twenty eight samples were tested by MCT and one hundred thirteen samples by FCM. PCR-SSP for HLA-B27 subtyping or Amplification Refractory Mutation System-PCR(ARMS-PCR) for HLA-B typing was performed on twenty four discrepant samples. RESULTS: The concordance rate between MCT and PCR-SSP was 92.9%(305/328) and the concordance rate between FCM and PCR-SSP was 99.1%(112/113). Twenty four(5.4%) out of four hundred forty one samples showed discrepancy between serologic methods and PCR-SSP method. Fourteen out of one hundred MCT positive samples and only one out of forty FCM positive samples showed negative by PCR-SSP. Nine samples showed PCR-SSP positive and MCT negative. CONCLUSIONS: The false positive rate of MCT was quite high and there were some false positive and negative results by PCR-SSP, too. From the above findings, we suggest that FCM is the most accurate method for HLA-B27 typing in those laboratory equipped with flow cytometry.

HLA Class I Typing for Umbilical Cord Blood / 대한임상병리학회지

BACKGROUND: Accurate HLA typing is important for umbilical cord blood (UCB) transplantation as well as bone marrow transplantation. However, HLA class I typing by standard microcytotoxicity method has been often unsatisfactory for UCB samples. This study was conducted to investigate the characteristics of cytotoxic reaction in HLA class I microlymphocytotoxicity testing for UCB. METHODS: We compared the strength index (SI) and the frequencies of HLA antigens with weak reaction between 87 UCB and 103 adult peripheral blood samples by analysis of reaction score in microlymphocytotoxicity typing using Terasaki Oriental HLA-ABC (72) Well Tray (One Lambda, USA). RESULTS: The mean SI of UCB samples in HLA class I typing was significantly lower than that of adult blood (78.5% vs. 96.8%). The SI of 100% among UCB and adult blood samples were 24.1% and 71.8%, respectively. Among HLA-A, B, C antigens, those showing significantly high frequencies of weak reaction in UCB were HLA-A31, B48, B51, B59, B61, Cw3 and Cw7. Cytotoxic reactions of Bw4 and Bw6 in UCB were significantly weaker than those in adult blood. CONCLUSIONS: The strategy of using a supplementary DNA typing method in selected cases with doubtful or unreliable results in serological typing would be effective for an accurate HLA class I typing of UCB samples.

INVESTIGATION ON DISTRIBUTION OF HLA-DR DQ ANTIGENS JN NORTH-CHINESE CHILDREN OF HAN NATIONALITY / 西安交通大学学报(医学版)

The distribution of HLA antigens was investigated in 32 North-Chinese Han children aged 12～14 years, 20 of whom are male, the others are female. Except HLADR1, the frequencies of HLA-DR, DQ antigens in those children were almost identical to those in Han people in Beijing, but the difference of HLA-DR1 between the two groups was not significant statistically (Pc≥0.197). The difference of HLA-DR, DQ antigens between those children and Han people in Shanghai was obvious. This suggests that there is a differerce in distribution of HLA antigens beween North and Sourth Han Chinese people. Compared with the date of other minority nationalities, this result approaches that of North-Chinese minority pationalities such as Korean, Mongolian, etc.