Comparison between enzyme-linked immunosorbent assay and indirect immunofluorescence for detection of antineutrophil cytoplasmic antibodies.

OBJECTIVE: To evaluate the performance of enzyme-linked immunosorbent assay and indirect immunofluorescence methods for the detection of antineutrophil cytoplasmic antibodies in a routine clinical laboratory setting. METHODS: A total of 227 samples were tested by indirect immunofluorescence and enzyme-linked immunosorbent assay with antigen specificity for antiproteinase 3 and antimyeloperoxidase. The proportions of positive samples were compared by McNemar hypotheses and agreement was described by Cohen's Kappa coefficient. RESULTS: The agreement of the tests was 96.5%, and the Kappa coefficient obtained was 0.70 (95%CI: 0.50-0.90; p<0.001). Considering indirect immunofluorescence as the gold standard, the sensitivity of the enzyme-linked immunosorbent assay was 0.62 and the specificity was 0.99, with diagnostic accuracy in 96% of cases. Some samples were negative in enzyme-linked immunosorbent assay and positive in indirect immunofluorescence. This situation occurred in all immunofluorescence patterns, but particularly in atypical patterns. Two samples with antiproteinase 3 positivity were considered negative in indirect immunofluorescence. CONCLUSION: The enzyme-linked immunosorbent assay had high specificity but lower sensitivity. The performance of indirect immunofluorescence increases diagnostic sensitivity, while the search for antiproteinase 3 by enzyme-linked immunosorbent assay may also add diagnostic power.

Performance evaluation of an indirect immunofluorescence kit for the serological diagnosis of dengue.

OBJECTIVE: To evaluate the performance of indirect immunofluorescence for serological diagnosis of dengue virus in a population with high prevalence of arboviruses. METHODS: Two-hundred serum samples from patients with clinical suspicion of dengue fever were tested by immunoenzymatic and indirect immunofluorescence assay BIOCHIP® mosaic. Specificity, sensitivity and Kappa coefficient were calculated. Discordant samples were tested by polymerase chain reaction for confirmation. RESULTS: Of the 200 samples, 20% were positive and 80% negative for anti-dengue virus IgM antibodies in the immunoenzymatic test. Of the 40 positives, 25% were negative in indirect immunofluorescence. Of these ten discordant results, only 20% were also negative in the polymerase chain reaction (PCR). Of the 160 negatives in the immunoenzymatic test, 5% were positive in indirect immunofluorescence. Of these nine discordant results, 33% were positive in the PCR. The Kappa coefficient was 0.7 (0.572-0.829). Sensitivity and specificity of indirect immunofluorescence were respectively 75% and 94%. For anti-dengue virus IgG antibodies, of the 200 samples, 15.5% were positive and 84.5% were negative in the immunoenzymatic test. Of the 31 positives, 12.9% were negative in indirect immunofluorescence. Of these four discordant results, 25% were negative in the PCR. Of the 169 negatives, 8% were positive in indirect immunofluorescence. Of these 14 discordant results, 64% were also positive in the PCR. The Kappa coefficient was 0.695 (0.563-0.83). Sensitivity and specificity of indirect immunofluorescence were 87.1% and 91.7%, respectively. CONCLUSION: For diagnosis of acute infection, the immunoenzymatic test is enough, and the use of additional methods is not warranted. Replacing the immunoenzymatic test by indirect immunofluorescence would compromise the sensitivity for IgM. However, indirect immunofluorescence can distinguish three arboviruses simultaneously, an advantage during concomitant epidemics.

Performance evaluation of an indirect immunofluorescence kit for the serological diagnosis of dengue / Avaliação do desempenho de kit de imunofluorescência indireta para o diagnóstico sorológico de dengue

ABSTRACT Objective: To evaluate the performance of indirect immunofluorescence for serological diagnosis of dengue virus in a population with high prevalence of arboviruses. Methods: Two-hundred serum samples from patients with clinical suspicion of dengue fever were tested by immunoenzymatic and indirect immunofluorescence assay BIOCHIP® mosaic. Specificity, sensitivity and Kappa coefficient were calculated. Discordant samples were tested by polymerase chain reaction for confirmation. Results: Of the 200 samples, 20% were positive and 80% negative for anti-dengue virus IgM antibodies in the immunoenzymatic test. Of the 40 positives, 25% were negative in indirect immunofluorescence. Of these ten discordant results, only 20% were also negative in the polymerase chain reaction (PCR). Of the 160 negatives in the immunoenzymatic test, 5% were positive in indirect immunofluorescence. Of these nine discordant results, 33% were positive in the PCR. The Kappa coefficient was 0.7 (0.572-0.829). Sensitivity and specificity of indirect immunofluorescence were respectively 75% and 94%. For anti-dengue virus IgG antibodies, of the 200 samples, 15.5% were positive and 84.5% were negative in the immunoenzymatic test. Of the 31 positives, 12.9% were negative in indirect immunofluorescence. Of these four discordant results, 25% were negative in the PCR. Of the 169 negatives, 8% were positive in indirect immunofluorescence. Of these 14 discordant results, 64% were also positive in the PCR. The Kappa coefficient was 0.695 (0.563-0.83). Sensitivity and specificity of indirect immunofluorescence were 87.1% and 91.7%, respectively. Conclusion: For diagnosis of acute infection, the immunoenzymatic test is enough, and the use of additional methods is not warranted. Replacing the immunoenzymatic test by indirect immunofluorescence would compromise the sensitivity for IgM. However, indirect immunofluorescence can distinguish three arboviruses simultaneously, an advantage during concomitant epidemics.

RESUMO Objetivo: Avaliar o desempenho da imunofluorescência indireta no diagnóstico sorológico de dengue em uma população com alta prevalência de arboviroses. Métodos: Duzentas amostras de soro de pacientes com suspeita clínica de dengue foram testadas por ensaio imunoenzimático e imunofluorescência indireta mosaico BIOCHIP®. Foram calculados especificidade, sensibilidade e coeficiente Kappa. Nas amostras discordantes, realizou-se reação em cadeia da polimerase como método confirmatório. Resultados: Das 200 amostras, 20% foram positivas e 80% negativas para IgM antivírus da dengue no ensaio imunoenzimático. Das 40 positivas, 25% foram negativas na imunofluorescência indireta. Destas dez negativas, apenas 20% eram também negativas na reação em cadeia da polimerase. Das 160 negativas no ensaio imunoenzimático, 5% foram positivas na imunofluorescência indireta. Por fim, dentre as nove discordantes, 33% tiveram vírus da dengue detectado na reação em cadeia da polimerase. O coeficiente Kappa foi 0,70 (0,57-0,82). Sensibilidade e especificidade por imunofluorescência indireta foram, respectivamente, 75% e 94%. Para IgG antivírus da dengue, de 200 amostras, 15,5% foram positivas e 84,5% negativas no ensaio imunoenzimático. Das 31 positivas, 12,9% foram negativas na imunofluorescência indireta. Destas quatro discordantes, 25% apresentaram vírus da dengue não detectado na reação em cadeia da polimerase. Das 169 negativas, 8% foram positivas na imunofluorescência indireta. Destas, 64% foram positivas também na reação em cadeia da polimerase. O coeficiente Kappa foi 0,695 (0,56-0,83). Sensibilidade e a especificidade por imunofluorescência indireta foram, respectivamente, 87,1% e 91,7%. Conclusão: Ensaio imunoenzimático seria suficiente para diagnóstico sorológico de infecção aguda, não justificando a incorporação da imunofluorescência indireta. Substituir ensaio imunoenzimático pela imunofluorescência indireta poderia comprometer a sensibilidade para IgM. Contudo, a imunofluorescência indireta auxilia diferenciar três arboviroses simultaneamente, sendo vantajoso em epidemias concomitantes.

Comparison between enzyme-linked immunosorbent assay and indirect immunofluorescence for detection of antineutrophil cytoplasmic antibodies / Comparação entre os métodos de ensaio imunoenzimático e imunoflurescência indireta para a pesquisa de anticorpos anticitoplasma de neutrófilos

ABSTRACT Objective To evaluate the performance of enzyme-linked immunosorbent assay and indirect immunofluorescence methods for the detection of antineutrophil cytoplasmic antibodies in a routine clinical laboratory setting. Methods A total of 227 samples were tested by indirect immunofluorescence and enzyme-linked immunosorbent assay with antigen specificity for antiproteinase 3 and antimyeloperoxidase. The proportions of positive samples were compared by McNemar hypotheses and agreement was described by Cohen's Kappa coefficient. Results The agreement of the tests was 96.5%, and the Kappa coefficient obtained was 0.70 (95%CI: 0.50-0.90; p<0.001). Considering indirect immunofluorescence as the gold standard, the sensitivity of the enzyme-linked immunosorbent assay was 0.62 and the specificity was 0.99, with diagnostic accuracy in 96% of cases. Some samples were negative in enzyme-linked immunosorbent assay and positive in indirect immunofluorescence. This situation occurred in all immunofluorescence patterns, but particularly in atypical patterns. Two samples with antiproteinase 3 positivity were considered negative in indirect immunofluorescence. Conclusion The enzyme-linked immunosorbent assay had high specificity but lower sensitivity. The performance of indirect immunofluorescence increases diagnostic sensitivity, while the search for antiproteinase 3 by enzyme-linked immunosorbent assay may also add diagnostic power.

RESUMO Objetivo Avaliar o desempenho das metodologias de ensaio imunoenzimático e imunofluorescência indireta para a detecção de anticorpos anticitoplasma de neutrófilos em um contexto de laboratório clínico de rotina. Métodos Foram testadas 227 amostras pelas metodologias de imunofluorescência indireta e ensaio imunoenzimático com especificidades para anticorpos antiproteinase-3 e antimieloperoxidase. As proporções de amostras positivas foram comparadas por hipóteses de McNemar, e a concordância foi descrita pelo coeficiente Kappa de Cohen. Resultados A concordância dos testes foi 96,5%, e o coeficiente Kappa obtido foi 0,70 (IC95%: 0,50-0,90; p<0,001). Utilizando a imunofluorescência indireta como padrão-ouro, a sensibilidade do ensaio imunoenzimático foi de 0,62 e a especificidade, 0,99, com acurácia diagnóstica em 96% dos casos. Algumas amostras apresentaram resultados negativos por ensaio imunoenzimático e positivos por imunofluorescência. Isso ocorreu em amostras com vários padrões de fluorescência, mas particularmente nos casos com padrões atípicos. Duas amostras com positividade antiproteinase 3 foram consideradas negativas por imunofluorescência. Conclusão Os métodos de ensaio imunoenzimático tiveram alta especificidade, mas sensibilidade inferior. A realização da imunofluorescência indireta aumenta a sensibilidade diagnóstica, ao mesmo tempo que a pesquisa de antiproteinase 3 por ensaio imunoenzimático também pode agregar poder diagnóstico.