ABSTRACT
Objetivos: Establecer la validez del test de TUNEL en la determinación de la fragmentación del ADN espermático, la relación entre el grado de fragmentación con los parámetros seminales y la muestra a emplear para efectuar la prueba. Materiales y métodos: Se emplearon muestras de semen de varones sanos fértiles (n = 33), pacientes que consultaron por infertilidad con prescripción de test de TUNEL (n = 77) y pacientes con fracaso en ICSI (n = 20), analizadas según OMS 2010. Se efectuó el test de TUNEL/ioduro de propidio por citometría de flujo, en muestras basales y post swim up. Resultados: El valor de corte de TUNEL (curvas ROC) fue de 26% con sensibilidad y especificidad de 85 y 89% respectivamente. Las medianas de los resultados de TUNEL pre y post swim up no mostró diferencia significativa (17,0 vs. 12,9%). Sin embargo, el 39,1% expuso una diferencia superior a 15 en valor absoluto entre los resultados del TUNEL basal y post swim up. El estudio de correlación lineal de la morfología, movilidad y vitalidad con el TUNEL post swim up mostró una correlación mayor que el pre selección, con resultados significativos (r: -0,394, p < 0,0001; r: -0,461, p < 0,0001; r: -0,526, p < 0,0001). Conclusiones: La prueba de TUNEL es una prueba válida para su empleo en la clínica. La fragmentación del ADN es un factor independiente de las pruebas tradicionales del semen. Hallamos mayor susceptibilidad al daño generado en los procedimientos de laboratorio en las muestras de peor calidad. La muestra de elección para evaluar la fragmentación del ADN dependerá de si se trata de una fertilización natural o asistida
Objectives: To establish the validity of the TUNEL assay in determining sperm DNA fragmentation, the relationship between the degree of fragmentation and the seminal parameters and the sample needed to conduct the test. Material and methods: We used semen samples from healthy fertile men (n = 33), patients who consulted for infertility with a prescription for the TUNEL assay (n = 77) and patients with intracytoplasmic sperm injection failure (n = 20), analyzed according to the 2010 WHO. The TUNEL/propidium iodide test was performed by flow cytometry, on baseline and post-swim-up samples. Results: The cutoff value for the TUNEL assay (ROC curves) was 26%, with a sensitivity and specificity of 85% and 89%, respectively. The pre-swim-up and post-swim-up medians of the results from the TUNEL assay showed no significant differences (17.0% vs. 12.9%, respectively). However, 39.1% of the samples showed a difference greater than 15 in absolute value between the results of the baseline and post-swim-up TUNEL assays. The linear correlation study of the morphology, mobility and vitality using the post-swim-up TUNEL assay showed a greater correlation than preselection, with significant results (r: -0.394, P < .0001; r: -0.461, P < .0001; r: -0.526, P < .0001). Conclusions: The TUNEL assay is a valid test for clinical use. DNA fragmentation is a factor independent from traditional semen tests. We found a greater susceptibility to damage generated in the laboratory procedures in the samples with lower quality. The sample of choice for evaluating DNA fragmentation will depend on whether the clinician is treating a natural or assisted fertilization
Subject(s)
Humans , Male , DNA Fragmentation , In Situ Nick-End Labeling/methods , Semen Analysis/methods , Reproducibility of Results , Specimen Handling/methods , Reproductive Techniques, AssistedABSTRACT
OBJECTIVES: To establish the validity of the TUNEL assay in determining sperm DNA fragmentation, the relationship between the degree of fragmentation and the seminal parameters and the sample needed to conduct the test. MATERIAL AND METHODS: We used semen samples from healthy fertile men (n=33), patients who consulted for infertility with a prescription for the TUNEL assay (n=77) and patients with intracytoplasmic sperm injection failure (n=20), analyzed according to the 2010 WHO. The TUNEL/propidium iodide test was performed by flow cytometry, on baseline and post-swim-up samples. RESULTS: The cutoff value for the TUNEL assay (ROC curves) was 26%, with a sensitivity and specificity of 85% and 89%, respectively. The pre-swim-up and post-swim-up medians of the results from the TUNEL assay showed no significant differences (17.0% vs. 12.9%, respectively). However, 39.1% of the samples showed a difference greater than 15 in absolute value between the results of the baseline and post-swim-up TUNEL assays. The linear correlation study of the morphology, mobility and vitality using the post-swim-up TUNEL assay showed a greater correlation than preselection, with significant results (r: -0.394, P<.0001; r: -0.461, P<.0001; r: -0.526, P<.0001). CONCLUSIONS: The TUNEL assay is a valid test for clinical use. DNA fragmentation is a factor independent from traditional semen tests. We found a greater susceptibility to damage generated in the laboratory procedures in the samples with lower quality. The sample of choice for evaluating DNA fragmentation will depend on whether the clinician is treating a natural or assisted fertilization.
