Comprehensive analysis of sperm DNA fragmentation by five different assays: TUNEL assay, SCSA, SCD test and alkaline and neutral Comet assay

Abstract

SUMMARYSperm DNA fragmentation (SDF) is becoming an important test to assess male infertility. Several different tests are available, but no consensus has yet been reached as to which tests are most predictive of infertility. Few publications have reported a comprehensive analysis comparing these methods within the same population. The objective of this study was to analyze the differences between the five most common methodologies, to study their correlations and to establish their cut-off values, sensitivity and specificity in predicting male infertility. We found differences in SDF between fertile donors and infertile patients in TUNEL, SCSA, SCD and alkaline Comet assays, but none with the neutral Comet assay. The alkaline COMET assay was the best in predicting male infertility followed by TUNEL, SCD and SCSA, whereas the neutral COMET assay had no predictive power. For our patient population, threshold values for infertility were 20.05% for TUNEL assay, 18.90% for SCSA, 22.75% for the SCD test, 45.37% for alkaline Comet and 34.37% for neutral Comet. This work establishes in a comprehensive study that the all techniques except neutral Comet are useful to distinguish fertile and infertile men.

Fluctuation of “sperm DNA integrity” in accordance with semen parameters, and it’s relationship with infertility

“…% Tail DNA represented a threshold value of 15 % for predicting male infertility with an area under the curve 0,837 and Tail Moment represented a threshold value of 15,21 % with an area under the curve 0,796. However, these results are not comparable with previous studies because it is not mentioned which comet measurement were used in these studies [15,18,31]. Higher area under the ROC curve has been shown by % Tail DNA with a high sensitivity and specificity of 0,717 and 0,800 respectively (Table 1, Fig.…”

“…Statistical analysis SPSS v. 15 software was used to analyze and to graph the data collected in the study. Descriptive statistics was used to describe the basic features of semen volume, sperm count, motility, morphology, % Tail DNA and Tail Moment.…”

The pellet swim-up is the best technique for sperm preparation during in vitro fertilization procedures

“…We chose this commercial kit for the following reasons: the test was easy to set up and it did not require access to expensive equipment, which would have been unavailable in the routine laboratory where the study was carried out; this test also bears the CE marking. Moreover, it has been demonstrated that the sperm chromatin dispersion test is an effective tool for assessing sperm DNA fragmentation when compared with other tests [11,16,17]. The SCD test and TUNEL assay have the capacity to give similar results in terms of DNA fragmentation, and it has been demonstrated that the SCD test is even more sensitive than the TUNEL assay [18].…”

Paternal contribution to development: Sperm genetic damage and repair in fish

“…Ribas-Maynou and colleagues (Ribas-Maynou et al 2013) proposed that the best method to assess chromatin lesions in sperm is the alkaline comet assay, followed by TUNEL, SCD and SCSA. So in conclusion we can determine that the rational choice of a method should rely on the particular objective and that a combination of different methodologies would provide a more informative diagnostic about sperm quality or about the damages promoted by a particular agent.…”

“…The DNA fragmentation index (DFI) (ratio between red fluorescence and total fluorescence) is used to express the results. This technique is widely used in mammals and it is considered a useful approach for the clinical diagnosis of human fertility, mainly for assisted reproductive technologies (ART) (Evenson et al 1999;Ribas-Maynou et al 2013;Bungum et al 2011;Farah et al 2013;Palermo et al 2014). However the application of SCSA for the analysis of teleost spermatozoa does not provide reliable results, probably due to the requirement of modifications in the denaturation process, in accordance to the specific chromatin packaging.…”