In the glycolytic process, 2 enzymes in the same pathway have been identified as PHA-848125 S-nitrosation targets, and their Snitrosation levels were relatively higher than the other targets. In the translation process, 8 proteins, including 6 ribosomal proteins and 2 elongation factors, were identified as S-nitrosation targets. These results indicate that S-nitrosation may function by regulating multiple pathways. Recently an iTRAQ-based quantitative method for S-nitrosation detection has been reported, however, it has not yet been applied to endogenous analysis. The advantage of iTRAQ approach is that it can be widely used for analysis of cell, tissue and animal samples. However, since the labeling strategy on peptide was carried out after multi-steps of sample preparation, which may introduce significant quantification error, the parallel and accuracy of quantification were compromised. Being different from it, our SILAC-based ESNOQ method shows significant advantages in the parallel and accuracy of quantification because treatment and control group cells can be mixed as intact cells and processed together throughout the experimental procedure. Therefore, sample losses at a particular step do not affect the quantitative accuracy. The follow-up steps including blocking, reducing, labeling and LC-MS analysis are all performed on the same sample. Therefore, ESNOQ has high accuracy for quantification of endogenous SNOs. The disadvantage of our method is that it can not be easily used for animal and tissue samples. The ESNOQ method described here may be used for analyzing S-nitrosation profiles in cellular LY2784544 processes such as apoptosis or differentiation. It could also be used for dynamic studies by labeling with a range of different isotopes. Moreover, the ESNOQ method lends itself to the study of S-nitrosated modification networks since multiple SNO targets can now be evaluated using the quantitative information obtained. Thus, the ESNOQ method takes us one step closer to revealing the dynamic endogenous roles of S-nitrosation. In internally fertilizing animals sperm are usually accompanied by seminal fluid, forming the ejaculate.