The seminal plasma proteins Peptidyl arginine deaminase 2, rRNA adenine N (6)-methyltransferase and KIAA0825 are linked to better motility post thaw in stallions

Comprehensive Flow-Cytometric Quality Assessment of Ram Sperm Intended for Gene Banking Using Standard and Novel Fertility Biomarkers

“…Moreover, the Group 1 involved samples with significantly higher sperm concentration than Group 2 ( Figure 2 ). Similar distribution of semen samples according to sperm viability and motility was performed in the study focused on the quality of thawed stallion spermatozoa [ 44 ]. The viability of ram spermatozoa in this study was assessed by the SYBR-14 probe, which is most widely used in combination with PI or 7-AAD [ 5 ].…”

“…However, to distinguish dead spermatozoa in our samples, a novel far-red dead cell dye DRAQ7 was employed into presented flow-cytometric analysis to fully eliminate the possible spectral overlap of used fluorescent dyes. This dye was successfully used previously for the viability assessment of ram or stallion spermatozoa [ 44 , 45 ]. On the other hand, SYTOX Green, a green dead cell dye, was used in combination with red fluorescent dyes to analyze some sperm-specific attributes in this study.…”

“…Similarly, Caspase 3/7 probe was useful for apoptosis detection in fresh ram sperm samples exposed to different centrifugal forces [ 51 ] or in frozen-thawed samples with various antioxidant supplementation [ 52 ]. Moreover, this probe found significantly more positive spermatozoa in cryopreserved stallion semen samples with poor viability and motility [ 44 ]. In addition, these stallion samples also displayed significantly decreased mitochondrial activity (high MMP observed by JC-1 dye) in poor samples compared to samples with good viability, similar to our study comparing ram semen samples with good (Group 1) or poor (Group 2) viability ( Figure 4 ).…”

Dataset of the sperm proteome of stallions with different motility

“…The same protocol as described in [3] , [4] , [5] , [6] , [7] , [8] , [9] was used. Stallion ejaculates were washed in PBS (600gx 10’), then the pellet formed by the spermatozoa was frozen immediately in liquid nitrogen and kept frozen at -80 °C.…”

“…The separation and analysis were conducted in an Agilent 1290 Infinity II Series UHPLC (Agilent Technologies, Santa Clara, CA, USA) equipped with an automated multisampler module and a High Speed Binary Pump, coupled to a Mass Spectrometer (Agilent 6550 Q-TOF, Agilent Technologies, Santa Clara, CA, USA) using an Agilent Jet Stream Dual electrospray (AJS-Dual ESI) interface, controlled by the MassHunter Workstation Data Acquisition software (Agilent Technologies, Rev. B.06.01), following previously published protocols [ 2 , 4 , 5 , 9 ]. The digested proteins were injected onto an Agilent Advance Bio Peptide Mapping HPLC column (2.7 μm, 150 × 2.1 mm, Agilent technologies), at 55 °C, at a flow rate of 0.4 ml/min.…”