de Souza F F, Barreto C S, Lopes M D
FertCani, Serviço Veterinário Especializado em Reprodução de Pequenos Animais, Incubadora Tecnológica de Botucatu, Botucatu, SP, Brazil.
Theriogenology. 2007 Jul 1;68(1):100-6. doi: 10.1016/j.theriogenology.2006.11.016. Epub 2007 May 21.
The objectives were to separate canine seminal plasma proteins (with SDS-PAGE) and to determine the correlation between specific proteins and semen characteristics. Three ejaculates from 20 mixed-breed dogs, of unknown fertility, were collected by digital manipulation. Ejaculate volume and color, sperm motility, sperm vigor, percentage of morphologically normal spermatozoa, and membrane integrity (hypoosmotic swelling test and fluorescent staining) were assessed. For each dog, seminal plasma was pooled from all three ejaculates and proteins were separated with SDS-PAGE, using polyacrylamide concentrations of 13% and 22% in the separation gels. After staining, gel images were digitized to estimate molecular weights (MW) and integrated optical density (IOD) of each lane and of individual bands. Total seminal plasma protein concentration was 2.19+/-1.56 g/dL (mean+/-SD; range 1.12-5.19 g/dL). A total of 37 protein bands were identified (although no dog had all 37 bands). In the 13% gel, molecular weights ranged from 100.6 to 17.1 kDa, with four bands (49.7, 33.2, 26.4, and 19.5 kDa) present in samples from all dogs. In the 22% gel, molecular weights ranged from 15.6 to 3.6 kDa, with nine bands (15.6, 13.5, 12.7, 11.7, 10.5, 8.7, 7.8, 5.6, and 4.9 kDa) present in samples from all dogs. Combined for both gels, the majority of bands (85%) had molecular weights <17 kDa, with B20 (15.6 kDa) in high concentrations in samples from all dogs. There were positive correlations (P < or = 0.01) between two bands, B4 (67 kDa) and B5 (58.6 kDa), and sperm motility (r=0.66 and r=0.46), sperm vigor (r=0.56 and r=0.66), percentage of morphologically normal spermatozoa (r=0.55 and r=0.59), the hypoosmotic swelling test (r=0.76 and r=0.68), and fluorescent staining (r=0.56 and r=0.59), respectively. In conclusion, 37 proteins were identified in seminal plasma; two were significantly correlated with semen characteristics.
目的是（通过十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳）分离犬精液中的蛋白质，并确定特定蛋白质与精液特征之间的相关性。通过手法采集了20只生育能力未知的混种犬的三份射精样本。评估了射精量、颜色、精子活力、精子活率、形态正常精子的百分比以及膜完整性（低渗肿胀试验和荧光染色）。对于每只犬，将三份射精样本的精浆混合，使用分离胶中聚丙烯酰胺浓度为13%和22%的十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳分离蛋白质。染色后，将凝胶图像数字化以估计每条泳道和各个条带的分子量（MW）和积分光密度（IOD）。精浆总蛋白浓度为2.19±1.56 g/dL（平均值±标准差；范围1.12 - 5.19 g/dL）。共鉴定出37条蛋白条带（尽管没有一只犬具有全部37条带）。在13%的凝胶中，分子量范围为100.6至17.1 kDa，所有犬的样本中均存在四条条带（49.7、33.2、26.4和19.5 kDa）。在22%的凝胶中，分子量范围为15.6至3.6 kDa，所有犬的样本中均存在九条条带（15.6、13.5、12.7、11.7、10.5、8.7、7.8、5.6和4.9 kDa）。综合两种凝胶来看，大多数条带（85%）的分子量<17 kDa，所有犬的样本中B20（15.6 kDa）浓度较高。两条条带B4（67 kDa）和B5（58.6 kDa）与精子活力（r = 0.66和r = 0.46）、精子活率（r = 0.56和r = 0.66）、形态正常精子的百分比（r = 0.55和r = 0.59）、低渗肿胀试验（r = 0.76和r = 0.68）以及荧光染色（r = 0.56和r = 0.59）之间分别存在正相关（P≤0.01）。总之，在精浆中鉴定出37种蛋白质；其中两种与精液特征显著相关。
