Preliminary investigations on the standardisation and quality control for the determination of gamma-glutamyltranspeptidase activity in seminal plasma

The present study was designed to assess the effects of centrifugation velocity, standing time after dilution, freezing-thawing and chymotrypsin on the determination of gamma-glutamyltranspeptidase (gamma-GT) activities in seminal plasma, and to establish an instruction for the standardisation and quality control for the determination of gamma-GT within the same laboratory and among different laboratories. The gamma-GT level and sperm concentration of each of 72 samples of seminal plasma obtained by centrifugation at 1000 g for 10 min or 3000 g for 15 min were assayed. In addition, gamma-GT activities in diluted seminal plasma with different standing time and in samples with or without chymotrypsin were measured. The results showed that there was a significant difference of gamma-GT levels in seminal plasma obtained by centrifugation at different velocities (P < 0.001), and that gamma-GT activities in seminal plasma measured after standing for 30 min after dilution were notably lower than those measured immediately after dilution (P < 0.001). However, the data indicated that both chymotrypsin and freezing-thawing had no apparent effect on the determination of seminal gamma-GT. In conclusion, standing time after dilution and centrifugation velocity should be standardised, and frozen seminal plasma could serve as quality control products for the determination of gamma-GT activity among different laboratories.     

The effect of chymotrypsin on the determination of total alpha-glucosidase activity in seminal plasma and the correlation between alpha-glucosidase level and semen parameters

To evaluate the effect of chymotrypsin on the examination of alpha-glucosidase activity in seminal plasma, thirty-nine samples of fresh liquefied semen with or without chymotrypsin and forty-eight samples of fresh un-liquefied semen with chymotrypsin were determined for the total alpha-glucosidase activity in seminal plasma. The total alpha-glucosidase level of each sample was assayed by the method of glucose oxidase. The correlations between alpha-glucosidase level and semen parameters, including semen volume, pH, sperm concentration, grade a and b motility and total motility, were analyzed with SPSS 11.0 software. The results showed that chymotrypsin had no effect on seminal alpha-glucosidase activity determination. Chymotrypsin could improve the liquefaction for un-liquefied semen, and there was no significant difference of alpha-glucosidase activity between liquefied and un-liquefied semen samples. There were significantly positive correlations between seminal alpha-glucosidase activity (U/ml) and sperm concentration (r = 0.338, p = 0.015) and between total alpha-glucosidase activity (U/ejaculate) and semen volume (r = 0.677, p = 0.000). However, there was no significant correlation between alpha-glucosidase level (U/ml) and semen volume, pH, sperm motility or grade a and b motility (r = -0.234 approximately 0.077, p = 0.099 approximately 0.993). The data indicated that chymotrypsin could be added into the un-liquefied semen samples for alpha-glucosidase activity determination, and there were different correlations between seminal alpha-glucosidase level and various semen parameters.     

南京市精液分析质量控制的初步研究

目的:调查并分析南京市各家医院对精液分析项目中的精子浓度、精浆果糖、α葡糖苷酶及酸性磷酸酶(ACP)的检测结果,为进一步开展省内乃至全国范围内的精液分析的外部质量控制打下基础。方法:制备低、高浓度的4种检测项目的质控品8份,分装后检测每份质控品的结果并计算变异系数(CV)。质控品分发至南京市11家医院,收集并分析质控品回收结果。以分装后检测结果为参考值,计算不同实验室测得的质控品结果的总相对误差(RE)。结果:8份质控品分装后检测的CV为3.83%~11.16%。11家医院回报的精子浓度结果11份,精浆α葡糖苷酶、果糖和ACP结果5份。不同实验室高、低浓度精浆果糖的检测结果差异最小(CV分别为8.99%和3.95%),其次为α葡糖苷酶(CV分别为16.66%和18.41%),而精浆ACP的检测结果差异最大,CV分别为54.12%和65.58%。RE亦有相同的趋势,不同实验室精浆果糖检测的总RE分别为11.99%和20.31%,α葡糖苷酶为22.92%和27.26%,而ACP为7.34%和318.35%,即高浓度ACP检测结果的RE最大。11家医院中,6家用计算机辅助精液分析(CASA)系统检测精子浓度,5家用改良牛鲍血球计数池的方法检测。血球计数池计数的精子浓度结果[分别为(62.74±16.63)×106/m l和(163.32±36.24)×106/m l]和RE值(分别为148.47%和187.59%)显著高于CASA系统检测的结果分别为[(24.88±4.16)×106/m l和(54.24±23.06)×106/m l]和RE值(分别为13.97%和10.48%)。结论:目前男科学实验室精浆α葡糖苷酶和果糖的检测方法比较稳定,结果的变异在可接受的范围内,而ACP检测的方法可能不适合于临床常规使用,尚需进一步改进。血球计数池明显高估精子浓度,不适合用作精子浓度分析。     

精浆中游离L-肉碱水平与附属性腺生化指标的相关性研究

目的:研究精浆中游离L-肉碱水平与α-葡糖苷酶、果糖、酸性磷酸酶3项附属性腺生化指标之间的相关性,探讨L-肉碱作为一项生化指标在男性生殖功能评价中的作用。方法:采集30例正常生育男性和222例不育患者精液样品,分别以计算机辅助精液分析系统进行精液常规分析,同时测定精浆中游离L-肉碱、果糖水平以及α-葡糖苷酶和酸性磷酸酶活性。用SPSS 12.0统计软件包分析两组间各项生化指标之间的差异,以及精浆中游离L-肉碱与果糖水平、α-葡糖苷酶和酸性磷酸酶活性之间的相关性。结果:正常生育组精浆游离L-肉碱水平明显高于不育组(P<0.01)。两组间α-葡糖苷酶活性差异有统计学意义(P<0.05),而果糖水平和酸性磷酸酶活性差异无统计学意义。相关性分析结果显示,精浆游离L-肉碱水平与精浆中α-葡糖苷酶活性具有较强的正相关关系(r=0.504,P<0.001);而与精浆中果糖水平以及酸性磷酸酶活性之间无相关关系。结论:精浆中游离L-肉碱水平测定可作为评估附睾功能的一项生化指标,其在正常生育组与不育组间的水平差异较两组精浆中α-葡糖苷酶活性差异更为显著,可为男性不育检查及临床诊治和进行有关男性生殖功能机制研究提供参考。     

Evaluation of a commercially available kit for the colorimetric determination of zinc in human seminal plasma

A kit from Wako Pure Chemical Industries for colorimetric determination of zinc has been evaluated for its possible use in the determination of zinc in human seminal plasma. The within-assay variation for 15 replicates of each of two seminal plasma samples having zinc concentrations (mM) of 0.43 +/- 0.025 and 6.06 +/- 0.125 (mean +/- SD) was 5.7% and 2.1%, respectively. The between-assay variation after analysis of 15 replicates of a seminal plasma sample (zinc conc. 5.6 mM) on different days was 2.3%. No interference from other metal ions present in seminal plasma was observed. The average % recovery of zinc added to seminal plasma was 102.7 +/- 1.77 (mean +/- SD). A close correlation (r = 0.996, n = 105) was found between the levels of zinc determined by the colorimetric method and that determined by atomic absorption spectrophotometry as reference method. It is concluded that the present colorimetric method, which is fast, sensitive and linear over the entire concentration range of zinc present in human seminal plasma, can be recommended for use in semen analysis laboratories.     

精索静脉曲张不育患者的精浆生化分析

目的 探讨精索静脉曲张不育患者精浆中酸性磷酸酶、果糖、锌和α-糖苷酶水平的变化.方法 分别检测120例精索静脉曲张不育患者、180例非精索静脉曲张不育患者和36例正常男性的精浆中酸性磷酸酶、果糖、锌和α-糖苷酶含量.结果 精索静脉曲张不育组和非精索静脉曲张不育组精浆中酸性磷酸酶含量均显著低于正常对照组（P＜0.01）,但精索静脉曲张不育组和非精索静脉曲张不育组之间的差异无显著性意义（P＞0.05）;各组精浆果糖活性无显著性差异（P＞0.05）;精浆中锌和α-糖苷酶含量随精索静脉曲张程度的增加而降低,且明显低于正常对照组（P＜0.05）,但与非精索静脉曲张不育组之间的差异无显著性意义（P＞0.05）.结论 精索静脉曲张可通过某些因素引起精浆中酸性磷酸酶、锌和α-糖苷酶含量降低,从而造成男性不育.     

精浆内脂质运载蛋白型前列腺素D合成酶与α-葡萄糖苷酶的相关性研究

目的分析脂质运载蛋白型前列腺素D合成酶(L-PGDS)与精浆其他参数之间的关系,探讨L-PGDS在男性生殖系统中的作用。方法分析92份精液中的精子密度、精子活力、L-PGDS浓度、酸性磷酸酶活力以及α-葡萄糖苷酶活力。依据精子密度,将标本分为3组:正常组(精子密度>20×106/ml)、寡精子组(精子密度<20×106/ml)及无精子组(精子密度为0)。彩色精子质量分析系统测定精子密度及活力,双抗体夹心酶联免疫吸附试验(ELISA)检测精浆内的L-PGDS浓度,分光光度计测定α-葡萄糖苷酶活力。结果正常组、寡精子组以及无精子组患者精浆L-PGDS浓度依次降低,差异显著(P<0.001)。L-PGDS的浓度与α-葡萄糖苷酶、精子密度及精子活力呈正相关,相关系数(r)分别为0.426、0.813和0.380。结论精浆L-PGDS浓度可作为少精子症的辅助诊断指标。     

男性不育症患者精浆酸性磷酸酶与精浆生化指标的相关性研究

目的：探讨男性不育症患者精浆酸性磷酸酶与精浆生化指标的相关性,为精液质量的变化和前列腺炎的诊断、治疗寻找一个可靠的检测指标。方法：收集我院2009年1月～2011年3月治疗的资料完整的男性不育症患者精液分析记录616份,建立Excel数据库,采用SPSSl6．0统计软件,对精浆酸性磷酸酶和精浆生化指标进行相关性分析;并将精浆酸性磷酸酶48．8~208．6U／ml设为正常组,将〈48．8U／mI设为异常组,比较两组之间精浆生化指标的差异性。结果：①精浆酸性磷酸酶与精浆弹性蛋白酶呈负相关（P〈0．05）,与精浆锌呈正相关（P〈0．05）,与精浆a一糖苷酶、果糖无明显相关性（P〉0．05）;②按精浆酸性磷酸酶水平分组比较,显示精浆酸性磷酸酶异常组的精浆弹性蛋白酶明显增高,精浆锌明显降低（P〈0．05）;而两组精浆a一糖苷酶、果糖的表达差异则无统计学意义（P〉O．05）。结论：精浆酸性磷酸酶是前列腺炎可靠判断指标,与精浆锌、精浆弹性蛋白酶一道可作为生殖道感染及精液质量变化的检测指标。、     

半自动生化分析仪检测精浆α葡糖苷酶活性的研究

目的:建立半自动生化分析仪检测精浆α葡糖苷酶活性的方法。方法:同时用半自动生化分析仪法(速率法)和手工葡萄糖氧化酶法检测51例精液常规分析参数均正常的男性精浆α葡糖苷酶活性,计算批内和批间变异系数以及正常参考值范围,并分析两种方法检测结果的相关性。结果:2例α葡糖苷酶活性不同的精浆样本用半自动生化分析仪法检测的批内变异系数分别为12.63%和9.13%,批间变异系数分别为10.67%和13.49%,正常参考值范围为102.28～555.08U/L。半自动生化分析仪法和手工法检测的精浆α葡糖苷酶活性呈显著正相关(r=0.792,P<0.01)。结论:半自动生化分析仪检测精浆α葡糖苷酶活性法简便、省时,节省试剂,结果稳定可靠。建议在男科实验室推广使用半自动生化分析仪法检测精浆α葡糖苷酶活性。     

全自动精浆γ-L-谷氨酰转肽酶检测方法的建立及评价

目的:建立精浆γ-L-谷氨酰转肽酶(GGT)全自动检测方法并对其准确性、重复性和线性范围等进行评价。方法:利用速率法检测精浆GGT活性,并在全自动生化分析仪上建立检测参数,并评估该方法的试剂空白吸光度、准确性、重复性和线性范围,同时与目前临床上常用的精浆GGT检测化学比色法试剂盒(南京欣迪)进行比较。结果:试剂空白吸光度平均为0.047 6,空白吸光度变化率(△A/min)平均为0.000 168。3份高、中、低GGT活性的精浆样本重复测定10次的变异系数值分别为0.26%、4.83%和1.60%。采用比对试验的方法来评价准确性,40份精浆标本每个浓度点的相对偏差范围为-13.38%¹1.05%,均低于15%的要求。精浆GGT活性在29911.05%,均低于15%的要求。精浆GGT活性在299¹ 833 U/L范围时,具有良好的线性关系(r>0.99)。以精浆GGT检测化学比色法试剂盒作为对照试剂,全自动检测法作为试验试剂,两者对115例精浆样本的检测结果呈显著正相关(r=0.981,P<0.01),Kappa值为0.776(P<0.05),符合率为90.43%。结论:本研究建立的精浆GGT全自动检测法有较低的试剂空白,重复性和准确性较好,且与目前临床上使用的化学比色法有很好的一致性。该法操作简单、快速,精浆样本无需稀释,适合临床上大批量样本的检测筛查,大大节省了人力和试剂成本。     

The accuracy of sperm concentration determination by the automated CellSoft semen analyzer before and after discontinuous Percoll gradient centrifugation: Die Genauigkeit der Spennatozoendichte-Bestimmung mittels der automatischen Cell-Soft-Samenanalyse vor und nach diskontinuierlicher Percoll-Gradienten-Zentrifugierung

The automated CellSoft semen analyzer identifies human spermatozoa on the basis of user-defined values for cell size and luminosity. Previous studies have shown that the non-sperm particles usually present in seminal plasma would interfere with the computerized determination of sperm concentration by the CellSoft system. Therefore, an effective method to separate the sperm from non-sperm particles would be desirable to obtain accurate determination of sperm concentration. In the present study, sperm concentrations in 72 semen samples before and after discontinuous Percoll gradient centrifugation were determined by the automated CellSoft system and the results compared with those obtained with the routine procedure using the haemocytomer according to the World Health Organization laboratory manual. The computerized measurement caused a significant overestimation when the sperm concentration in semen was less than 80 x 10(6)/ml. Processing of human semen sample by the simple two-layer discontinuous Percoll gradient centrifugation removed the majority of non-sperm particles and the overestimation by the automated CellSoft system became significant only when the sperm concentration in the final Percoll sperm preparation was less than 10 x 10(6)/ml. These findings indicate that the automated CellSoft semen analyzer has to be improved to allow for the correction of non-sperm particles in seminal plasma or processed sperm samples before it can be used to provide sufficiently accurate sperm concentration results for routine laboratory service or research purposes.     

Comparison of glycosidase levels in bovine seminal plasma

Seven glycosidases (beta-N-acetylglucosaminidase, alpha-fucosidase, beta-galactosidase, acid alpha-glucosidase, beta-glucuronidase, acid and neutral alpha-mannosidase) were analysed in seminal plasma from the first and second successive ejaculates in normal Ayrshire bulls. In comparison to our previous data the results indicate that beta-N-acetylglucosaminidase, beta-galactosidase and beta-glucuronidase are derived mainly from epididymal secretions, while alpha-fucosidase and particularly neutral alpha-mannosidase originate additionally from the spermatozoan cytoplasmic droplets. The seminal vesicles appear to contribute particularly to the seminal plasma acid alpha-glucosidase and acid alpha-mannosidase activities. The seminal plasma enzymes derived from the epididymis and cytoplasmic droplets were suppressed in semen samples with low sperm density or with high numbers of abnormal spermatozoa. The epididymal and seminal vesicle enzymes could be utilized in assessment of the secretory/functional capacity of these glands.     

Prolactin and α-1, 4-glucosidase activity in normal and poorly coagulated human semen

Prolactin and alpha-1,4-glucosidase levels in seminal plasma were measured in poorly coagulated (I), deficiently coagulated (II) and normally coagulated (III and IV) human ejaculates having 0-20%, 21-50% and 51-100% coagulum respectively 4 min after emission. The prolactin concentration (ng ml-1, mean +/- SEM) in poorly coagulated (5.2 +/- 0.48) and deficiently coagulated (7.6 +/- 0.72) samples was significantly lower than in the normally coagulated groups III (51-75% coagulum, 8.2 +/- 0.43) and IV (76-100% coagulum, 9.9 +/- 0.59) as well as the presumably fertile samples (9.2 +/- 0.74). A highly significant positive correlation was observed between the prolactin level and the percentage coagulum of the ejaculates (r = 0.686, n = 58, P less than 0.001). In contrast, the epididymal marker, alpha-glucosidase showed no relationship to seminal coagulation.     

Sperm motility inhibitor from human seminal plasma: presence of a precursor molecule in seminal vesicle fluid and its molecular processing after ejaculation

Human seminal plasma contains a protein factor that has the capacity to inhibit the movement of demembranated and intact spermatozoa. This factor 'seminal plasma motility inhibitor' (SPMI) has been shown to originate exclusively from the seminal vesicles. The present results demonstrate that the biological activity of SPMI in semen decreases rapidly from 1000 U/d, immediately after ejaculation, to 220 U/ml 2 h later. Immunoblots of seminal plasma proteins probed with an antibody against human SPMI, revealed the rapid processing of a predominant 52 kDa SPMI antigen, present in the seminal vesicle secretions. This precursor was degraded initially into intermediate molecular mass fragments of 25–40 kDa, and subsequently into smaller fragments of 17–21 kDa. When seminal vesicle fluid was mixed with prostatic secretions (3: 1 v/v), proteases present in prostatic secretions were shown to be responsible for processing of the SPMI precursor. Addition of protease inhibitors such as phenylmethylsulphonyl fluoride (PMSF, 5 mM), benzamidine (100 mM) or ethylenediaminetetraacetic acid (EDTA, 5 mM) to the mixture of seminal vesicle and prostate secretions partially prevented the loss of activity of SPMI by 54%, 27% and 9%, respectively. However, the simultaneous addition of PMSF and benzamidine conferred almost total stability to the SPMI precursor activity. These results demonstrate that SPMI exists as a predominant 52 kDa precursor form in the seminal vesicles and is processed rapidly after ejaculation into less active, lower molecular mass forms by one or more serine proteases and/or metallo-proteases of prostatic origin which are present in liquefied semen.     

Hemoglobin test result variability and cost analysis of eight different analyzers during open heart surgery

The purpose of this study was to compare the variation in hemoglobin (Hgb) values among various point-of-care (POC) analyzers available on the market. Eight analyzers (Gem 3000, ABL 720, ABL 77, Rapidpoint 405, IL 682, GemOPL, Hb 201+, and manual/centrifugation) were compared with the Hgb values from the Beckman Coulter LH750. A total of 72 patient samples were analyzed on each test instrument. The samples were obtained after intubation, after heparinization, during cardiopulmonary bypass, and after protamine administration. Four of the samples were excluded from the study because of delayed sample analysis. The calculated mean differences of reference test method Hgb (mean +/- SD) for all samples (n = 68) were Gem 3000 = 1.431 +/- 0.396 g/dL; ABL 720 = -0.224 +/- 0.240 g/dL; ABL 77 = 0.341 +/- 0.578 g/dL; Rapidpoint 405 = 0.001 +/- 0.205 g/dL; IL 682 = -0.137 +/- 0.232 g/dL; GemOPL = 0.774 +/- 0.427 g/dL; Hb 201+ = 0.110 +/- 0.524 g/dL; and manual/ centrifugation = 0.547 +/- 0.499 g/dL. Cumulative results indicated that the bias in Hgb values from the Gem 3000, ABL720, ABL 77, IL 682, GemOPL, and the manual method were statistically significant (p < .05), compared with the Coulter LH750. Additionally, only the Rapidpoint 405 and Hb 201+ most closely matched the values from the Coulter LH750 (p > .05). Some of the methodologies have previously been shown to be affected during hemodilution, hypoproteinemia, and/or after blood transfusion. There is variability among methodologies, which can give rise to statistically different Hgb values, and one should consider the "ideal" instrument based on this and many other factors. Based on our results, the rank order of closest approximation to the Coulter LH750 measurement was Rapidpoint 405, Hb 201+, IL 682, ABL 720, ABL 77, manual/centrifugation, GemOPL, and Gem 3000.     

精浆α葡糖苷酶全自动检测方法的建立及评价

目的:建立精浆α葡糖苷酶活性全自动检测方法并对其准确性、重复性和线性范围等进行评价。方法:利用速率法检测精浆α葡糖苷酶活性,并在全自动生化分析仪上建立检测参数,并评估该方法的试剂空白吸光度、准确性、重复性和线性范围。结果:试剂空白吸光度平均为0.002 14,空白吸光度变化率(△A/min)平均为0.015 26。3份高、中、低α葡糖苷酶活性的精浆样本重复测定10次的CV值分别为2.52%、0.36%和0.76%。采用比对试验的方法来评价准确性,40份精浆标本每个浓度点的相对偏差范围为-5.52%~7.54%,均低于15%的要求。精浆α葡糖苷酶活性在76~647 U/L范围时,具有良好的线性关系(r0.99)。结论:本研究建立的全自动精浆α葡糖苷酶活性检测法有较低的试剂空白,重复性和准确性较好,且有较宽的线性范围。该法操作简单、快速,精浆样本无需稀释,适合临床上大批量样本的检测筛查,大大节省了人力和试剂成本。     

精浆尿酸的检测及其与精液参数的相关性研究

目的:建立精浆尿酸(UA)的检测方法并探讨精浆UA水平与精液参数的相关性。方法:根据检测血清UA的方法加以改良建立检测精浆UA的方法,并观察批内变异和不同技术人员检测结果之间的差异以评价方法的可接受性。同时分析138例男性精浆UA水平与患者年龄、禁欲时间、精液量、pH、精子密度、活动率、a+b级精子百分率和正常形态精子百分率之间的相关性。结果:精浆UA检测方法的批内变异为9.16%,2位技术人员的检测结果没有显著性差异(P=0.541)。精浆UA水平与正常形态精子百分率呈正相关(r=0.350,P=0.025),与精子活动率和a+b级精子百分率有呈正相关的趋势(r=0.147和0.156,P=0.085和0.068),而与精液分析其他参数如精液量、pH、精子密度、禁欲时间以及患者年龄等无相关性。结论:通过对血清UA检测方法加以改良可以建立可接受的精浆UA检测方法。精子形态学、活动率及a+b级精子百分率可能与精浆UA水平高低有关。     

Why do we determine α-glucosidase activity in human semen during infertility work-up?

alpha-Glucosidase is a normal constituent of human semen, produced mainly in the epididymis. It is significantly correlated to sperm count. Its activity is low in cases of epididymal obstruction. We evaluated alpha-glucosidase activity in 653 semen samples of patients, who attended our department for marital infertility, with respect to associations to clinical and other seminal parameters. The normal range (mean +/- 2 SD) in samples with normal parameter values was 7.2-46.4 mU ml-1. The determination in patients with azoospermia revealed mean values of 7.7 +/- 9.5 mU ml-1 in obstructive azoospermia, and 15.8 +/- 11.5 mU ml-1 in nonobstructive azoospermia. The difference was not statistically significant in that the sensitivity of determination with respect to the presence of obstruction was only 0.66, and the specificity 0.83. A significant correlation (r = 0.34) of alpha-glucosidase activity with log sperm count was observed. The mean alpha-glucosidase activity was not significantly different in groups formed according to sperm motility, according to leucocyte count or according to semen volume. A difference between smokers and nonsmokers with comparable sperm count, as reported in the literature, did not occur. We conclude from our results that the determination of alpha-glucosidase activity does not give additional information of the fertility status exceeding that of other clinical investigations or parameters of semen analysis.     

精浆酸性磷酸酶和γ-L-谷氨酰转肽酶检测的比较及其与精液参数的相关性研究

目的:对精浆酸性磷酸酶(ACP)和γ-L-谷氨酰转肽酶(γ-GT)活性检测进行比较,并分析ACP和γ-GT活性与精液参数的相关性。方法:133例精浆标本,分别检测ACP活性和γ-GT活性。随机留取2例精浆标本,1例用于ACP批内检测,另1例用于γ-GT批内检测。随机留取4例标本,2例用于ACP批间检测,另2例用于γ-GT批间检测。用计算机辅助的精液分析(CASA)系统分析精液标本的精液量、pH、精子密度、活动率、a+b级活动精子百分率等参数。同时分析ACP和γ-GT活性与精液参数的相关性。结果:精浆ACP活性和γ-GT活性呈显著性正相关(r=0.570,P=0.000)。ACP的批内变异系数(CV)为13.72%,批间CV分别为13.80%和15.49%。γ-GT批内CV为7.68%,批间CV分别为7.76%和9.73%。精浆ACP活性和γ-GT活性均与pH值呈显著负相关(r=-0.330,P=0.000;r=-0.388,P=0.000)。γ-GT活性与精子密度呈显著正相关(r=0.165,P=0.045),而ACP活性与精子密度无显著相关(r=0.048,P=0.546)。ACP活性和γ-GT活性均与精子活动率、(a+b)级活动精子百分率、精液量、禁欲时间以及年龄无显著相关性。结论:精浆γ-GT活性检测的精确性高于ACP活性检测,两者与精液参数的相关性基本类似。提示精浆γ-GT活性检测比ACP活性检测更适合用来评价前列腺功能。     

Quick recovery and characterization of cell-free DNA in seminal plasma of normozoospermia and azoospermia： implications for non-invasive genetic utilities

We established a quick and reliable method for recovering cell-free seminal DNA （cfsDNA）, by using the binding-washing-elution procedure on the DNA purification column. Low variations （below 15%） among the triplicate values of cfsDNA quantity verified the reproducibility of our cfsDNA recovery method. Similar cfsDNA yield and size distribution between seminal plasma acquired by filtration and centrifugation confirmed the presence of cfsDNA. To investigate the general characterization of cfsDNA, the quantitation and size distribution of cfsDNA from normozoospermic and azoospermic semen were analyzed by real-time PCR and electrophoresis, respectively. CfsDNA concentration in semen with normozoospermia （n = 11） was 1.34 ± 0.65 μg ·mL^-1, whereas a higher cfsDNA concentration was observed in azoospermia （2.56 ± 1.43 μg ·mL^-1, n = 9）. The continuous distribution of DNA fragments ranging from -1 kb to 15 kb and a spectrum of multiples of 180-bp fragments were observed in each normozoospermic and azoospermic sample. Distinct characteristic DNA ladder fragmentations in some azoospermic samples implicated that cfsDNA originate partly from apoptotic cells. CfsDNAs of 36 selected azoospermic patients with known information of Y chromosome microdeletion were subjected to the same microdeletion analysis by multiplex PCR and PCR amplification of sY114 （1 450 bp）. All multiplex PCR reactions with cfsDNA amplified successfully and provided the same result as leukocyte DNA. PCR amplification of sY114 gave a 1 450-bp amplicon as expected. Our data suggested the potential use of cfsDNA in search of biomarker or diagnostic procedures.