混合斑中精斑ABO血型的检验

<正> 1988年,壹岐裕志等报告了用吸附抗α_2-SGP 血清的硝化纤维素膜(NCF)检验混合斑中的精斑 ABO 血型,但耗时。本文作者通过对此方法的改进,采用常彩琴等研制的抗人精特异蛋白血清(anti-human seminalpeculiar protein,ASPP),采用蛋清粘片热解离法检验混合斑中精斑 ABO 血型,耗时短,效果好。现介绍如下。     

A sandwich enzyme-linked immunosorbent assay for human seminal γ-glutamyl transpeptidase

A sandwich enzyme-linked immunosorbent assay (ELISA) was developed for detecting human seminal γ-glutamyl transpeptidase (γ-GTP) using a combination of anti-seminal γ-GTP monoclonal antibodies. These monoclonal antibodies did not react with human ovary or uterus in immunohistochemical study. Optimal assay condition, resulting in a sensitive assay with a low background, is presented. The detection limit of this assay was estimated to be 1 ng/ml of seminal γ-GTP corresponding to about 100 000 times dilution of seminal sample. This ELISA was specific for seminal γ-GTP, without cross-reactivity to renal or hepatic gg-GTP, normal blood serum, non-coital vaginal fluid or saliva. The recovery of seminal γ-GTP added to various biological fluids were also examined.     

自制与商业化抗人精液蛋白P30金标试剂条的法医学应用比较

目的　检测比较自制和商业销售的抗人精液蛋白P30 金标试剂条 ,探讨它们法医学应用的可靠性。方法　以人精液和人前列腺特异蛋白检测效价及灵敏度 ,以人阴道分泌物、人血清、人唾液和人初乳检测特异性 ,以保存不同年限的陈旧人精斑和斑痕类检材测定检验实际检材的能力。结果　自制抗人精液蛋白P30 金标试剂条效价及灵敏度与商业销售的试剂条中较好的基本相当 ,特异性则相当或好于商业试剂条 ,对非特异性测定均为阴性 ,而商业化试剂条则多有非特异性反应。结论　使用商业金标试剂条 ,要确定其是否合乎法医学检验标准与要求。     

The detection of inherited enzyme polymorphism in semen (author's transl)]

Our investigation of the occurrence of the enzymes phosphoglucomutase (PGM), glutamate-pyruvate-transaminase (GPT), adenylatkinase (AK), adenosine-desaminase (ADA), and 6-phophogluconate-dehydrogenase (6-PGD) produced the following results: The phosphoglucomutase type was demonstrated in the most sperm samples and seminal stains in accordance with the corresponding blood type. This enzyme is rather stable and could still be demonstrated well in 1-month old stains. The glutamate-pyruvate-transaminase can only seldom be determined in semen and seminal stains. We only found the GPT 1 type, which is known to have usually the strongest activity. The adenylatekinase was demonstrable in the most fresh ejaculates (not older than 24 h) and in about half the seminal stains (not older than 7 days)--The AK--2-band gets weak with increasing lay days, which may lead to incorrect determinations. The adenosine-desaminase could not be determined in sperm. On the contrary, 6-phosphogluconate-dehydrogenase could be demonstrated in fresh semen samples and also partly in seminal stains up to 7 days. The demonstration of the enzymes did not depend in any system on the secretor type.     

Glucose phosphate isomerase variants in human bloodstains and seminal stains

Glucose phosphate isomerase (GPI) variants occurring in human red cells were also demonstrated in human semen. Phenotyping was possible from bloodstains of 6 weeks storage and seminal stains of 12 weeks storage. The GPI system may be a supplemental tool for medicolegal individualization of seminal stains.     

The LAP test. A new method for identification of seminal stains by a qualitative color reaction of leucine aminopeptidase.

A new simple method for identification of seminal stains is described. It employs a qualitative color reaction based on histochemical technique for demonstration of leucine aminopeptidase (LAP), which is extremely abundant in human semen. The method herein reported (the LAP test) is quite suitable for medicolegal examination of seminal stains as a preliminary test.     

A multi-enzyme electrophoretic system for the identification of seminal fluid from postmortem specimens

The screening of autopsy specimens, vaginal, buccal, and rectal swabs, for the presence of seminal fluid in rape homicide investigations utilizing classical techniques can lead to erroneous results. In the absence of spermatozoa, techniques are needed which can help to identify seminal fluid. This report illustrates the use of a multi-enzyme electrophoretic approach identifying seminal acid phosphatase (SAP) and lactic acid dehydrogenase (LDH-X) as an initial screening procedure. Subsequent analyses for the presence of acid and alkaline phosphatase (semiquantitative) yield information which can help identify false-positive SAP's. Additionally, salivary amylase can be tentatively identified using the same multi-enzyme procedure which informs the investigator of possible salivary contamination of the sample and possible erroneous PGM results. Statistics utilizing the multi-enzyme approach in case work are also presented.     

alpha-L-fucosidase phenotyping in human placentae, semen and seminal stains

The polymorphism of alpha-L-fucosidase (Fu) was investigated in a Japanese population from samples of placentae and semen, using isoelectric focusing. The gene frequencies of placental types were Fu1 = 0.748 and Fu2 = 0.252, and those of seminal types were Fu1 = 0.739 and Fu2 = 0.261. The coincidence in the distribution between the placental and seminal types suggests that the Fu types occurring in placentae and in semen are controlled by the same Fu alleles. The Fu typing was possible in seminal stains stored at 4 degrees C for up to 9 weeks, at room temperature for up to 7 weeks and at 37 degrees C for up to 4 weeks. The Fu types were still detectable at semen dilutions of up to 1:4. This polymorphism would provide a useful genetic marker for the medicolegal grouping of seminal stains.     

Detection of human seminal γ-glutamyl transpeptidase in stains using sandwich ELISA

A sensitive and specific sandwich ELISA for human seminal γ-glutamyl transpeptidase (γ-GTP) was developed using a combination of monoclonal antibodies, SG1 and SG3, which we produced. For semen identification in forensic samples, we modified the assay so as to be more sensitive and to establish efficient extracting conditions. After testing the extracting abilities of several detergents, CHAPS and deoxy-BIGCHAP were chosen as the solubilizer. Polystyrene beads coated with SG1 were incubated with samples extracted by the detergents, and further with biotinylated SG3, followed by peroxidase-labeled streptavidin. γ-GTP was detected only in seminal samples. The sensitivity of this assay was 0.01 ng/ml of seminal γ-GTP equivalent to 10⁷ times diluted semen, which was ten times as compared with the previous plate assay. No significant seminal γ-GTP was detected in other biological stains such as blood, saliva and vaginal smear. The extract of a 500 fold diluted seminal stain, 8 months old, showed the detection limit. Seminal γ-GTP was detectable even in 14-year-old stains.     

用酶标抗人精液单克隆抗体A_(10)C_6建立ELISA斑点法快速鉴定人精(液)斑

作者用过氧化物酶标记自己研制的抗人精液单克隆抗体A10C6建立了简便快速鉴定人精斑的ELISA斑点法，直接通过酶标抗体上的酶催化底物显示颜色变化，来判断结果，阳性呈灰色斑点，阴性为无色。其结果只对人精斑浸出液显示阳性斑点，对人的其他组织器官及体液均呈阴性，对动物精斑也无交叉反应，精斑浸出液稀释到1：3200倍仍呈阳性结果。     

分泌抗人精液特异蛋白P_(30)单克隆抗体杂交瘤细胞系的建立和初步应用

作者通过杂交瘤技术建立了9株产生抗精浆特异蛋白 P_(30) 单克隆抗体的杂交瘤细胞系。它们是由 SP2/0骨髓瘤与经 P_(30) 免疫的 BALB/C 小鼠脾细胞按常规方法进行细胞融合、并经克隆化筛选得出.这些细胞株均经体外培养3个月以上能够稳定分泌抗 P_(30) 单克隆抗体。该抗体只能识别纯化的 P_(30) 和精液中的 P_(30) ;与人精液以外的其他体液和多种人体组织无交叉反应;与几种常见动物的精液和血液无交叉反应。这些 P_(30) 单克隆抗体均属 IgG 类和 IgG_1亚类。其培养上清液和腹水的抗体效价最高分别达到320和128,000。以 ELISA 法应用这些单克隆抗体能很好地鉴定精液和精斑。     

Semen finger print.

A correlation between testosterone (T) and dihydrotestosterone (DHT) exists in seminal plasma. These androgens play a role in sperm maturation. The T/DHT ratio is different from one person to the other, due to the heterogenicity of seminal plasma which stems for the most part from the male accessory sex glands, the prostate and seminal vesicles, and also from the 'epididymo-testicular unit'. This ratio is useful in identifying the person's semen. Consequently, the steroid values from assailant semen or accused persons and semen on the victim's clothes are of cardinal importance in matching. The results reported include data on the validation of this technique as a tool for semen identification. Coital and masturbated semen were correlated, and consecutive coital semen were also analysed. It may be concluded that the T/DHT value is specific for each person.     

人精浆蛋白双向谱型特征及其器官特异性的研究

本文应用双向聚丙烯酰胺凝胶电泳（2D—PAGE）技术分析了60例正常人精浆中的蛋白成分。阐明了人精浆蛋白双向电泳谱型的特征，并将人精浆蛋白双向电泳谱型与其他体液蛋白双向谱型进行了比较，证明人精浆蛋白双向电泳谱型具有器官特异性。本文还用2D－PAGE对不同条件下保存的精斑进行了认定。并将其应用于强奸案的鉴定。     

gamma-Glutamyltransferase test as a new tool for identification of seminal stains

A simple qualitative method for identification of seminal stains based on a high activity of gamma-glutamyltransferase (gamma-GTP) in human semen is described. It employs the release of alpha-naphthylamine from N-gamma-glutamyl-alpha-naphthylamide by the gamma-GTP action: alpha-naphthylamine couples with Fast Garnet GBC salt to produce a strong brownish-red color. The data on its simplicity, specificity, and stability show that the present method is suitable for medicolegal examination of seminal stains as a preliminary test.     

PCR-RFLP法检验混合斑中精斑ABO血型及基因型

运用PCR-RFLP检验方法,对21份精斑样品、5份混合斑检材进行了检验,结果表明该方法能够很好地检测混合斑中精斑的ABO血型及基因型,有效地解决了非分泌型精斑的ABO血型及基因型,拓宽了混合斑中精斑的检验范围.     

The use of scanning electron microscopy in the examination of seminal stains

Samples of human seminal stains on fabrics, paper and wood were observed with a scanning electron microscope (S.E.M). It seems a reliable, rapid and simple technique for the routine examination of seminal stains.     

The simultaneous identification of seminal acid phosphatase and phosphoglucomutase by starch gel electrophoresis

Although elevated acid phosphatase (AP) activity in vaginal fluid is a consistent indicator for semen, differentiation between vaginal AP and seminal AP provides a more meaningful result. Detection of seminal AP in mixtures of vaginal AP, feces, and blood is accomplished by starch gel electrophoresis, employing the substrate thymolphthalein monophosphate as a selective visualization agent. Genetic phosphoglucomutase isoenzymes are simultaneously separated by this method and allow differentiation in some semen/vaginal fluid mixtures.     

用ELISA检测精液及精斑中精子抗原的研究

本实验应用单克隆抗人精子抗体和酶标记羊抗人精子抗体,采用ELISA方法确定精子抗原成份的存在。对10份新鲜精液,15份精斑进行了验测,其结果阳性率为100%。新鲜精液(精子数约10,000万个/ml)稀释100万倍,精斑浸出液稀释50万倍,均可出现阳性。对唾液斑、尿斑、乳汁斑、阴道斑、汗斑及输精管结扎的精液均为阴性。实验结果表明,本法检验精子抗原具有灵敏度高,特异性好的优点。     

人精浆特异蛋白P_30的分离纯化与鉴定

本文介绍人精浆特异蛋白P_(30)分离纯化及鉴定的方法。人精浆经蒸馏水透析后经Sephadex G100、G150和G75三种不同的柱层析即可获得精浆特异蛋白。SDS-PAGE、免疫电泳、免疫双扩散证明其纯度和特异性均符合要求。它与已知抗P_(30)血清呈强阳性反应,用其免疫制备的抗血清和已知抗P_(30)血清,在免疫电泳中只与精浆或精浆特异蛋白形成一条完全相同的沉淀线,SDS-PAGE测得分子量约为30,000。故笔者将其亦定名为P_(30)。     

一种改良酸性磷酸酶试验鉴定精斑的方法

本文报道了一种改良的酸性磷酸酶试验方法。用该法鉴定精斑，可克服非特异性反应，排除其它体液和植物汁液的影响。