流式细胞术在细胞凋亡研究中的应用

流式细胞术做为一种高新生物技术,在动物医学的各个领域,包括细胞生物学、病毒学、肿瘤学、免疫学和病理学中都得到广泛应用,为细胞凋亡研究提供了有效的技术手段。该技术具有简便、快速、多参数分析等优点,可针对细胞在凋亡时产生的一系列形态学、生物化学及分子生物学性质的变化,包括细胞皱缩,核染色质凝聚,细胞膜通透性改变,Caspases 激活,线粒体跨膜电位降低,膜磷酯酰丝氨酸外化,胞质 Ca2 浓度升高,DNA片段化及含量变化等特点,进行定性、定量测试分析,从而实现对细胞凋亡的准确测定。     

犬子宫蓄脓的临床诊治

犬子宫蓄脓是目前临床上常见的产科病之一,在宠物饲养中危害较大,2009年3～5月,乌鲁木齐市畜牧科学院动物医院共接诊了40例产科病,其中犬子宫内蓄脓占10例.本文结合这10例病例,从该病的发病原因、症状表现、诊断方法和治疗方面进行阐述,并就相关内容进行讨论.     

Kinematic Motion Patterns of the Cranial and Caudal Canine Cervical Spine

Objective: To define the kinematic motion patterns of the canine cervical spine, with a particular emphasis on identifying differences between the cranial (C₂–C₄) and caudal (C₅–C₇) segments, and to determine the significance of coupled motions (CM) in the canine cervical spine. Study Design: Cadaveric biomechanical study. Sample Population: Cervical spines of 8 Foxhounds. Methods: Spinal specimens were considered free of pathology based on radiographic, computed tomography, and magnetic resonance imaging examinations. All musculature was removed without damaging ligaments or joint capsules. Spines were mounted in a customized pure‐moment spine testing jig, and data were collected using an optoelectronic motion capture system. Range of motion, neutral zone and CM in flexion/extension, left/right lateral bending and left/right axial rotation were established. Data were analyzed using mixed‐effects maximum likelihood regression models. Results: Total flexion/extension did not change across the 4 levels. There was no difference between flexion and extension, and no CM was identified. Lateral bending was not different across levels, but tended to be greater in the cranial spine. Axial rotation was ～2.6 times greater in the caudal segments. Lateral bending and axial rotation were coupled. Conclusions: Kinematics of the cranial and caudal cervical spine differed markedly with greater mobility in the caudal cervical spine.     

流式细胞技术在哺乳动物性别控制中的研究进展

在动物性别控制的各种技术中,应用流式细胞仪分离的X、Y精子授精可较准确地控制动物后代的性别.文章简单阐述了流式细胞仪分离X、Y精子的原理及其在动物性别控制中的应用,并结合最新研究进展详细地探讨了这项技术在实际生产与应用中的注意事项,旨在为这项技术的研究与应用提供必要的参考与建议.     

琉式细胞技术在哺乳动物性别控制中的研究进展

在动物性别控制的各种技术中,应用流式细胞仪分离的X、Y精子授精可较准确地控制动物后代的性别.文章简单阐述了流式细胞仪分离X、Y精子的原理及其在动物性别控制中的应用,并结合最新研究进展详细地探讨了这项技术在实际生产与应用中的注意事项,旨在为这项技术的研究与应用提供必要的参考与建议.     

应用流式细胞技术鉴定桑树倍性的样品选择

利用流式细胞仪对桑树不同叶位的叶片、同一叶位的不同部位、叶芽和种子胚根等进行倍性测定,筛选合适的桑树倍性鉴定材料。结果表明,未完全展开幼叶的检测效果优于第1叶位叶和第2叶位叶,第1叶位叶的检测效果优于第2叶位叶;同一叶位幼叶叶基部分的检测效果优于叶尾部分;叶芽的检测效果最好,而且制备样品的杂质较少,是用于流式细胞术研究的最佳材料。胚根也可以作为流式细胞仪检测的材料,这为更早检测桑树的倍性提供了候选材料。研究结果为选取不同时间段的不同材料进行桑树倍性检测提供了依据,也为其他植物倍性鉴定取材提供了参考。     

流式细胞仪在快速测定鲤春病毒血症病毒滴度中的应用

本研究建立了流式细胞仪快速检测鲤春病毒血症病毒（spring viraemia of carp virus,SVCV）滴度的方法。运用荧光激活细胞分选（fluorescence-activated cell sorting, FACS）技术检测SVCV A1株对草鱼性腺细胞系（GCO）的感染情况。用SVCV病毒单克隆抗体为一抗,FITC标记的羊抗鼠抗体为二抗,运用FACS来检测感染后不同时间点,以及不同病毒接种量的阳性细胞率。感染第3天时为最佳的病毒滴度测定时间点,测得SVCV的病毒滴度为8.31×105 FIU/mL,最低检测病毒滴度为（1000 FIU/mL）,与传统空斑试验（plaque assay,PA）相比,两种方法测得的结果基本一致。实验结果表明,FACS是一种简捷、高效、直接的检测SVCV滴度的方法,是一种新型的病毒滴度测定方法。     

流式细胞术在活化T淋巴细胞检测中的应用

免疫细胞的活化和增殖是免疫应答产生的一种标志.静止T淋巴细胞经过抗原刺激活化后,表现出一系列的特征,如细胞膜表面活化分子的表达、细胞分裂和产生细胞因子等.近年来,已有利用流式细胞术(FCM)通过检测免疫细胞活化后的特性,来分析抗原特异性免疫应答及其应答规律的报道.论文分别从几个不同的角度综述了FCM在活化T淋巴细胞检测中的应用.     

流式细胞术检测不同试验条件对猪T 淋巴细胞亚群的影响

通过不同方式处理猪外周血淋巴细胞,对不同保存时间的淋巴细胞进行流式细胞术检测,探讨其对T淋巴细胞亚群中CD3^＋、CD4^＋、CD8^＋亚群的影响。采用肝素钠、EDTA、枸橼酸钠3种抗凝剂抗凝猪外周静脉血,分别在第1、2、4、6、8、10天用流式细胞仪检测。结果表明,肝素钠抗凝,第1、2、4天内检测差异不显著(P＞0.05),第6天检测CD4^＋、CD8^＋细胞显著减少(P＜0.05),第8、10天极显著减少(P＜0.01)。EDTA抗凝,第1、2、4天内检测差异不显著(P＞0.05),第6天检测CD3^＋细胞显著减少(P＜0.05),第8、10天极显著减少(P＜0.01);枸橼酸钠抗凝,第1、2、4、6、8天内检测差异不显著(P＞0.05),第10天检测CD3^＋、CD4^＋细胞极显著减少(P＜0.01),CD8^＋细胞显著减少(P＜0.05)。3种抗凝剂检测比较,处理后的第1、2、4、6天肝素钠抗凝管检测结果CD3^＋、CD4^＋、CD8^＋均高于EDTA和枸橼酸钠抗凝。本研究表明,流式细胞术检测猪外周血优选肝素钠抗凝,样本采集后4 d内检测完毕。     

Effects of Seminal Fluid Fractions on Plasma and Acrosome Membrane Integrity and Mitochondrial Membrane Potential Determined by Flow Cytometry in Chilled Canine Spermatozoa

Depending on the mammal species, the use of seminal plasma during semen processing for cryopreservation has been found to have both beneficial and detrimental effects. This study was designed to determine the effects of the second (SF) and third [prostatic fluid, (PF)] ejaculate fractions on plasma membrane and acrosome integrity, mitochondrial membrane potential (ΔΨm), phosphatidylserine (PS) translocation and sperm motility in chilled canine spermatozoa by flow cytometry. After pooling the second sperm‐rich fraction of ejaculates from six dogs, samples for each assay were preserved at 5°C for 72 h in egg yolk‐TRIS extender (EYT) alone (control) or supplemented with seminal fluid from the second (EYT‐SF) or third (EYT‐PF) ejaculated fractions. After cold storage, groups EYT‐SF and EYT‐PF showed significantly higher percentages of sperm cells with an intact acrosome [68.8 ± 1.4%, 69.6 ± 2.6% (p < 0.01)] and intact plasma membrane [48.1 ± 2.8%, 50.4 ± 8.2% (p < 0.001)] than that observed in EYT [51.7 ± 3.2% and 33.3 ± 4.1% respectively]. Only in EYT‐SF was PS translocation significantly reduced compared to EYT‐PF and EYT [3.9 ± 0.4%, 10.2 ± 2.2% and 9.0 ± 1.5%, respectively (p < 0.001)]. However, significantly diminished sperm motility was observed in EYT‐SF and EYT‐PF compared to EYT [36.8 ± 2.1%, 35.5 ± 2.3% and 78.4 ± 4.7% (p < 0.001)]. No significant differences were detected in ΔΨm (p > 0.05). In conclusion, supplementing semen extenders with seminal fluid from the second or third fractions of the ejaculate supplementation helps to preserve the integrity of the plasma and acrosome membranes along with the mitochondrial membrane potential but seems to compromise the motility of canine spermatozoa chilled for 72 h.     

藏獒细小病毒病的诊断

藏獒作为犬类的一种,也受到犬类病毒感染的威胁。据调查,侵害藏獒的主要病毒有犬细小病病毒和犬瘟热病毒两大类。作者对青海某藏獒养殖户的病死藏獒进行临床症状观察,病理剖检及实验室诊断,通过病毒的分离培养及鉴定,确诊病死藏獒为犬细小病毒感染。鉴于养殖场中该病的存在及对养犬业的危害,建议加强对犬细小病毒的诊断及监控。     

一例犬会阴疝的诊治报告

会阴疝（perineocele）是盆隔肌组织缺陷，不能支撑直肠，使盆腔及腹腔内容物经盆膈和直肠间脱至会阴部皮下。此病在临床上发病率低，死亡率小，但病程长，诊断和治疗困难。就一例会阴疝患犬的诊断和治疗进行分析和总结，尤其对手术治疗展开深入探讨，目的是为该病的临床诊疗提供一定的理论和实践指导意义。     

犬血丝虫病诊疗病例

犬血丝虫病是由犬血丝虫寄生于犬类动物右心室或肺动脉引起的犬类动物的一种重要的寄生虫病,以循环障碍、呼吸困难、贫血为主要特征。犬血丝虫又叫犬心丝虫或犬恶丝虫,犬为其终末宿主,蚊、蚤等吸血昆虫是中间宿主。当寄生的虫体影响心脏器质功能（比如三尖瓣或肺动脉瓣功能不全）时,患犬会出现呼吸困难、胸腹水、下体浮肿等较为严重的临床症状,甚至发生急性死亡。诊断本病以外周血中查到犬血丝虫的幼虫微丝蚴为主。治疗本病则以驱虫为主,补血、输液等对症治疗作为辅助措施。     

The Utility of PCR for Antigen Receptor Rearrangement (PARR) in Staging and Predicting Prognosis in Canine Lymphoma

Introduction:  In lymphoid neoplasia, molecular assays to confirm clonality rely on the fact that lymphoid cells normally contain DNA regions that are unique in sequence, resulting from recombination of the V, D, and J genes. Based on the belief that malignancies are clonal, the presence of a single, clonal rearrangement can indicate neoplasia. In humans with lymphoid malignancies, PCR for antigen receptor rearrangement has been studied as a molecular staging test and a therapeutic monitoring tool. The purpose of this study was to determine the utility of PARR for molecular staging and predicting prognosis in canine lymphoma.
Methods:  Patients with lymphoma who had complete clinical staging were included in the study. All had PARR of pre treatment lymph node, blood and/or bone marrow. All were assigned a clinical stage (WHO 1–5) and a PCR stage (PS) where 0 indicated no clonal expansion in the blood, and 5 indicated the presence of a clonal expansion. In addition to stage, other factors that could affect prognosis were evaluated.
Results:  Eighty six patients were included. Clinical stage was distributed as follows: 3 = 44, 4 = 21, and 5 = 21. Sixty two patients had clonal rearrangements detected in the lymph node and were able to be PCR staged. Seventeen were PS 0 and 45 were PS 5. PCR stage was not prognostic for DFI or survival.
Conclusion:  PARR is sensitive at detecting blood/bone marrow infiltration in canine lymphoma. It is not a useful prognostic tool in the diverse population in this study.     

犬瘟热的诊断及其预防免疫的研究进展

本文对犬瘟热（CD）的诊断、预防免疫和免疫失败的影响因素及犬瘟热病毒（CDV）的宿主范围进行了综述。CDV不仅感染陆生食肉动物，而且也感染水生食肉动物，并且其宿主范围还在不断扩大。CDV感染主要采用病毒分离、特异性病毒抗原或特异性核酸检测等方法确诊。疫苗包括灭活的CDV疫苗、麻疹病毒（MV）异源苗及CDV弱毒活苗。疫苗接种犬的免疫反应主要取决于毒株特性及犬的应答能力，只有弱毒活苗能诱导产生持久而坚强的保护力。尽管多年来CDV弱毒活苗的使用控制了CD的发生，但最近免疫过的犬发生CD的病例并不少见。分析免疫失败的原因，主要是母源抗体干扰、疫苗质量差、其它病毒的免疫抑制以及CDV流行株可能发生了变异等因素的影响。     

猪精子质量的流式细胞检测分析

本试验将流式细胞术应用于猪精子分析,同时检测精子功能的多个特性,以保证精子具备多种特性且功能完整,顺利完成授精过程。分别采用SYBR-14/PI（Propidium Iodide）、YO-PRO-1/PI、PNA/PI及Mito Tracker/YO-PRO-1 4种染色方法对荣昌猪精子的质膜完整性、细胞凋亡、顶体完整性及线粒体功能进行了检测和分析;以SYBR-14⁺/PI^-表示质膜完整的活精子,YO-PRO-1^-/PI^-表示活精子,PNA^-/PI^-表示未发生顶体反应的活精子,Mito Tracker⁺/YO-PRO-1^-表示线粒体活性高的活精子,以上述检测指标的综合结果判定精子功能的完整性。本试验通过流式细胞术对荣昌猪精子进行质量检测和分析,结果表明,该方法可为临床诊断种猪不孕症提供理论依据,对有效治疗不孕症有重要意义。     

Detection and Analysis of the Pig Sperm Quality by Flow Cytometry

The flow cytometry was used in this trail to determine the pig sperm quality,in order to ensure the sperm had complete functions during the fertilization process.Four stain methods of SYBR-14/PI (Propidium Iodide),YO-PRO-1/PI,PNA/PI,Mito tracker/YO-PRO-1 were used to detect the plasma membrane integrity,apoptosis,top body ntegrity and mitochondrial function of Rongchang pig sperm.SYBR-14⁺/PI^- expressed live sperm with complete plasma membrane,YO-PRO-1^-/PI^- expressed live sperm,PNA^-/PI^- expressed live sperm had no top body reaction,and Mito Tracker⁺/YO-PRO-1^-expressed live sperm with high mitochondrial activity.The functional integrity of the tested sperm was evaluated according to the above comprehensive results.Through the study of Rongchang pig sperm quality detection by flow cytometry,it suggested that the method could provide theoretical basis for clinical diagnosis,and was very important for swine infertility treatments.     

犬瘟热实验室诊断方法研究进展

犬瘟热病毒(CDV)为副粘病毒科,麻疹病毒属成员,由carré于1905年首次分离发现。随着近年来研究的不断深入,发现犬瘟热病毒的感染范围在不断的扩大。犬瘟热病毒不仅可以感染犬科(如犬、狐、狼等)、鼬科、浣熊科、猫科(如狮、虎、豹)等多种肉食动物[1-6],在实验室条件下还可以感染灵长类,引起脑脊髓炎,以及感染人的神经细胞[7,8]。目前已有日本猕猴和野猪自然感染CDV引起的致死性脑炎的临床病例报道[9-10]。Hoyland J A等[11]通过原位RT-PCR技术证实了患Pa-grts骨病的病人组织中含有犬瘟热病毒核酸,但犬瘟热是否会成为继狂犬病之后通过…