清瘟败毒散的质量标准提升研究

为完善清瘟败毒散的质量标准,采用薄层色谱鉴别清瘟败毒散中的黄连、地黄、栀子、连翘、知母、淡竹叶;采用HPLC法测定清瘟败毒散中的栀子苷含量,色谱条件为：ODS C18色谱柱（4.6 mm ID×250 mm,5 μm）,乙腈-水（13：87）为流动相,检测波长为238 nm。研究结果表明,在建立的薄层色谱鉴别结果中,在与对照药材色谱相对应的位置上,供试品色谱显示相同颜色的斑点,且阴性样品无干扰;栀子苷在0.062~0.370 μg范围内具有良好的线性关系,其回归方程为y=1 789.1x-14.296,R²=0.99984,平均回收率为99.60%,RSD为0.76%。本方法简单、准确、重复性好,能更好地控制清瘟败毒散的质量,可作为清瘟败毒散的质量标准。     

伊维菌素微乳中伊维菌素的含量测定方法研究

为建立伊维菌素微乳中伊维菌素含量的高效液相色谱（HPLC）测定方法,选用Hypersil ODS2 （5 μm,4.6 mm×250 mm）色谱柱,流动相为甲醇∶乙腈∶水为35∶60∶5（V/V/V）,检测波长为244 nm,柱温为30 ℃,流速为1 mL/min进行测定。结果显示,伊维菌素在该色谱条件下,系统适应性良好,在80~320 μg/mL浓度范围内线性关系良好,回归方程为：Y=22 700X+2 510,R²=0.9998,总平均回收率为101.90%±2.94%,RSD为2.88%,对中试生产的3批伊维菌素微乳进行含量测定,RSD为1.86%。表明该含量测定方法准确可靠,重现性好,可用于伊维菌素微乳中伊维菌素含量的测定,并为该新型制剂的质量标准的制定和质量评价提供依据,也为后期的临床安全应用提供可靠的参考。     

Content Determination of Ivermectin in the Ivermectin Microemulsion Injection

In order to establish the determination method of ivermectin (IVM) in the ivermectin microemulsion injection by high performance liquid chromatography (HPLC), Hypersil ODS2 colunm (5 μm,4.6 mm×250 mm) was used in this study. The mobile phase composed of methanol, acetonitrile and water (35:60:5,V/V/V) at a flow rate of 1 mL/min. The detection wave length was set at 244 nm and the column temperature was 30 ℃. The results showed that the HPLC system suitability of IVM was good. A good linear correlation of IVM was observed within the concentration range 80 to 320 μg/mL, and the average recovery rate was 101.90%±2.94% with RSD was 2.88%, the regression equation was Y=22 700X+2 510 (R²=0.9998). The RSD of IVM content in ivermectin microemulsion injection was 1.86%. The method was accurate and reliable,reproducible,easy to operate, which could be used in new type of ivermectin microemulsion injection. The method could be used as the basis of quality control and establishing a quality standard, and to provide basis for quality evaluation, also can provide reliable reference for safe veterinary clinical application in the future.     

基于实时荧光定量PCR技术的猪伪狂犬病活疫苗含毒量评价

为定量分析贵州省临床用猪伪狂犬病(pseudorabies,PR)活疫苗中病毒含量,根据GenBank中公布的伪狂犬病病毒(pseudorabies virus,PRV)gB基因序列(登录号:M17321.1)设计1对特异性引物,建立实时荧光定量PCR方法,并对来自6个不同厂家的猪伪狂犬病活疫苗进行含毒量分析。结果显示,试验成功建立了可用于PRV检测的实时荧光定量PCR方法,标准曲线中标准品各稀释度质粒拷贝数与Ct值呈良好的线性关系,标准曲线方程为:Y=-0.97X+33.66(R^2=0.999),该方法最低检测病毒含量为3.19×10~1拷贝/μL,敏感性高且具有良好的重复性和特异性。应用所建立的实时荧光定量PCR方法对贵州省临床用6个厂家生产的猪伪狂犬病活疫苗的病毒含量进行检测,结果显示,6种市售猪伪狂犬病活疫苗(A～F)中病毒含量分别为1.67×10~7、4.83×10~5、2.64×10~6、4.27×10~7、3.39×10~6和3.68×10~5拷贝/μL,来自不同厂家的疫苗病毒含毒量存在明显差异,最大差异可达116倍,其中来自A、D厂家的两种猪伪狂犬病活疫苗具有较高的含毒量。本试验所建立的实时荧光定量PCR方法可用于猪伪狂犬病活疫苗病毒含量的快速检测和评价,对猪伪狂犬病活疫苗的质控和临床用疫苗选择具有一定的指导意义。     

Determination of Acetaminophen in Bactrian Camel Plasma by RP-HPLC

To establish the reversed phase high-performance liquid chromatography (RP-HPLC) method for determination of plasma concentration of acetaminophen in Bactrian camel, the chromatography was carried on a C18 column (150 mm×4.6 mm,5 μm).The mobile phase was composed of methanol-water (20:80),the flow rate was 0.5 mL/min,and the detection wavelength was set at 248 nm for acetaminophen.The retention time of acetaminophen was 6.5 min, and was not disturbed by other miscellaneous peaks.The calibration curve was linearity in the range of 0.08 to 10.24 μg/mL (R²=0.9997),and the quantitation limit of acetaminophen was 0.10 μg/mL.Both the inter-day and intra-day RSD were under 7.0%, the absolute recovery of the method was 80.39% to 93.56%,the relative recovery of the method was 94.66% to 104.73%.Therefore, the established method was simple, reproducible and accurate,which could be used for determination of plasma acetaminophen concentration and in vivo activities of CYP1A enzyme in Bactrian camel.     

反向高效液相色谱法检测双峰驼血浆中对乙酰氨基酚的含量

试验旨在建立测定双峰驼血浆中对乙酰氨基酚浓度的反相高效液相色谱法（reversed phase high-performance liquid chromatography,RP-HPLC）。选用Supelco Discovery C18色谱柱（150 mm×4.6 mm,5 μm）,以甲醇：水=20：80为流动相,流速为0.5 mL/min,紫外检测波长为248 nm,采用甲醇直接沉淀蛋白方法处理血浆样品。结果表明,对乙酰氨基酚的保留时间为6.5 min,分离度良好,无血浆杂峰干扰,且在0.08~10.24 μg/mL浓度范围内线性关系良好（R²=0.9997,n=8）,检测限为0.10 μg/mL,日内和日间精密度RSD均小于7.0%,绝对回收率为80.39%~93.56%,相对回收率为94.66%~104.73%。因此,本试验所建立的RP-HPLC法操作简便、准确,重复性好,专属性强,适用于检测双峰驼血浆中对乙酰氨基酚浓度,为后续的借助特异性探针药物--对乙酰氨基酚研究双峰驼CYP1A酶的体内活性提供了可靠而准确的定量方法。     

注射液中非法添加地塞米松磷酸钠UPLC-PDA检查方法的建立

本试验建立了鱼腥草、柴胡、维生素C、盐酸林可霉素、黄芪多糖等注射液中非法添加地塞米松磷酸钠的超高效液相色谱检查方法。采用Waters ACQUITY UPLC^® BEH C18色谱柱（2.1 mm×50 mm,1.7 μm）,以0.75%三乙胺溶液（磷酸调节pH 3.0）-甲醇-乙腈（50：45：5）为流动相,流速0.3 mL/min,二极管阵列检测器,波长采集范围为190~400 nm,记录色谱图波长242 nm,柱温25℃,进样量10 μL。地塞米松磷酸钠质量浓度在10.0~200.0 mg/L内峰面积呈良好线性关系（R²=0.9999）,在鱼腥草、柴胡、维生素C、盐酸林可霉素、黄芪多糖等注射液中地塞米松磷酸钠的平均回收率和相对标准偏差（RSD）分别为97.54%和1.14%、101.59%和0.19%、100.92%和0.92%、99.82%和0.73%、100.08%和0.62%。本方法简单、快速、灵敏、可靠,适用于鱼腥草等5种注射液中非法添加地塞米松磷钠的检查。     

Establishment of Detection Methods for Dexamethasone Sodium Phosphate in Injections by UPLC-PDA

A detection method of dexamethasone sodium phosphate in houttuynia injection,bupleurum injection,vitamin C injection,lincomycin hydrochloride injection,and asragulus polysacharin injection was established by UPLC-PDA.The Waters ACQUITY UPLC^® BEH C18 column(2.1 mm×50 mm,1.7 μm) was used,the mobile phase consisted of 0.75% triethylamine phosphatic liquor/solution (pH 3.0)-methyl alcohol-acetonitrile(50:45:5),the flow rate was 0.3 mL/min,the column temperature had remained 25℃,photodiode array detector wavelength range was 190 to 400 nm,the recorded wavelength was 242 nm,the injection volume was 10 μL.The linear range of dexamethasone sodium phosphatel was 10.0 to 200.0 mg/L (R²=0.9999),the mean recovery and RSD for dexamethasone sodium phosphate in houttuynia injection,bupleurum injection,vitamin C injection,lincomycin hydrochloride injection and asragulus polysacharin injection were 97.54% and 1.14%,101.59% and 0.19%,100.92% and 0.92%,99.82% and 0.73%,100.08% and 0.62%,respectively.This method was accurate,easy and fast,it was suitable for detection of dexamethasone sodium phosphatein houttuynia injection and other four kinds of injections.     

基施硒肥对不同生育期紫花苜蓿吸收、转化及利用硒的影响

采用土壤施硒方法,研究基施硒肥对不同生育期紫花苜蓿(Medicago saliva)吸收、转化和利用硒的影响.结果表明:基施硒肥能显著提升紫花苜蓿吸收外源硒的能力(P<0.05);并随生育期延伸呈先升后降的倒"V"型变化,初花期吸收硒能力最强;而紫花苜蓿硒含量随生育期的推移而降低.基施硒肥显著提高紫花苜蓿有机硒的转化率(P<0.05),当施硒量≥0.45 mg·kg^-1时,有机硒转化率 > 50%.紫花苜蓿有机硒转化率随生育期的发展呈下降趋势;紫花苜蓿对硒肥的利用率很低,不超过1.5%;并随生育期的进程先升后降,初花期硒肥利用率最高.施硒量与不同生育期苜蓿硒含量之间存在极显著的线性关系(P<0.01),基施硒肥与苗期牧草硒线性方程为:y=1.9912x+0.1827(R²=0.9696),基施硒肥与分枝期牧草硒线性方程为:y=1.7394x+0.1724(R²=0.9670),基施硒肥与孕蕾期牧草硒线性方程为:y=1.5045x+0.1542(R²=0.9694),基施硒肥与初花期牧草硒线性方程为:y=1.2547x+0.1588(R²=0.9835),基施硒肥与盛花期牧草硒线性方程为:y=1.0044x+0.1500(R²=0.9904).     

3种染料在小鼠胃肠推进试验中的运用

为了筛选胃排空和肠推进试验最佳示踪染料,本试验选取葡聚糖蓝2000、伊文思蓝和墨汁3种染料,用微量倍比稀释法测定一系列浓度各染料在一系列波长的吸光度值,以确定各染料的最大吸收波长和可测浓度范围。根据各染料可测范围确定小鼠最佳灌胃剂量,测定各染料在胃残留率、小肠推进率及小肠均4段各段染料残留率,确定各染料在消化道总残留率。结果显示,在本试验测定波长中,葡聚糖蓝2000、伊文思蓝和墨汁最大吸收波长分别630、630和405 nm;可测浓度范围分别为0.16~5.00 mg/mL（R² =0.9863）、0.78~6.25 μg/mL（R²=0.9984）和0.02%~0.16%（R²=0.9979）。3种染料对胃排空率的影响差异不显著（P > 0.05）,而肠推进率伊文思蓝显著快于其他两种染料（P < 0.05）;3种染料总残留率葡聚糖蓝2000最高,其次为墨汁,伊文思蓝最低,分别为93.6%±4.5%、71.5%±8.5%和18.7%±2.8%。本试验结果提示,在检测受试物对在体消化道运动影响研究中,如果仅要求测定肠推进率可用墨汁,如果要精确检测胃和小肠各段排空情况则用葡聚糖蓝2000为宜。     

奶牛用利福昔明子宫注入剂有关物质反相高效液相色谱检测方法的建立及应用

本试验旨在建立利福昔明有关物质的检测方法。采用反相高效液相色谱法,紫外检测器进行检测,采用WondaSil C18-WR 250 mm×4.6 mm 5 μm色谱柱,柱温:40 ℃,流动相及其比例为:V（甲醇） ∶V（乙腈）∶V（3.16 g/L甲酸铵,浓氨水调pH至7.2）=31.5∶31.5∶37,检测波长276 nm。采用面积归一化法计算利福昔明子宫注入剂的检测限、定量限及其浓度;考察了方法精密度、线性相关性及有关物质专属性,并通过极端条件的破坏、有关物质的含量、分离度等对主成分峰面积降解程度进行了比较。利福昔明子宫注入剂主成分峰在浓度为40~150 μg/mL的范围内,峰面积与浓度呈现良好的线性关系（R²=0.9998）,其有关物质浓度对峰面积线性回归方程为y=23 510x+7 182,其进样精密度RSD为0.13%,方法精密度RSD为0.46%,制剂一定时间内经过强酸、强碱、氧化、高温条件破坏后均有不同程度降解,以破坏前的样品主成分含量为100.00%,氧化破坏和高温破坏主成分降解达13%,酸、碱破坏主成分降解在18%左右,主成分峰保留时间在以上4种条件破坏前后一致,且主成分峰与杂质峰之间及杂质峰与杂质峰之间分离度良好（主成分峰与杂质峰之间分离度均大于1.5,杂质峰与杂质峰之间分离度均大于1.2）。本试验建立的利福昔明子宫注入剂有关物质检测方法操作简便,专属性和重复性好,可适用于利福昔明子宫注入剂有关物质的测定。     

Establishment and Application of Reversed-phase High Performance Liquid Chromatography Method for Detection of Related Substances of Rifaximin Uterine Infusion in Dairy Cows

The study was aimed to establish a method for detection of rifaximin related substances by reversed-phase high-performance liquid chromatography with UV detector. The instrument methods was WondaSil C18-WR 250 mm×4.6 mm 5 μm column, mobile phase was methanol∶acetonitrile∶3.16 g/L ammonium formate (concentrated aqueous ammonia adjusted to pH 7.2)=31.5∶31.5∶37, with detection wavelength 276 nm. Area normalization method was used to calculate the limit of detection, limit of quantification and concentration of rifaximin uterine injectant; The precision of the method, linear correlation and related substances specificity were explored. And the degradation degree of the peak area of the principal components was compared by the destruction of the extreme conditions, the content of the related substances and the separation degree. 40 to 150 μg/mL concentration of rifaximin, peak area and concentration showed a good linear relationship (R²=0.9998), the related substances concentration on peak area linear regression equation was y=23 510x+7 182, within a certain period of time after preparation of acids, alkalis, oxidation, high temperature degradation damage in varying degrees, in the main ingredient content in the sample before the damage was 100.00%, after oxidative damage and heat damage the main component was 13%, acids, alkalis destruction of the main component degradation was about 18%, the main component peak retention time was consistency before and after the destruction. The retention time and resolution of rifaximin and related-substance were good. (The separation between the main component peak and the impurity peaks were greater than 1.5, the degree of separation between the impurity peak and impurity peaks greater than 1.2). The substance detection methods of rifaximin uterine injectants was simple and had specificity and good repeatability, selective and sensitive, which could be used to analyze the related substance of rifaximin uterine injectants.     

实时荧光定量RT-PCR检测小反刍兽疫病毒方法的建立

本试验旨在建立一种快速、灵敏的诊断小反刍兽疫的方法。本研究通过RT-PCR方法扩增小反刍兽疫病毒N基因,连接到pMD19-T克隆载体上,构建质粒标准品。根据GenBank中中国流行毒株及Nigeria 75/1疫苗株N基因保守序列设计引物,利用SYBR Green Ⅰ法进行实时荧光定量PCR,建立标准曲线,并进行特异性试验、敏感性试验和重复性试验。结果表明,在2.82×10⁰~2.82×10⁷拷贝/μL范围内,Ct值与质粒拷贝数对数值呈良好的线性关系,标准曲线线性关系R²值为0.992;其他病毒无特异性扩增曲线,特异性良好;批内变异系数为0.27%~2.77%,批间变异系数为0.41%~3.39%,重复性较好;检测灵敏度可达2.82拷贝/μL,是普通PCR的1 000倍。用该方法对12份cDNA样品进行检测,9份为阳性,3份为阴性,而普通PCR检测,7份为阳性,5份为阴性,说明本方法比普通PCR灵敏度高。本检测方法的建立对快速、灵敏诊断小反刍兽疫,防止疫情的扩散具有重要意义。     

猪瘟病毒TaqMan实时荧光定量PCR检测方法的建立及初步应用

根据GenBank中猪瘟病毒（classical swine fever virus,CSFV）5′NTR的保守序列,设计1对特异性引物和TaqMan探针,以CSFV全长基因重组质粒pGEM-CSFV为标准品,建立TaqMan实时荧光定量PCR标准曲线,进行特异性、敏感性和重复性试验,并检测人工感染CSFV后不同时期猪血浆中CSFV的载量。结果表明,标准曲线循环阈值与模板浓度具有良好的线性关系,R²=0.999;敏感性高,最低检测限为1×10¹拷贝/μL;特异性强,与猪圆环病毒2型、猪细小病毒、猪伪狂犬病病毒和猪繁殖与呼吸综合征病毒无交叉反应性;重复性好,组内和组间变异系数均小于2%。人工感染CSFV后第7天猪血浆中CSFV载量达到峰值,之后逐渐降低。结果表明,建立的TaqMan实时荧光定量PCR方法具有特异、敏感、重复性好等优点,为CSFV的定量分析及临床诊断奠定基础。     

Application of Three Kinds of Dyes in Gastrointestinal Propulsion in Mice

In order to select the best tracer dye for gastric emptying and intestinal propulsion experiment,three kinds of dyes including of dextran blue 2000,direct blue 53 and ink were selected.The absorbance values of the series of concentration of dyes were tested in a range of wavelength by micro double dilution method.Moreover,the maximum absorption wavelength and measurable concentration range of three dyes were determined.According to measurable range of three kinds of dyes,the optimal gavage doses were obtained in mice.The gastric residual rates,small intestinal propulsion rates and residual rates in four parts of equal small intestine were tested.The results showed that the maximum absorption wavelengths of dextran blue 2000,direct blue 53 and ink were 630,630 and 405 nm,respectively.Their measurable concentration ranges were 0.16 to 5.00 mg/mL (R²=0.9863),0.78 to 6.25 μg/mL (R²=0.9984) and 0.02% to 0.16% (R²=0.9979).The difference of gastric emptying rates among the three dyes was not significant (P > 0.05),and intestinal propulsion rate of direct blue was significantly higher than those of the other two dyes (P < 0.05).Total residual rates of dextran blue 2000,ink and direct blue were 93.6%±4.5%,71.5%±8.5% and 18.7%±2.8%,respectively.The results suggested that ink was the best tracer dye if only the intestinal propulsion rate in vivo needed to be determined,while dextran blue 2000 is appropriate if a precise determination of the gastric and intestinal emptying needed to be done.     

APSIM苜蓿模型在宁夏半干旱地区的适应性

基于宁夏3个试验点的数据和同期气象资料,对APSIM模型在宁夏半干旱地区的适应性进行研究。根据试验条件建立相应模型,通过“试错法”对模型相关参数进行校准,实现参数的本地化;并运用多个检验统计指标验证紫花苜蓿（Medicago sativa）土壤含水量及其分别在同心和银川试验点的生育期和干草产量。结果表明：各生育期模拟值和实测值显著相关,决定系数R²分别为0.99和1.00,D值均为0.99,NRMSE分别为2.4%和7.8%;干草产量表现出良好的相关性和一致性,决定系数R²分别为0.82和0.98,D值均为0.99,NRMSE分别为7.1%和9.3%。模拟值比实测值平均高估了7.1%;土壤含水量整体表现出较好的相关性和一致性,决定系数R²为0.82,D为0.85,NRMSE为10%,模拟值比实测值平均高估了10.1%。产量的高估很有可能与水分的高估相关,需要更详细的测量田间试验数据,改善模型土壤参数描述,提高模型预测准确度。校准后的APSIM模型在宁夏半干旱地区具有较好的适应性,可以用于指导该区半干旱区苜蓿生产及栽培措施优化管理。     

A SYBR Green Ⅰ Based on Real-time Quantitative RT-PCR Assay for Specific Detection of Peste des Petits Ruminants Virus

The study was aimed to establish a rapid and sensitive diagnostic method for the prevention and control of peste des petits ruminants.In this study,a fragment of PPRV N gene was amplified and cloned into pMD19-T cloning vector.Real-time quantitative PCR assay was performed using SYBR premix Ex Taq.The standard curve was plotted and the specificity,sensitivity and reproducibility of the assay were assessed.The generated standard showed linearity over the entire range from 2.82×10⁰ to 2.82×10⁷ copies/μL with a linear correlation（R²）of 0.992.The specificity of the assay showed that other viruses failed to show an amplification signal.The coefficient of variation（CV）values for intra- and inter-assay variability were low,ranging from 0.27%~2.77% and 0.41%~3.39%,respectively.The lower detection limit,based on plasmid copy number,achieved was 2.82 copies/μL and was 1 000 times more sensitive than conventional PCR assay.cDNA of 12 samples were tested using this method,9 were positive,and 3 were negative.The samples were also tested using conventional PCR,7 were positive and 5 were negative,proving that the two-step SYBR Green Ⅰ based Real-time quantitative RT-PCR assay reported here were more sensitive than conventional PCR.The establishment of this detection method is of great significance to rapid and sensitive diagnosis of peste des petits ruminants and preventing the spread of peste des petits ruminants.     

治疗乳房炎的中药芩红灌注剂质量标准的建立

芩红灌注剂是一种新研制的用于治疗奶牛乳房炎的中药复方制剂，为了确保芩红灌注剂临床用药的安全性和生产工艺的稳定性，研究建立了其质量标准；采用薄层色谱法（TLC）对芩红灌注剂中黄芩、金银花和红花进行定性鉴别，参照《中国兽药典》中灌注剂的检查方法，对芩红灌注剂的相对密度、pH、无菌、有关物质和装量等进行检查，采用高效液相色谱法（HPLC）对黄芩苷进行定量检测。试验结果表明，芩红灌注液样品与对照药材及对照品在相应位置上呈现相同颜色的斑点，表明用于鉴别黄芩、金银花和红花的TCL法专属性、重现性和耐用性均良好，芩红灌注剂的相对密度、pH、无菌、有关物质和装量均符合相关规定，并确立芩红灌注剂的相对密度为不低于1.00（30 ℃），pH为5.0~7.0，测定芩红灌注剂中黄芩苷含量的HPLC法精密度、稳定性、重复性、线性、专属性和耐用性均较好，含量测定中黄芩苷在0~9.20 μg范围内与峰面积线性关系良好（R²=0.9999），平均回收率为100.04%（RSD=1.34%，n=9）。所建立的质量标准准确可行，重复性好，可用于芩红灌注剂的质量控制，为其大规模的生产和临床应用奠定基础。     

利用体外模拟瘤胃发酵技术预测奶牛生产性能的研究

本试验旨在利用全混合日粮（TMR）体外发酵参数预测奶牛的生产性能,采集2头体重550 kg、安装永久瘤胃瘘管的健康荷斯坦奶牛瘤胃液,并取40种不同的TMR进行体外发酵试验,测定体外发酵24 h的乙酸、丙酸、丁酸、氨态氮（NH₃-N）含量,以及甲烷（CH₄）排放量、微生物蛋白质（MCP）、体外干物质消失率（IVDMD）、体外蛋白质消失率（IVCPD）和体外有机物消失率（IVOMD）等指标。记录饲喂不同TMR的奶牛对应的生产性能（产奶量、乳蛋白率、乳脂率）,并与奶牛生产性能之间建立预测模型。结果显示：①产奶量与丙酸含量（r=0.37,P<0.05）、MCP（r=0.40,P<0.05）均呈显著正相关,但与IVCPD（r=-0.44,P<0.01）呈极显著负相关;乳脂率与乙酸含量（r=0.55,P<0.01）、甲烷排放量（r=0.36,P<0.05）、MCP（r=0.40,P<0.05）呈极显著或显著正相关;乳蛋白率与MCP（r=0.91,P<0.01）、IVDMD（r=0.44,P<0.01）、IVCPD（r=0.45,P<0.01）呈极显著正相关,但是与IVOMD（r=-0.56,P<0.01）呈极显著负相关。②奶牛的生产性能可用体外发酵参数作为预测因子进行预测。产奶量（kg/d）=29.72-0.86IVCPD（R²=0.73,RSD²=47.26,P<0.001）;乳脂率（%）=0.40+0.06AA（R²=0.91,RSD²=0.15,P<0.001）;乳蛋白率（%）=1.52+0.06MCP（R²=0.78,RSD²=0.01,P<0.001）。奶牛的生产性能预测值与实际值的误差在允许的范围内,表明预测方程是可行的。     

牛血清淀粉样蛋白A时间分辨荧光免疫层析检测试剂盒研制

本研究旨在研制一种牛血清淀粉样蛋白A（SAA）时间分辨荧光免疫层析定量检测试剂盒,用于牛奶中SAA含量的临床快速检测。采用双抗体夹心法结合荧光免疫层析技术,在结合垫上固定荧光微球标记的抗SAA单克隆抗体及荧光微球标记的鸡IgY的混合物,在硝酸纤维素膜的检测区包被另一株抗SAA单克隆抗体,在硝酸纤维素膜的质控区包被山羊抗鸡IgY。经抗体原料筛选及荧光微球标记抗体的工艺优化后,绘制标准曲线并对试剂盒的空白限、精密度、稳定性及样本测试性能进行初步评估。结果显示,Medix SAA-2单克隆抗体包被与YBX SAA-3单克隆抗体标记为最适抗体配对原料。荧光微球标记抗体的工艺中,荧光微球与抗体的质量投料比为40∶1、偶联剂与荧光微球羧基摩尔比为2∶1的条件为最优组合。试剂盒标准曲线的四参数拟合曲线方程为y=（1.03947-0.00182）/[1＋（x/12.08222）×（-0.84692）]＋0.00182,线性相关系数R²=0.9997。研制的牛SAA检测试剂盒空白限为0.052 mg/L。精密度测试结果显示,批内变异系数 < 15%,批间变异系数 < 20%。室温稳定性试验表明,试剂盒在室温密封存放6个月的荧光T/C值相对跌幅约15%。自制试剂盒与上海蓝基试剂盒的样本对比测试相关系数R²为0.97。综上所述,本试验研制的试剂盒具有操作简便、灵敏度高、成本低廉等优点,能满足临床测定需求,可作为一种新型牛SAA检测的快捷、准确的检测手段。