钴(Ⅱ)配合物[Co(2,2′-bipy)2(NO3)]2SO4·(H2O)5的水热合成和晶体结构

以2,2′-联吡啶和硝酸根为混合配体,通过水热反应合成了一个新的钴(Ⅱ)配合物[Co(2,2′-bipy)2(NO3)]2SO4·(H2O)5,用元素分析、红外光谱、X-射线单晶衍射表征了配合物的结构.标题配合物晶体属单斜晶系,P2(1)/n空间群,晶体学参数:a=0.709 32(9) nm,b=1.952 8(2) nm,c=1.684 1(2) nm,β=96.835(2)°,V=2.316 1(5) nm3,Z=2,D=1.495 g/cm3.配合物中每个钴(Ⅱ)离子与两个2,2′-联吡啶分子中的4个N原子、1个硝酸根的两个O原子形成六配位的变形八面体结构.     

电化学发光探针与生物分子的定量标记与表征

电化学发光生物分析是在电极表面经过生物识别反应后检测电化学发光信号,实现对目标物的高灵敏检测,此类分析方法在疾病的早期诊断、筛选等方面具有广泛的应用前景。本研究中,选用牛血清蛋白(BSA)为模型蛋白,用电化学发光分子联吡啶钌衍生物(联吡啶-4'-甲基-4-羰基吡啶钌-N-琥珀酰亚胺酯双六氟磷酸酯,RuNHS)对BSA进行定量标记和表征,并考察了形成的复合物(BSARu)的电化学发光行为。结果表明:RuNHS和BSA的摩尔投料比为5:1、10:1时,标记率分别为1.1、2.1,BSA回收率分别为54%、61%,标记率随着投料比的增加而升高;同浓度下标记率越高电化学发光信号越强,且标记率为1.1的发光强度与BSA~(Ru)浓度在3.1×10~(-10)~3.1×10~(-6) g/mL范围内呈线性相关,为后续RuNHS标记抗体用于免疫分析奠定基础。     

2,2′-联吡啶合成工艺研究

以Ni-Fe/硅藻土为催化剂,固定床为反应器,吡啶直接偶联合成2,2′-联吡啶.系统考查了Ni含量、Fe含量、催化剂用量、异丙醇用量、反应温度、反应压力的影响,筛选出合适的反应条件:采用M-3催化剂,空速2 h-1,异丙醇用量2％,反应温度450℃,反应压力2 MPa,此条件下吡啶转化率达到65.7％,2,2′-联吡啶收率达到65％.该工艺操作简单,安全性高,产品收率高,适合工业化生产.     

2,2'-联吡啶的合成工艺研究

采用γ-Al₂O₃载体负载的Pd-Cu合金纳米催化剂,以高压釜为反应器,吡啶直接偶联合成2,2′-联吡啶。考查了催化剂中Pd-Cu摩尔比、催化剂用量、反应温度、反应时间的影响,筛选出反应的最优条件为Pd-Cu摩尔比1∶1.5,催化剂占原料2.5wt%,反应温度260℃,反应时间14 h,2,2′-联吡啶的收率可达到39%。该工艺操作简单,安全性高,适合工业化生产。     

4,4′-(4,4′-砜基二苯氧基)二苯甲酰氯(SODBC)的合成

以对甲基苯酚、4,4′-二氯二苯砜为原料,通过亲核取代反应合成了4,4′-二(4-甲基苯氧基)二苯砜,用高锰酸钾将甲基氧化得到4,4′-(4,4′-砜基二苯氧基)二苯甲酸(SODBA),后者在二氯亚砜和路易斯碱的催化下合成了4,4′-(4,4′-砜基二苯氧基)二苯甲酰氯(SODBC)白色固体.用FT-IR、1H-NMR、13C-NMR、DSC等对其进行了表征,实验证明该化合物具有预期的结构和较高的纯度.     

无溶剂合成2,2′-联吡啶桥联4,4′-联吡啶二元醇

将2-羟乙氧乙基对甲苯磺酸酯与4,4′-联吡啶按物质的量比1∶10混合,加热到熔化,保温10 min,形成N-羟乙氧乙基-4,4′-联吡啶,收率88%;然后与α,α-′(二溴甲基)-2,2′-联吡啶按物质的量比2∶1混合,加热到熔化,保温10 min,形成标题化合物,收率82%。     

4-(4-甲基苯基)-6-苯基-2,2′-二联吡啶芳炔铂(II)络合物的合成及光物理性质

设计合成以4-甲基苯基-6-苯基-2,2′-二联吡啶为主配体、芳炔为辅助配体的铂(II)络合物1-3.与苯乙炔、萘乙炔为辅助配体的络合物1,2相比,蒽乙炔4-(4-甲基苯基)-6-苯基-2,2′-二联吡啶铂(II)络合物MLCT激发态的能量升高.     

4-(反-4′-正丙基环己基) 环己基甲酸的合成

以w(NaOH)=10%的水溶液为反应介质,Ru/C为催化剂,4-(反-4′-正丙基环己基)苯甲酸(3HPA)进行加氢反应合成了4-(反-4′-正丙基环己基) 环己基甲酸(3HHA).研究了工艺条件对3HPA转化率的影响,得出最佳合成反应条件是m(Ru/C) m(3HPA) =5 100;反应温度105 ℃;反应压力5 MPa,3HPA的转化率接近100.0%.     

锌(Ⅱ) - 2,2''-联吡啶络合吸附波法测定血清白蛋白

在pH值6.3的HAc-NaAc介质中,血清白蛋白使锌(Ⅱ) - 2,2′-联吡啶络合物在-1.20 V(vs.SCE)处的络合吸附波还原峰电流降低,峰电流降低值与加入的牛血清白蛋白(BSA)或人血清白蛋白(HSA)的浓度在一定范围内呈线性关系.BSA和HSA的线性范围分别为0.5～40.0 mg·L-1、0.5～50.0 mg·L-1,检出限均为0.2 mg·L-1.应用该法测定了人血清样品中总蛋白含量,结果令人满意.     

4-[P-（1-辛基-4，4＇-联吡啶）甲基苯基]-6-苯基-2，2-联吡啶多齿配体的合成

以溴辛烷和4,4’-联吡啶为原料合成了4-[P-（1-辛基-4,4’-联吡啶）甲基苯基]-6-苯基-2,2-联吡啶多齿配体,通过。HNMR确定了所合成的化合物的结构。用循环伏安法和紫外可见分光光度法对化合物的电化学性质、光谱性质进行了研究。     

Ultrasensitive Electrochemiluminescence Immunoassay for Protein Specific Detection Based on Dendrimer-Encapsulated Gold Nanoparticles Labels

A sensitivity-enhanced electro chemiluminescence immunoassay (ECLIA) was fabricated by covalently immobilizing a monoclonal prostate specific antigen (PSA) antibody (anti-PSA, Ab2) and a luminophore (luminol) on dendrimer-encapsulated gold nanoparticles (Den/AuNPs) as electrochemiluminescence labels. The primary antibody was immobilized on Fe₃O₄@SiO₂ NP support, and the antibody-loaded Fe₃O₄@SiO₂ NP was placed onto an indium tin oxide working electrode in a home made electrochemiluminescence (ECL) cell. PSA and Ab2/Luminol/Den/AuNP was successively injected into the cell, and conjugated to form a sandwich-type immunocomplex. Under optimized experimental conditions, the proposed ECLIA provided a linear response range from 0.001 to 100.0 ng/mL with a low detection limit of 0.3 pg/mL. The PSA assay results in clinical serum samples were in good agreement with the commercially available electro chemiluminescence assay. The ECL immunosensor has the advantages of high sensitivity, specificity and stability and may be a promising technique for tumor marker detection.     

Electrochemical immunoassay for CA125 based on cellulose acetate stabilized antigen/colloidal gold nanoparticles membrane

A novel separation-free electrochemical immunosensor for carcinoma antigen-125 (CA125) was proposed based on the immobilization of CA125 antigen on colloidal gold nanoparticles that was stabilized with cellulose acetate membrane on a glassy carbon electrode. A competitive immunoassay format was employed to detect CA125 antigen with horseradish peroxidase (HRP) labeled CA125 antibody as tracer, o-phenylenediamine and hydrogen peroxide as enzyme substrates. After the immunosensor was incubated with a mixture of HRP labeled CA125 antibody and CA125 sample at 35 °C for 50 min, the amperometric response decreased with an increasing CA125 concentration in the sample solution. The decreased percentage of the electrocatalytic current was proportional to CA125 concentration ranging from 0 to 30 U ml⁻¹ with a detection limit of 1.73 U ml⁻¹ (S/N = 3). The proposed immunosensor showed good stability, acceptable accuracy, and would be applicable to clinical immunoassay of CA125.     

An Ultrasensitive Electrochemiluminescence Immunoassay for Carbohydrate Antigen 19-9 in Serum Based on Antibody Labeled Fe3O4 Nanoparticles as Capture Probes and Graphene/CdTe Quantum Dot Bionanoconjugates as Signal Amplifiers

The CdTe quantum dots (QDs), graphene nanocomposite (CdTe-G) and dextran–Fe₃O₄ magnetic nanoparticles have been synthesized for developing an ultrasensitive electrochemiluminescence (ECL) immunoassay for Carcinoembryonic antigen 19-9 (CA 19-9) in serums. Firstly, the capture probes (CA 19-9 Ab1/Fe₃O₄) for enriching CA 19-9 were synthesized by immobilizing the CA 19-9’s first antibody (CA 19-9 Ab1) on magnetic nanoparticles (dextran-Fe₃O₄). Secondly, the signal probes (CA 19-9 Ab2/CdTe-G), which can emit an ECL signal, were formed by attaching the secondary CA 19-9 antibody (CA 19-9 Ab2) to the surface of the CdTe-G. Thirdly, the above two probes were used for conjugating with a serial of CA 19-9 concentrations. Graphene can immobilize dozens of CdTe QDs on their surface, which can emit stronger ECL intensity than CdTe QDs. Based on the amplified signal, ultrasensitive antigen detection can be realized. Under the optimal conditions, the ECL signal depended linearly on the logarithm of CA 19-9 concentration from 0.005 to 100 pg/mL, and the detection limit was 0.002 pg/mL. Finally, five samples of human serum were tested, and the results were compared with a time-resolved fluorescence assay (TRFA). The novel immunoassay provides a stable, specific and highly sensitive immunoassay protocol for tumor marker detection at very low levels, which can be applied in early diagnosis of tumor.     

Synthesis of metal-carbonyl-dendrimer-antibody immunoconjugates: towards a new format for carbonyl metallo immunoassay

We report the preparation of metal-carbonyl-dendrimer-antibody conjugates. These metal-carbonyl-multilabeled antibodies are designed to be used in a new solid-phase-format carbonyl metallo immunoassay (CMIA). A fourth-generation polyamidoamine dendrimer was labeled with 10-25 (eta5-cyclopentadienyl)iron dicarbonyl (eta1-N-succinimidyl) entities. An antibody was chemically modified at its carbohydrate chains by a site-directed process used to preserve the antigen-antibody binding site. The antibody was then coupled with the dendrimer labeled with 10 metal carbonyl groups. An average of 1.4 labeled dendrimers were grafted per antibody molecule. These metal-carbonyl-dendrimer-antibody conjugates were used as new universal detection reagents that recognize their specific antigens. The antigens were spotted onto nitrocellulose membranes and detected by using the conjugates in combination with Fourier transform infrared spectroscopy. A detection level in the range 5-200 pmol per membrane was achieved. This approach opens the way to a new CMIA format.     

前列腺特异抗原光激发化学发光免疫检测方法的建立

目的建立前列腺特异抗原(Prostate specific antigen,PSA)光激发化学发光免疫(Light induced chemilumines-cent immunoassay,LICA)定量检测方法。方法用PSA多克隆抗体包被受体微粒,PSA单克隆抗体标记生物素,两者与链霉亲和素包被的供体微粒共同组成检测试剂检测PSA,优化测定条件并对方法进行验证。分别采用本方法与Roche公司电化学发光免疫分析法对82份临床标本进行检测,并进行比较分析。结果本方法的分析灵敏度为0.31 ng/ml;批内变异系数为4.66%～6.75%,批间变异系数为5.68%～8.42%;回收率为95.1%～105.2%。两种方法具有较好的相关性,其R2为0.981 5。结论建立的均相化学发光免疫测定方法能够用于血清PSA的定量测定,且其性能指标符合临床要求。     

Gold nanoparticle/DNA/methylene blue nanocomposites for the ultrasensitive electrochemical detection of carcinoembryonic antigen

A new signal amplification strategy employing nanocomposites composed of gold nanoparticles (GNPs) with DNA and methylene blue (MB) as labels has been developed to construct a novel immunosensor for the highly sensitive bioanalysis of carcinoembryonic antigen (CEA). To fabricate the label, DNA was first linked to GNPs through the formation of an Au–S bond. Next, the resulting bioconjugates were sequentially bound with MB via the guanine (G) bases in DNA specifically and then with secondary CEA antibody (Ab₂) via the crosslinking of glutaraldehyde (GA). The receptor–analyte complex is formed by the biorecognition of the as-obtained MB labeled Ab₂ with the primary CEA antibody (Ab₁) immobilized on the surface of a modified GNP/Chitosan (Chits) composite electrode. After the receptor–analyte complex has formed, square wave voltammetry (SWV) was employed to oxidize the labeled MB in order to detect the CEA. This immunosensor shows a linear response from 0.10 to 2.0 pg mL⁻¹ CEA with an improved detection limit of 0.05 pg mL⁻¹ as compared to the conventional immunoassay. In addition, this new protocol shows acceptable stability, reproducibility and good recovery (97.5–110.0%) for CEA in human serum with great potential for clinical applications.     

农药人工抗原的合成研究进展

免疫分析法是以抗原、抗体的特异性识别和可逆性结合反应为基础,以抗体作为生物检测器而建立的一种灵敏、简便、快速的超微量分析方法。抗原的质量决定了抗体的特异性和亲和性,最终影响免疫分析的质量。在农药残留的分析中,建立免疫分析方法的关键是农药人工抗原的合成,包括半抗原的设计(强调连接臂的位置、长度、性质和功能基团的引入)、半抗原的合成、载体的选择、半抗原与载体的偶联方法和条件、最佳结合比的讨论、人工抗原的纯化与鉴定。     

新型癌抗原125化学发光免疫试剂盒的制备

目的制备新型癌抗原125(CA125)化学发光免疫试剂盒,并进行验证。方法用1株针对CA125蛋白特异性位点的单克隆抗体作为包被抗体,1株针对CA125分子糖链的碱性磷酸酶(ALP)标记单抗和另1株针对蛋白特异性位点的ALP标记单抗分别作为酶标抗体,采用双抗体夹心法,金刚烷增敏化学发光体系作为酶底物制备试剂盒,检测CA125,并对试剂盒进行验证。结果新型CA125试剂盒检测灵敏度达0.1U/ml,测定范围在0.1~430U/ml之间,与传统CA125试剂盒一致;与CA199、CA153和CA50糖蛋白无交叉反应;精密性和准确性良好。与传统CA125试剂盒比较,检测结果有显著性差异,特别是在卵巢癌标本检测方面,糖链单抗酶标抗体制备的试剂盒与普通抗体酶标抗体制备的试剂盒检测结果的比值与其他标本两者的比值之间差异具有统计学意义。结论已制备了新型CA125化学发光免疫试剂盒,其与传统试剂盒检测标本结果的比值更适用于对卵巢癌进行特异性诊断。     

应用ELISA检测小鼠肝炎病毒抗体

以 DBT 细胞增殖的小鼠肝炎病毒 MHV-3经差速离心浓缩后作为抗原,用辣根过氧化物酶标记的金黄色葡萄球菌 A 蛋白(HRP-SPA)作为第二抗体,通过酶联免疫吸附试验(ELISA)检测 MHV抗体。经本中心及三个外单位共检测362份普通群小鼠血清样本,MHV 抗体检出率为149/362(41.16%)屏障系统内小鼠血清样本121份,均为阴性。     

人催乳素化学发光免疫分析试剂盒的制备及验证

目的制备人催乳素(hPRL)化学发光免疫分析(CLIA)试剂盒,并进行验证。方法采用2株不同结合位点的抗hPRL单克隆抗体,1株用于包被微孔板,另1株用于标记HRP,建立双抗体夹心检测系统,配合化学发光底物组装成试剂盒,并进行各项技术指标的验证。结果试剂盒最佳定量范围在5～100ng/ml,标准曲线的相关系数r=0.9999,灵敏度为0.49ng/ml,平均回收率为100.9%,试验内和试验间变异系数分别小于6.3%和10.9%。试剂盒有效期可达1年,特异性良好,出现Hook效应的样品浓度为1500ng/ml。与进口化学发光试剂盒和放射免疫分析试剂盒对比检测84份临床标本的结果呈高度相关,相关系数分别为0.9537和0.9486。结论已成功制备灵敏、特异的hPRLCLIA试剂盒,可用于人催乳素的临床检测。