Changes of serum p53 antibodies and clinical signifi cance of radiotherapy for esophageal squamous cell carcinoma

AIM： To explore the relationship between serum p53 antibodies （p53-Abs） and clinicopathological characteristics and therapeutic effect in patients with esophageal carcinoma （EC）, and to investigate sequential changing regularity of serum pS3-Abs after radiotherapy. METHODS： The serum pS3-Ab levels were detected in 46 EC patients and 30 healthy adults by enzyme linked immunosorbent assay （ELISA）. The blood samples were collected on the day before radiotherapy and on the administration of an irradiation dose of 20 Gy/10 f/12 d, 40 Gy/20 f/24 d and 60 Gy/30 f/36 d after radiotherapy. RESULTS： The level and positive rate of serum pS3-Abs in EC patients were significantly higher than those in normal individuals （P 〈 0.05）. Serum anti-p53 antibodies were positive in 18 of 46 EC patients （39.1%）. The positive rate of pS3-Abs in EC was related to histological grade, disease stage and lymph node metastasis （P 〈 0.05）, but it was not significantly related to sex, age and to the size and site of tumor. The level and positive rate of p53-Abs had significant differences between before radiotherapy and after administration of an irradiation dose of 40 Gy/20 f/24 d and 60 Gy/30 f/36 d （P 〈 0.05 orP 〈 0.01）. The positive rate of p53-Abs in EC patients with effect was significantly lower than that in those without effect after radiotherapy （P 〈 0.0001）.CONCLUSION： Detection of serum p53-Abs is helpful to the diagnosis of esophageal carcinoma. Monitoring for sequential change of serum p53-Abs before and after radiotherapy in patients with esophageal carcinoma is also useful to evaluate the response to the treatment and prognosis of the patients.

Serum Helicobacter pylori Kat A and Ahp C antibodies as novel biomarkers for gastric cancer

AIM: To investigate catalase (KatA) and alkyl hydroperoxide reductase (AhpC) antibodies of Helicobacter pylori as biomarkers for gastric cancer (GC). METHODS: This study included 232 cases and 264 controls. Recombinant KatA and AhpC proteins were constructed and the levels of antibodies were tested by indirect enzyme-linked immunosorbent assay (ELISA). Logistic regression was applied to analyze the relationships between KatA, AhpC and GC. The chi(2) trend test was used to evaluate the dose-response relationships between serum KatA and AhpC antibody levels and GC. Receiver operating characteristic (ROC) curve was used to evaluate the screening accuracy of KatA and AhpC as biomarkers. Combined analysis was used to observe screening accuracy of predictors for GC. RESULTS: In all subjects, the association between KatA and AhpC and GC risk was significant (P < 0.001) with odds ratio (OR) = 12.84 (95%CI: 7.79-21.15) and OR = 2.4 (95%CI: 1.55-3.73), respectively. KatA and AhpC antibody levels were strongly related to GC risk with a dose-dependent effect (P for trend < 0.001). The area under the ROC (AUC) for KatA was 0.806, providing a sensitivity of 66.81% and specificity of 86.36%; and the AUC for AhpC was 0.615, with a sensitivity of 75.65% and specificity of 45.49%. The AUC was 0.906 for KatA and flagella protein A (FlaA) combined analysis. CONCLUSION: Serum KatA and AhpC antibodies are associated with GC risk and KatA may serve as a biomarker for GC. KatA/FlaA combined analysis improved screening accuracy.

<i>Blastomyces dermatitidis</i>Antibody Responses in Serial Serum Specimens from Dogs with Blastomycosis: Comparison of Different Yeast Lysate Antigens

The systemic fungal organism, Blastomyces dermatitidis causes blastomycosis in animals and hu-mans. This study was designed to evaluate antibody detection in 55 serial serum specimens from 9 dogs with blastomycosis using B. dermatitidis yeast lysate antigens produced from two human isolates (B5896;B5931) and two dog isolates (ERC-2;T-58) with the indirect enzyme linked im-munosorbent assay (ELISA;peroxidase system) to determine an optimal lysate antigen(s) for use in the ELISA to detect antibody in the dog serum specimens. The mean absorbance values when the lysate antigens were compared with respect to their ability to detect antibody in the day 0 sera from the 9 dogs were 1.024 (ERC-2), 1.351 (B5896), 1.700 (B5931) and 2.084 (T-58) respectively. All of the reagents exhibited a high level of sensitivity and in all instances the amount of antibody declined as the time interval post-treatment increased, but the T-58 lysate prepared from the dog isolate from Tennessee was the optimal reagent. We continue to evaluate antigens for B. derma-titidis antibody detection in different immunodiagnostic assays.

Screening of Serum Dilution and Determination of Normal Range of Antibodies against Rabies Viruses in Dog Sera

[ Objective] To screen a suitable serum dilution and determine the normal range of antibodies against rabies viruses （RV） in dog serum. [Method]A sensitive, specific and suitable serum dilution was screened. A total of 812 dog serum samples were collected from Changchun, Nanning and Uuzhou regions, and then they were diluted with above screened serum dilution and evaluated by indirect enzyme-linked immunosorbont assay （ELISA）. The positive and negative standard dog sara from the World Organization for Animal Health （OIE） were used as controls. The serum samples with different A450 were randomly selected and the anti-RV serum titers were detected by rapid fluorescent focus inhibition test （RFFIT）. According to the A450 of dog sera, the normal A450 range of negative serum and 95% confidence intervals were calculated with SPSS 10.0 software, and the regression equation of OIE standards was established. [ Result] The screened serum dilution was sensitive and specific; at least 756 negative serum samples were obtained; the normal range and 95% confidence interval of negative serum were 0.127 3 ± 0.059 8 and 0.078 1 -0.172 3, respectively; and the regression equation was A450 = 1.139 serum titer （IU） ±0.470. [ Conclusion] These results lay a foundation for thecontrolling of dog rabies endemic and the development of ELISA kits of dog antibodies against RV.

Detection of Serum Antibodies of Porcine Pseudorabies Virus(PRV)in Binzhou City in 2014

To understand immunization effect of pseudorabies vaccine and infection status of porcine pseudorabies （PR） in pig farms and guide prevention and control against PR, 453 copies of blood samples collected from 43 different scale pig farms in four counties （districts） of Binzhou City were detected with ELISA to investigate PRV gB antibody and gE antibody. Detection results showed that the gB antibody positive rate of sows was 75.58%, and that of fattening pigs was 68.67% ; the pig farms with positive rate higher than 70% accounted for 74.42% of total survey pig farms. The PRV gE antibody positive rates of sows and fatte- ning pigs were 25.41% and 26.67%, and the positive rates of pig farms were 46.51% and 44.33%, respectively. There were regional differences among counties （districts）.

Role of GM3 ganglioside in the pathology of some progressive human diseases and prognostic importance of serum anti-GM3 antibodies

Glycosphingolipids(gangliosides)have been characterized as important biological molecules with a key role as regulators in many physiological processes on cellular,tissue,organ,and organism levels.The deviations in their normal amounts,production,and metabolism are very often related to the development of many multi-factor socially important diseases.GM3 ganglioside,as a small molecule,plays important roles in the cascade regulatory pathways in the pathology of many disorders like neurodegenerative diseases,autoimmune diseases,inflammation,diabetes,malignant transformation,and others.Ganglioside GM3 and its derivatives are membrane-bound glycosphingolipids composed of an oligosaccharide head structure containing one sialic acid residue.These molecules transduce signals involved in cell surface events,including the phosphorylation of transmembrane receptors.This ganglioside is the most widely distributed among tissues,and it serves as a precursor for most of the more complex ganglioside species.GM3 inhibits the function of fibroblast growth factor receptor,and cell growth is regulated by GM3-enriched microdomain.GM3 is thought to inhibit immunologic functions,such as the proliferation and production of cytokines by T cells.On the other hand,the anti-ganglioside antibodies(AGAs)are important in many acquired demyelinating immunemediated neuropathies,like Multiple sclerosis(MS),Guillain–Barrésyndrome(GBS)and its variation,Miller–Fisher syndrome(MFS)and could be suggested as important diagnostic and prognostic markers about the describe diseases and their etiology.We show that the complexes of anti-ganglioside antibodies to GM3(detected by ELISA)may be useful diagnostic and prognostic tool markers for autoimmune diseases,neurodegenerative disorders,malignancy,diabetes,and inflammation.Our pilot studies suggest increased serum IgG anti-GM3 antibodies titers in patients with secondary progressive MS(SPMS),throat cancer,elder people with diabetes(89–96 years),old Lewis rats(30–33 months),and in the serum of subjected on lead intoxication BALB/c mice treated by salinomycin.We observed no changes in the titers in healthy elder people(89–96 years),in 70-year-old woman on dialysis,in relapsing-remitting MS(RRMS)patients on long-term treatment with Glatiramer acetate,Laquinimod,and Interferons,as well as in 18–22 months old Wistar rats and subjected on lead intoxication BALB/c mice treated by monensin and dimercaptosuccinic acid(DMSA).Considerable decrease of serum GM3 in early MS correlate with early damage and severe destruction of the blood–brain barrier,which provides impetus to initiate early therapy.

Development of a Multi-Plex Electrochemiluminescent Assay for the Detection of Serum Antibody Responses to Meningococcal Conjugate Vaccines

Neisseria meningitidis is a gram negative diplococcal bacterium. Worldwide, N. meningitidis is the leading cause of bacterial meningitis and sepsis, with five serogroups (A, B, C, Y, and W-135) responsible for the majority of the disease. Multivalent (A, C, Y, and W-135) polysaccharide and conjugate vaccines have been licensed in the United States and elsewhere and are widely available. We have developed a multi-plexed electrochemiluminescent assay to quantitate serum antibody responses to meningococcal polysaccharides A, C, Y, and W-135 to allow for rapid evaluation of li- censed and investigational vaccines. A 96-well plate containing a carbon electrode arrayed with polysaccharides A, C, Y, and W-135 on separate spots within each well has been developed for simultaneous detection of polysaccharidespecific antibodies in serum samples from vaccinated individuals. The assay conditions were optimized using the anti-meningococcal serogroup A/C reference serum pool, CDC 1992 (NIBSC 99/706), through evaluation of plate types, coating polysaccharide concentrations, and blocking and serum diluent buffers. Comparison of single and multiplex assays demonstrated the sensitivity, specificity, and speed of the multi-plex format for the quantification of serum antibody responses to N. meningitidis polysaccharides A, C, Y and W-135.

Emergent Serum Therapy and Antibody Medicine to Counteract Sudden Attacks of COVID-19 and Other Pathogenic Epidemics

The science behind two life-saving protocols to combat COVID-19 and future pathogenic epidemics is presented for immediate implementation.

Analysis of Detecting HIV-1 Antibody in Paired Urine and Serum Specimens from Drug Users by ELISA

Objective- To compare the consistency of the results from detecting HIV-1 antibody in the paired urine and serum specimens from drug users by ELISA. Methods: The paired urine and serum specimens from 273 drug users detained at a detoxification unit were collected, and the HIV-1 antibodies in the specimens of them were screened by urine and serum ELISA kits, respectively. Results: Of 273 serum specimens, 94 ones showed positive reaction and among 94 counterpart urine specimens, 93 ones also appeared positive reaction. Taking the results together,the consistent rate of HIV-1 antibody screened by urine and serum ELISA kits was 99.6%. Conclusion: The urine ELISA kit, which screened HIV-1 antibody of urine showing almost the same results tested by serum ELISA kit, is reliable. It is proposed that urine ELISA be introduced in many fields.

Potential of soluble CD26 as a serum marker for colorectal cancer detection

Colorectal cancer is characterized by a low survival rate even though the basis for colon cancer development,which involves the evolution of adenomas to carcinoma,is known.Moreover,the mortality rates continue to rise in economically transitioning countries although there is the opportunity to intervene in the natural history of the adenoma–cancer sequence through risk factors,screening,and treatment.Screening in particular accounted for most of the decline in colorectal cancer mortality achieved in the USA during the period 1975-2000.Patients show a better prognosis when the neoplasm is diagnosed early.Among the variety of screening strategies,the methods range from invasive and costly procedures such as colonoscopy to more low-cost and non-invasive tests such as the fecal occult blood test(guaiac and immunochemical).As a non-invasive biological serum marker would be of great benefit because of the performance of the test,several biomarkers,including cytologic assays,DNA and mRNA,and soluble proteins,have been studied.We found that the soluble CD26(sCD26)concentration is diminished in serum of colorectal cancer patients compared to healthy donors,suggesting the potential utility of a sCD26 immunochemical detection test for early diagnosis.sCD26 originates from plasma membrane CD26 lacking its transmembrane and cytoplasmic domains.Some 90%–95%of sCD26 has been associated with serum dipeptidyl peptidase IV(DPPIV)activity.DPP-IV,assigned to the CD26 cluster,is a pleiotropic enzyme expressed mainly on epithelial cells and lymphocytes.Our studies intended to validate this test for population screening to detect colorectal cancer and advanced adenomas are reviewed here.

System Engineering of Emergent Serum Therapy to Combat COVID-19 and Other Pathogenic Pandemics

Life-saving procedures of an emergency protocol to combat fatal ambushes of viral or bacterial pathogens are listed below for immediate dissemination and implementation. This unique protocol of serum therapy commands the highest benefits-versus-risks ratio, and is safe, efficacious, user-friendly and inexpensive to implement. A formal evaluation is presented to support upgrading convalescent plasma therapy to serum therapy in the context of presenting fewer donor antigens, thus evoking less lethal sensitization to obtain greater safety and efficacy in treatment.

肺炎痰液支原体DNA联合血清MP-IgM抗体检测在小儿肺炎支原体肺炎早期诊断中的价值

目的分析肺炎痰液支原体DNA(MP-DNA)联合血清MP-Ig M抗体检测在小儿肺炎支原体肺炎早期诊断中的价值。方法选择疑似肺炎支原体感染的患儿213例,分别使用痰液MP-DNA法、血清MPIg M抗体法进行检测,比较两种方法及检测方法联合使用的诊断价值。结果 213例疑似肺炎支原体(MP)感染患儿中,共确诊MP感染患儿100例(46.95%);两种方法联合检测特异度、灵敏度、阴性预测值、阳性预测值均优于单独一种方法检测(P<0.05);痰液MP-DNA检测准确率优于血清MP-Ig M抗体检测准确率,但差异无统计学意义(P>0.05)。结论在小儿肺炎支原体肺炎早期诊断中,使用痰液MP-DNA法联合血清MPIg M抗体法进行检测,能明显提高其诊断准确率,更利于临床用药指导。

Establishment and application of immunofluorescence using monoclonal antibody in diagnosis of gastric cancer

Monoclonal antibodies against gastric cancer(MG)were purified by ion-exchangechromatography on DEAE-cellulose(DE-52)column and coupled with CNBr-activated Sepharose4B.The Sepharose 4B linked with MG monoclonal antibodies was then incubated with serum andascitic fluid of patients with gastric cancer and benign diseases for 2h.After being blocked by nor-mal mouse serum,the MG-Sepharose was mack to react with the MG-labeled with fluoresceinisothiocyanate for 30min.According to immunoaffinity chromatography principle,the MG-labeledwith fluorescein isothiocyanate was separated from the MG-Sepharose with 0.01 mol/L of PBS-3mol/L of KCNS solution and then detected with a fluorospectrometer.The amount of fluorescencein the solution of separation represented the amount of the antigens defined by MG monoclonalantibodies.The mean value plus 2 standard deviations of the benign disease group was arbitrarilyset as their positive threshold.The positivity rates were 61.1%(22/36)in serum and 75%(18/24)inascitic fluid of patients with gastric cancer.It suggests that the determination of MG-Ags in the se-rum and ascitic fluid of patients may be helpful for the diagnosis of gastric cancer.

2017—2021年云南省猪瘟病原学和血清学监测情况分析

为全面了解2017—2021年云南省猪瘟(CSF)流行及免疫状况,预估疫病流行和发生风险,采集病原学样品63 224份、血清学样品146 547份,进行CSF病原和血清抗体监测。结果显示:2017—2021年云南省CSF免疫抗体合格率分别为82.94%、81.47%、80.79%、80.24%、82.53%,规模场合格率略高于散养户,但差异不显著(P>0.05);CSF病原核酸阳性率分别为0.52%、0.74%、1.10%、1.02%、2.42%,散养户、屠宰场阳性率高于规模场。结果表明:自2017年CSF退出强制免疫后,云南省CSF免疫抗体合格率均达到国家标准,但病原阳性率整体呈上升趋势,且分布范围广,免疫密度低和抗体合格率低的散养户发生CSF的风险较高,提示应对其加强监测和免疫。

Some biological characteristics of hybridomas and parental myelomas cultivated in serum-free medium for long-term period

It has been developed in this laboratory that a serum-free medium, designated asDMI,is adaptive for long-term culture,freezing and resurgence in liquid nitrogen,and cloningof hybridoma and parental myelomas as well as for cell fusions.In this report,it is describedthat the myeloma cells grown in DMI for more than 3 months maintained their biological charac-teristics such as induction of aseites and subcutaneous somatic tumor in BALB/c mice and toler-ance toward 8-azaguanine,ete..However,the ability of the tumor cells to attach to glass wallwas decreased.The selecting assay of these cells in HAT medium(medium containing hypoxan-thine,aminopterin and thymidine)showed that the death time in DMI was similar to that in theconventional 15％ newborn calf serum-supplemented medium(RPS15).The hybridoma cellsadapted in DMI secrete monoclonal antibodies in quantities comparable to those produced inRPS15.

H5N1亚型禽流感病毒神经氨酸酶蛋白在昆虫细胞中的表达与鉴定

研究旨在利用昆虫细胞-杆状病毒表达系统制备H5N1亚型禽流感病毒(AIV)的神经氨酸酶(NA)蛋白。试验为获得具有良好反应原性的NA蛋白,将H5N1亚型AIV(DK/YN/S4469/2022)密码子优化后的N1基因插入载体pFastBac1,构建的重组转移载体pFastBac1-N1转化至DH10Bac感受态细胞,经过蓝白斑筛选获得阳性重组杆粒rBacmid-N1,然后将重组杆粒转染至sf9昆虫细胞,获得N1亚型重组杆状病毒。结果显示:N1重组蛋白在sf9昆虫细胞中正确表达,并且该重组蛋白具有良好的反应原性,能够与N1蛋白单克隆抗体3F3发生特异性反应,其仅能识别N1亚型鸡血清,特异性较好。研究成功制备了N1重组蛋白,为NA蛋白表达方法提供参考并为临床血清样品的N1抗体检测方法的建立奠定基础。

新疆某规模化猪场3种病毒病抗体监测分析与免疫程序优化

该试验旨在了解当前新疆巴州某规模化猪场猪瘟(CSF)、猪蓝耳病(PRRS)和伪狂犬(PR)疫苗的免疫水平,收集当地某规模化生猪养殖场的467个血清样品,采用ELISA抗体检测试剂盒检测血清中猪瘟、猪蓝耳病和猪伪狂犬病毒的抗体水平。结果显示,该地区的猪瘟、猪蓝耳病和猪伪狂犬病毒平均血清抗体阳性率分别为93.68%、77.22%和92.65%,均超过我国强制性疫苗免疫抗体阳性率标准70%。该地区猪群免疫手段还有很大的进步空间,本研究结合该地区实际情况对免疫程序进行了相应的调整优化。

NT与血清β-hCG、PAPP-A、ADAM12单独及联合检测对孕早期DS的诊断价值

目的:探讨颈部透明层厚度(NT)与血清人绒毛膜促性腺激素(β-hCG)、妊娠相关蛋白A(PAPP-A)及解整合素金属蛋白酶12(ADAM12)单独及联合检测对孕早期唐氏综合征(DS)的诊断价值。方法:选取2019年1月—2023年1月在厦门大学附属第一医院杏林分院经羊膜穿刺确诊的47例DS孕妇作为DS组,另选取同期49例产前检查正常的孕产妇作为对照组,检测并比较两组NT及血清β-hCG、PAPP-A、ADAM12水平,并采用受试者工作特征(ROC)曲线分析NT与血清β-hCG、PAPP-A、ADAM12单独及联合检测对DS的诊断价值。结果:DS组NT及血清β-hCG水平高于对照组,PAPP-A、ADAM12水平均低于对照组,差异有统计学意义(P<0.05)。ROC曲线分析显示,NT与血清β-hCG、PAPP-A、ADAM12联合检测诊断DS的曲线下面积为0.993,预测敏感度、特异度分别为95.7%、98.0%,均高于上述指标单独检测。结论:NT与血清β-hCG、PAPP-A、ADAM12检测对孕早期DS均具有一定的诊断价值,且联合应用诊断效果更高,可作为孕早期DS筛查的有效指标。

白喉毒素突变体CRM197在大肠杆菌中优化表达及人群血清抗体水平分析

目的 构建白喉毒素突变体CRM197的原核表达载体,在大肠杆菌中表达CRM197蛋白。分析健康人群血清中的CRM197抗体水平。方法 对CRM197基因进行大肠杆菌密码子优化并克隆至pET28a(+)中,经表达条件优化后采用镍柱亲和层析和离子交换层析对重组蛋白CRM197进行纯化,对纯化的CRM197进行Western blot鉴定。最后,经纯化后的CRM197蛋白与健康人群血清进行反应,检测人群血清中针对该蛋白的抗体水平。结果 CRM197蛋白进行了密码子及表达条件优化后,获得了可溶性表达蛋白。经镍柱亲和层析和离子交换层析后其纯度可达95%以上,Western Blot鉴定后能得到单一的特异性条带。经纯化的CRM197蛋白与不同年龄阶段健康人群血清均发生不同程度的抗原抗体反应。结论 利用本研究的策略,CRM197得到高效的可溶性表达,但人群血清中含有高水平的CRM197抗体,这些抗体的存在可能会影响到应用CRM197结合多糖作为抗原进行特异免疫效果的评价以及作为诊断抗原的应用。