血清绿脓杆菌外毒素A抗体的测定

血清绿脓杆菌外毒素Ａ抗体的测定王厚东陈仁涉王小平李树芬缪竞智绿脓杆菌（ＰＡ）是院内感染的主要病原菌，外毒素－Ａ（ＰＥＡ）是ＰＡ毒性最强的毒力因子。我们以ＰＥＡ为抗原，建立了测定血清特异性抗ＰＥＡ抗体的酶联免疫吸附试验（ＥＬＩＳＡ）方法，对正常人、老年...     

孕妇IgG抗体效价的检测新方法探讨

目的:探讨在孕妇IgG抗体效价的检测预处理过程中不加入2-巯基乙醇(2-Me)处理液是否可行。方法:检测孕妇IgM和IgG抗体效价,对试验结果进行比较。结果:IgG抗体效价通常高于IgM抗体效价,IgM抗体效价通常在1∶64以下,IgM和IgG抗体在抗体效价检测过程中有协同凝集作用。结论:可以对IgM抗体效价通常在1∶64以下孕妇不用2-Me处理液处理直接检测IgG抗体效价,对个别IgM抗体效价通常在1∶64以上,IgG抗体效价未超过同一孕妇IgM抗体效价者则需要进行预处理。     

生物素—亲和素系统检测抗布鲁氏菌抗体的研究

本文将 ABC—ELISA 法引入布病检测中,通过不同人群的同一份血清与 SAT、2MET、Coomb's、SPA—ELIS 等方法平行比较分析,证实 ABC—ELISA 法对布病早期诊断与治疗,及流行病学检测,有现实意义。     

孕妇IgG血型抗体效价与ABO新生儿溶血病关系的探讨

目的:探讨孕妇IgG血型抗体效价与ABO新生儿溶血病发生率之间的关系。方法:对2741例妻O型Rh阳性夫非O型孕妇做IgG抗-A(B)效价测定,对其中286例有HDN危险的孕妇所生新生儿做HDN血型血清学检查(即ABO血型鉴定、Coomb's试验、游离和释放试验),以了解孕妇IgG血型抗体效价高低与ABO-HDN发病率之间的关系。结果:2741例O型孕妇中,IgG抗-A(B)效价<64、64、128、256和≥512各组分别占61.9%、17.0%、12.6%、6.1%和2.4%。286例母婴血型不合且有ABO-HDN危险的孕妇所生新生儿中共有71例HDN阳性,占24.8%;孕妇IgG抗-A(B)效价为64、128、256和≥512各组HDN阳性率分别为10.7%、28.3%、44.9%和64.3%(P<0.01)。结论:孕妇血清中IgG抗-A(B)效价高低与ABO-HDN的发生率成正相关,产前进行血清IgG抗体效价检测非常必要。     

血型A抗原模拟多肽与抗A抗体的分子对接

目的：从分子结构水平研究血型A抗原及其模拟多肽（肽P1 QIWYERTLPFTF,肽P2EYWYCGMNRTGC）与血型A抗体的相互作用。方法：利用Chimera软件构建血型A抗原模拟多肽（P1,P2）的三维结构;将天然血型A抗原及其模拟多肽通过Autodock软件与血型A抗体Fv段（PDB：1jv5）对接。结果：天然血型A抗原和多肽P1、P2都能与1jv5分子中一个由疏水性氨基酸组成的凹槽对接,且对接部位重叠。天然A抗原和P1可以与1jv5形成2个氢键,且天然抗原和模拟多肽都能够与1jv5中的多个氨基酸相合作用。结论：通过生物信息学计算软件成功模拟了血型A抗原及其模拟多肽与抗体1jv5的相互作用情况。此2个模拟多肽是从功能上对天然A抗原进行模拟的。     

RFFIT和MNT检测抗狂犬病毒中和抗体的比较研究

目的对RFFIT和MNT两种方法检测抗狂犬病毒中和抗体的相关性,敏感性和重复性进行比较。方法用MNT和RFFIT两种方法来检测已免疫的人血清52份,未免疫的人血清40份。结果显示这两种方法检测已免疫血清的一致性是88.5%,其相关性是r=0.886,MNT的敏感性和特异性分别是84.9%,100%;RFFIT的灵敏度和特异性分别是100%,89.0%。MNT和RFFIT重复检测一个样品的变异系数(CV)分别是62.3%、13.3%。结论RFFIT与MNT的相关性较好,RFFIT的灵敏度和重复性均比MNT好,因此,RFFIT在大多数需要检测RV中和抗体的情况下可替代MNT。     

微柱凝胶法与试管法检测新生儿溶血病IgG抗体效价的比较

目的：探讨微柱凝胶法在新生儿溶血病实验诊断中测定孕妇血清中IgG抗体效价的可行性及方法学。方法：孕妇血清用2-Me处理做倍量稀释后,采用微柱凝胶法测定IgG抗体效价。同时与试管法进行对照比较。结果：用微柱凝胶法和试管法同时检测孕妇血清中的IgG抗体效价,2位检验者检测结果符合率试管抗人球蛋白法为79．3％,微柱凝胶法为95．1％,效价〈64时,微柱凝胶法灵敏度75．6％（62／82）,试管抗人球蛋白法为86．6％（72／82）,效价≥64时,微柱凝胶法灵敏度24．4％（20／82）,试管抗人球蛋白法为13．4％（11／82）。采用Х^2。检验,Х^2=1.64,结果差异无统计学意义（P〉0．05）。结论：应用微柱凝胶法检测孕妇血清IgG抗体效价,能够使实验方法简单化、自动化,使实验操作标准化,判断结果直观,与试管法相比减少了肉眼判断结果时的人为误差。     

HIV-1中和抗体的作用与机制及实验室检测进展

人类免疫缺陷病毒Ⅰ型(HIV-1)中和抗体在防止病毒感染方面起到重要作用.最近的研究表明,中和抗体也能协同细胞免疫应答延缓感染者的疾病进程.HIV-1中和抗体主要通过结合HIV-1感染细胞过程中暴露的各种中间结构而干扰HIV-1进入靶细胞,从而阻断感染的发生.同时,病毒通过变异和外膜糖蛋白(Env)的高度糖基化发生免疫逃逸.目前中和抗体的检测主要依赖于体外中和试验,近年来研究者不断对该试验方案进行优化和标准化.对中和抗体作用的相关机制的逐渐阐明、相关检测技术的发展,必将推进疫苗研究的发展,并为疫苗评价提供新的可信的实验依据.     

抗P1抗体检测的血清学分析和方法探讨

目的:分析P1(-)表型个体产生抗P1抗体的血型、抗体鉴定的血清学检测结果、分子机制以及交叉配血策略,并回顾对比分析既往2例血清学检测方法和结果,对优化检测方法提高抗P1抗体检出率进行探讨。方法:采用盐水法、聚凝胺和微柱凝胶法进行血型鉴定、抗体筛选和交叉配血,采用PCR扩增α-1,4-半乳糖基转移酶基因(α-1,4-galactosyltransferase,A4GALT)和β-1,3-N-乙酰半乳糖氨基转移酶基因(β-13-N-acetylgalactosaminyltransferase,B3GALNT1)外显子编码区,分析DNA序列。结果:患者1经过盐水法、微柱凝胶法、抗人球蛋白法血清学分析鉴定为IgM抗P1抗体,经过PCR核酸检测确定为患者表型为P1(-);P型,验证与血清学的检测结果一致。患者2仅通过血清学检测鉴定为抗P1抗体。结论:抗P1抗体多为IgM型抗体,在进行抗体检测时需考虑到检测方法的适用范围和局限性,选择合适的方法,并进行多种方法联合检测相互验证的方式,以提高抗体的检出率。核酸检测基因分型技术的应用可以从分子生物学角度提供重要佐证,有助于提高抗体鉴定的准确率。     

铜绿假单胞菌外毒素A全长基因的扩增与克隆

目的　从绿脓杆菌中提取染色体DNA ,PCR扩增及克隆外毒素A全长基因 ,为实现外毒素A的高效表达奠定基础。方法　从绿脓杆菌培养物中提取染色体DNA ,PCR扩增外毒素A全长基因 ,A -T克隆入本室自行构建的pSK -T载体中 ,对重组质粒进行酶切与测序鉴定。结果与结论　成功地从高GC含量的绿脓杆菌染色体DNA中扩增到长片段的外毒素A全基因并进行了克隆与鉴定。     

Immunological Properties of Exotoxin A Toxoid - Detoxified Lipopolysaccharide - Gold Nanoparticles Conjugate Against Pseudomonas aeruginosa Infection

Background: Pseudomonas aeruginosa is an important opportunistic pathogen, especially in patients with compromised host defense. Objective: To prepare the conjugate of detoxified lipopolysaccharide (D-LPS) and exotoxin A toxoid (T-ETA) from P. aeruginosa in gold nanoparticles (Au NPs) in a mice model. Methods: LPS and ETA were purified from P. aeruginosa PAO1. D-LPS was conjugated with T-ETA via the amidation method. Au NPs were bound to D-LPS-T-ETA conjugate via electrostatic interaction. Mice were immunized with D-LPS, D-LPS-Au NPs, T-ETA, T-ETA-Au NPs, D-LPS-T-ETA, D-LPS-T-ETA-Au NPs, D-LPS-Au NPs+T-ETA-Au NPs, Au NPs, and phosphate-buffered saline (PBS), and specific IgG titers were determined by the ELISA and the whole-cell ELISA methods. Mice in the vaccinated and control groups were exposed to a 2×LD50 of P. aeruginosa and mortality rates were recorded for one week. Results: The results showed that vaccination by D-LPS, D-LPS-Au NPs, T-ETA, T-ETA-Au NPs, D-LPS-T-ETA, D-LPS-T-ETA-Au NPs and D-LPS-Au NPs+T-ETA-Au NPs induced specific IgG. Mice received the D-LPS-T-ETA-Au NPs conjugate showed significant protection against bacterial challenge. Conclusion: These data indicate that D-LPS-T-ETA-Au NPs conjugate has a significant immunogenicity potential to be applied as a new vaccine against Pseudomonas infections.     

重组铜绿假单胞菌外毒素A在大肠杆菌中的表达

目的实现铜绿假单胞菌外毒素A(PEA)全基因的原核载体构建及融合性表达。方法用基因重组技术构建PET-32a-PEA重组体,并在大肠杆菌BL21中进行表达。结果酶切鉴定及测序结果表明PET32a载体上成功地插入了PEA全基因。用IPTG诱导目的基因表达,沸水浴变性后进行SDS-PAGE电泳,在分子量78kD处有一条明显的新生蛋白带。表达的融合蛋白量约占细菌总蛋白的35%。结论成功地对PEA全基因进行了原核载体构建及表达,获得了稳定表达的工程菌,为进一步研出有效的基因工程疫苗和用于免疫导向治疗的重组毒素奠定了基础。     

Comparative efficacy of two doses of nebulized colistimethate for the eradication of Pseudomonas aeruginosa in children with cystic fibrosis

BACKGROUND:

Cystic fibrosis (CF) affects the respiratory and digestive systems. It evolves toward deterioration of pulmonary function through colonization with Pseudomonas aeruginosa. There is no consensus with respect to its eradication. Nebulized colistimethate is used for eradication treatment, but the optimal dose and duration is yet to be determined.

OBJECTIVES:

To compare the efficacy of two doses of nebulized colistimethate (30 mg versus 75 mg twice daily) for the eradication of P aeruginosa in children with CF and intermittent colonization.

METHODS:

A cohort study with both historical (30 mg) and prospective (75 mg) arms was conducted from 1999 to 2003. Medical records were used to collect data.

RESULTS:

Eighty-one patients were recruited in the retrospective group, for a total of 111 treatment courses. Twenty patients were recruited in the prospective group, for a total of 20 events. There was no statistically significant difference in the rate of eradication of P aeruginosa at days 28 and 90, neither when comparing the doses of colistimethate nor duration of treatment. There was a statistically significant difference (P=0.004) between days 1 and 90 in all analyzed subgroups (regardless of dose or duration of treatment) for forced vital capacity only. In the group of patients in whom eradication was achieved at day 28 (after receiving a three-week treatment course of colistimethate), 50% of patients developed a new infection 5.75 months later, on average, regardless of the dose administered. In the group of patients who achieved eradication at day 90 (after receiving a 15-week treatment course of colistimethate), 50% of the 14 patients developed a new infection after an average period of 7.3 months (P=0.28).

CONCLUSIONS:

There is no difference in the efficacy between a 30 mg dose and a 75 mg dose of colistimethate for P aeruginosa eradication in children with CF and intermittent colonization.     

Comparative Study of Efficacy of Plasma Exchange Versus Intravenous Gammaglobulin Treatment on Acute Postinfectious Polyradiculoneuropathy: A Preliminary Report

Abstract: We tried to compare the efficacy of plasma exchange (PE) with that of intravenous immunoglobulin (IVIG) in patients with postinfectious polyneuritis (Guil-lain-Barre syndrome [GBS] and cranial neuritis). Fifteen patients with postinfectious polyneuritis were divided into 2 groups. The IVIG group included 5 cases of GBS and 2 cases of postinfectious cranial neuritis (ophthalmoplegic type). The PE group included 5 cases of GBS and 3 cases of postinfectious cranial neuritis (ophthalmoplegic type). The changes and incidences of improvement of muscle strength scores (MSSs) and ocular movement scores (OMSs) were evaluated before treatment and 1, 2, 3, and 4 weeks after treatment. No significant differences between the IVIG and PE groups were found in the MSSs or OMSs at any time after treatment. These data suggested that PE and IVIG had equivalent efficacy. In the IVIG group, the proportion of suppressor-inducer T cells significantly increased (p ≤ 0.01) (before versus after treatment), and the proportion of suppressor-effector cells also increased but not significantly (before versus after treatment). In the PE group, the percentage of suppressor-inducer T cells significantly decreased (p ≤ 0.05) (before versus after treatment) while the proportion of suppressor/ cytotoxic T cells significantly increased (p ≤ 0.05) (before versus after treatment). The percentage of suppressor-effector T cells also increased (before versus after treatment) but not significantly.     

Comparative Investigation of Methods for Analysis of SARS-CoV-2-Spike-Specific Antisera

In light of an increasing number of vaccinated and convalescent individuals, there is a major need for the development of robust methods for the quantification of neutralizing antibodies; although, a defined correlate of protection is still missing. Sera from hospitalized COVID-19 patients suffering or not suffering from acute respiratory distress syndrome (ARDS) were comparatively analyzed by plaque reduction neutralization test (PRNT) and pseudotype-based neutralization assays to quantify their neutralizing capacity. The two neutralization assays showed comparable data. In case of the non-ARDS sera, there was a distinct correlation between the data from the neutralization assays on the one hand, and enzyme-linked immune sorbent assay (ELISA), as well as biophysical analyses, on the other hand. As such, surface plasmon resonance (SPR)-based assays for quantification of binding antibodies or analysis of the stability of the antigen–antibody interaction and inhibition of syncytium formation, determined by cell fusion assays, were performed. In the case of ARDS sera, which are characterized by a significantly higher fraction of RBD-binding IgA antibodies, there is a clear correlation between the neutralization assays and the ELISA data. In contrast to this, a less clear correlation between the biophysical analyses on the one hand and ELISAs and neutralization assays on the other hand was observed, which might be explained by the heterogeneity of the antibodies. To conclude, for less complex immune sera—as in cases of non-ARDS sera—combinations of titer quantification by ELISA with inhibition of syncytium formation, SPR-based analysis of antibody binding, determination of the stability of the antigen–antibody complex, and competition of the RBD-ACE2 binding represent alternatives to the classic PRNT for analysis of the neutralizing potential of SARS-CoV-2-specific sera, without the requirement for a BSL3 facility.     

A phase 2 study of aztreonam lysine for inhalation to treat patients with cystic fibrosis and Pseudomonas aeruginosa infection

BACKGROUND: Aztreonam lysine for inhalation (AZLI) is being developed for treatment of CF patients with Pseudomonas aeruginosa airway infection. METHODS: This double-blind, randomized, placebo-controlled Phase 2 study evaluated the safety, tolerability and efficacy of 75 and 225 mg AZLI administered BID for 14 days using the eFlow Electronic Nebulizer (Pari Innovative Manufacturers, Inc., Midlothian, VA). Patients were 13 years and older with FEV1>or=40% predicted, chronic P. aeruginosa infection, and had used no anti-pseudomonal antibiotics for 56 days. RESULTS: Of 131 patients screened, 105 received AZLI or placebo. Mean age was 26 years and mean FEV1 percent predicted was 77% at baseline. There was a statistically significant reduction, compared to placebo, in P. aeruginosa CFU density in each AZLI group at Days 7 and 14 (P<0.001). The planned primary analysis, percent change in FEV1 at Day 14, demonstrated no statistically significant difference. Post hoc analysis demonstrated significant increase in FEV1 at Day 7 for the subset of patients with baseline FEV1<75% predicted in the 225 mg AZLI group. Bronchodilator use was associated with greater improvement in FEV1, as well as greater reduction in P. aeruginosa bacterial density and higher plasma aztreonam concentrations in the 225 mg AZLI group. Adverse events were similar between placebo and AZLI although there was a trend toward increased respiratory symptoms in the 225 mg AZLI group. CONCLUSION: These data support the further development of AZLI and provide information for the design of subsequent studies.     

两种免疫抑制疗法治疗重型再生障碍性贫血比较研究

目的 :探讨重型再生障碍性贫血的适宜治疗方案。方法 :回顾性比较猪 -抗胸腺细胞球蛋白( P- ATG)单用 ( IST方案 )与联合方案兔 -抗人 T淋巴细胞球蛋白 ( R- ATL G)、环孢霉素 A( Cs A)、重组人粒 -巨噬细胞集落刺激因子 ( rhu GM- G- CSF) /重组人粒细胞集落刺激因子 ( rhu- G-CSF)及重组人红细胞生成素 ( rhu EPO) ( IST方案 )治疗 SAA疗效。结果 :IST方案 组不仅疗效高于 IST方案 组 ,且其有效者造血功能恢复更为迅速及良好。结论 :SAA患者更适宜于 IST方案 。