Measurement of salivary cortisol – effects of replacing polyester with cotton and switching antibody

Stable performance between‐runs are essential in longitudinal studies and when different studies are being compared. However, changes in analytical kits and laboratory material occur and have the potential to threaten analytical stability. In the present case, we examined how salivary cortisol measurements in our laboratory were affected by: 1) changes in the tampon material and 2) changes in the antibody of the analytical kit. In study 1, saliva from healthy subjects (n = 19) was split and spiked to Salivette^® polyester and cotton tampons, respectively, and treated as ordinary samples before being analysed for cortisol using a Spectria RIA kit for cortisol. In study 2, 68 anonymous saliva samples were analysed with the Spectria Cortisol RIA kit both before and after the manufacturer changed the antibody. The change from polyester to cotton tampons reduced the measured concentration of salivary cortisol by 62?%. A difference of 12?% between the two runs with different antibodies could not be attributed to differences in storage or in thawing and freezing of samples. To conclude, both a change in the material of the Salivette used for collecting saliva samples as well as a change of antibody in a kit can have a major impact on measurements, as illustrated here for concentrations of cortisol in saliva. It is therefore recommended always to check that the analysis stays in statistical control in one's own laboratory when changes are made, even if the manufacturer reports that the changes should have no effects.     

血清及胎盘中脂蛋白相关磷脂酶A2水平与妊娠期高血压疾病的关系

 目的 探讨血清及胎盘中脂蛋白相关磷脂酶A₂(Lp-PLA₂)与妊娠期高血压疾病的关系。方法 采用酶联免疫吸附法及免疫组织化学方法检测60例妊娠期高血压疾病患者(妊娠高血压19例、轻度子痫前期25例、重度子痫前期16例) 及20例正常妊娠妇女(对照组)血清及胎盘中Lp-PLA₂水平,并分析其与妊娠期高血压疾病的关系。结果 妊娠期高血压疾病组胎盘中Lp-PLA₂表达水平明显高于对照组(P<0.05)。轻度子痫前期组Lp-PLA₂表达水平显著高于对照组(P<0.05),重度子痫前期Lp-PLA₂表达水平显著高于轻度子痫前期组(P<0.05)。妊娠期高血压疾病组血清中Lp-PLA₂水平显著高于对照组(P<0.05)。随着病情的加重,Lp-PLA₂水平呈逐渐上升趋势,对照组、妊娠高血压、轻度子痫前期、重度子痫前期每两组之间比较,差异均有统计学意义(P均＜0.05)。妊娠期高血压疾病组血清与胎盘中Lp-PLA₂水平呈正相关(r=0.435,P<0.05)。结论 Lp-PLA₂水平的升高可能与妊娠期高血压疾病的发生有关,母血中Lp-PLA₂水平的变化有可能作为疾病的监测指标。     

几种猪囊尾蚴抗原用于EITB法诊断囊尾蚴病的观察

本研究对囊液10000g和100000g离心后的上清、头节抗原、全囊抗原及头节尿素溶解性抗原等5种抗原进行SDS－聚丙烯酰胺凝胶电泳。考马斯亮蓝染色依次出现10、10、13、14和5条显色带，银染结果依次出现20、20、21、21和13条显色带。分子量范围为＜6.6kDa－＞116kDa。选择囊液100000g（以下简称囊液）、全囊和头节尿素等3种抗原对105份各类血清进行酶联免疫电转移印斑（EITB）检测。结果囊尾蚴病患者血清与3种抗原反应可出现1－13条反应带，分子量范围6.6kDa－＞116kDa。如以≤21.5kDa条带作为判断标准，则囊液抗原、全囊抗原和头节尿素抗原的阳性率依次为54％、48％及58％，假阳性率依次为6％、10％及0％。头节尿素抗原对血吸虫、华支睾吸虫和肺吸虫病患者血清未出现交叉反应，10例包虫病患者血清中有2例出现交叉反应。     

Individual, household and community factors associated with HIV test refusal in rural Malawi

Objective To investigate individual, household and community factors associated with HIV test refusal in a counselling and testing programme offered at population level in rural Malawi. Methods HIV counselling and testing was offered to individuals aged 18–59 at their homes. Individual variables were collected by interviews and physical examinations. Household variables were determined as part of a previous census. Multivariate models allowing for household and community clustering were used to assess associations between HIV test refusal and explanatory variables. Results Of 2303 eligible adults, 2129 were found and 1443 agreed to HIV testing. Test refusal was less likely by those who were never married [adjusted odds ratio (aOR) 0.50 for men (95% CI 0.32; 0.80) and 0.44 (0.21; 0.91) for women] and by farmers [aOR 0.70 (0.52; 0.96) for men and 0.59 (0.40; 0.87) for women]. A 10% increase in cluster refusal rates increased the odds of refusal by 1.48 (1.32; 1.66) in men and 1.68 (1.32; 2.12) in women. Women counsellors increased the odds of refusal by 1.39 (1.00; 1.92) in men. Predictors of HIV test refusal in women were refusal of the husband as head of household [aOR 15.08 (9.39; 24.21)] and living close to the main road [aOR 6.07 (1.76; 20.98)]. Common reasons for refusal were fear of testing positive, previous HIV test, knowledge of HIV serostatus and the need for more time to think. Conclusion Successful VCT strategies need to encourage couples counselling and should involve participation of men and communities.     

IgA-mediated autoimmune hemolytic anemia in an infant

Autoimmune Hemolytic anemia (AIHA) a relatively uncommon form of hemolytic anemia in children, occurs due to the premature destruction of red blood cells caused by presence of autoantibodies directed against antigens on RBCs. Warm reactive AIHA is the most common form due to IgG isotype of immunoglobulin class binding to autologous RBCs at 37⁰C and confirmed with a positive DAT screening. We present a case of DAT-negative primary warm AIHA in an infant due to IgA antibody. A 10 month old male infant presented with dark colored urine and irritability for past two months, with associated history of fever, diarrhea and vomiting. He had received one red cell transfusion 10 days prior. On physical examination he had pallor with tachycardia without splenomegaly. On investigation his hemoglobin was 5.8 g/dl, WBC 25.9 × 10³/mm³ and normal platelets counts. Peripheral blood smear had spherocytes and biochemical values showed high bilirubin and LDH. Immunohematological work up revelaed polyspecific DAT was negative but monospecific DAT screening showed strong (4+) positivity for IgA and a weak IgG positivity. The patient was diagnosed as IgA-mediated Warm AIHA and was started on prednisolone at 2 mg/kg/day following which hemoglobin improved over the next 2 months. After 2 weeks, prednisolone was tapered and stopped by the end of 3 months. Patients with clinical and laboratory evidence of acute hemolysis, an additional screening for IgA antibody may be done even in cases where poly-specific DAT is negative. Early detection helps in avoiding further investigations and provide efficient management.     

Detection of small amounts of human adenoviruses in stools: comparison of a new immuno real-time PCR assay with classical tools

The detection of low virus concentrations in biological matrices, especially stool samples, is facing significant limitations as far as common diagnostic methods (enzyme-linked-immunosorbent assay (ELISA) or quantitative real-time PCR (qPCR)) are considered. Here the development of a new immuno real-time PCR (iPCR) is described and its performance in the detection of human adenoviruses (HAdVs) in spiked stools is compared with those of ELISA and qPCR assays. For the iPCR, detection of the sandwich formed by the complexation of capture antibody-antigen-detection antibody was performed by qPCR thanks to the substitution of peroxydase by a chimeric DNA. This modification increased the detection sensitivity 200-fold compared to ELISA. The direct qPCR results revealed that only 0.3–9.5% of the spiked HAdV were detectable, resulting from important losses of DNA occurring at the extraction step. This step was not necessary in the iPCR workflow, avoiding this drawback. The losses of viral particles occurred at the elution step from the stool only. The recovery rate of the iPCR was thus better and ranged between 21 and 54%. As a result, iPCR enabled the detection of lower virus concentrations in stool samples compared to those detected by ELISA and qPCR. The iPCR could be considered as a ‘hyper sensitive ELISA’ for early detection of HAdV infections, especially in the case of immunocompromised patients after haematopoietic stem cell transplant.