Synthetic Antigen Controls for Immunohistochemistry.

Immunohistochemistry (IHC) assays provide valuable insights into protein expression patterns, the reliable interpretation of which requires well-characterized positive and negative control samples. Because appropriate tissue or cell line controls are not always available, a simple method to create synthetic IHC controls may be beneficial. Such a method is described here. It is adaptable to various antigen types, including proteins, peptides, or oligonucleotides, in a wide range of concentrations. This protocol explains the steps necessary to create synthetic antigen controls, using as an example a peptide from the human erythroblastic oncogene B2 (ERBB2/HER2) intracellular domain (ICD) recognized by a variety of diagnostically relevant antibodies. Serial dilutions of the HER2 ICD peptide in bovine serum albumin (BSA) solution are mixed with formaldehyde and heated for 10 min at 85 °C to solidify and cross-link the peptide/BSA mixture. The resulting gel can be processed, sectioned, and stained like a tissue, yielding a series of samples of known antigen concentrations spanning a wide range of staining intensities. This simple protocol is consistent with routine histology lab procedures. The method requires only that the user have a sufficient quantity of the desired antigen. Recombinant proteins, protein domains, or linear peptides that encode relevant epitopes may be synthesized locally or commercially. Laboratories generating in-house antibodies can reserve aliquots of the immunizing antigen as the synthetic control target. The opportunity to create well-defined positive controls across a wide range of concentrations allows users to assess intra- and inter-laboratory assay performance, gain insight into the dynamic range and linearity of their assays, and optimize assay conditions for their particular experimental goals.

Depressive and Anxiety Symptoms among Pediatric In-Patients with Dengue Fever: A Case-Control Study.

BACKGROUND: Psychiatric symptoms have been reported in adult patients with dengue fever (DF); however, information on pediatric patients remains inadequate. We sought to identify the prevalence and predictors of depressive and anxiety symptoms and identify other psychiatric symptoms among pediatric patients with DF. This case-control study involved pediatric in-patients (n = 225) who had clinical or serologic-confirmed DF and healthy school-based controls (n = 260). Participants completed the Revised Child Anxiety and Depression Scale (RCADS). RESULTS: The prevalence of depressive (13.3%) and anxiety (34.2%) symptoms among pediatric patients with DF was significantly (p < 0.001) higher than that among controls (3.5% and 16.2%, respectively). Multiple linear regression analysis found that age, family history of DF, ≤2 days of hospitalization, myalgia, and arthralgia were predictors of increased depressive and anxiety symptoms among the patients. Further, 26.7% of pediatric patients reported irritability, agitation, visual hallucinations, and aggressiveness. CONCLUSION: Pediatric patients present depressive and anxiety symptoms whose levels were associated with social and clinical factors. However, whether these symptoms are present only during the infection or may still persist after recovery or are brought by children's adverse reactions to hospitalization are unknown, and thus, further studies are needed.

Knowledge, Attitude, and Practices Regarding Dengue Fever among Pediatric and Adult In-Patients in Metro Manila, Philippines.

BACKGROUND: Knowledge, attitude, and practice (KAP) of in-patients with dengue fever (DF) through hospital-based surveillance has not been done. This study aimed to assess and compare the KAP, identify its predictors, correlation, and protective factors among pediatric and adult patients with DF and community-based controls to structure proactive community-wide DF prevention and control programs. METHODS: This case-control study involved clinically or serologically confirmed patients (pediatrics n = 233; adults n = 17) with DF admitted in three public hospitals and community-based controls in Metro Manila, Philippines. A pretested structured KAP questionnaire was administered to participants to assess their KAP. RESULTS: Pediatric and adult patients had significantly lower mean scores in the practice (p < 0.001) domain compared with the pediatric and adult controls. Being in senior high school, having had days in hospital, and rash were predictors of KAP among pediatric patients. Knowledge and attitude of patients with DF did not correlate with their practices against DF. Use of mosquito-eating fish, screen windows, and dengue vaccine were protective factors against DF. CONCLUSION: The study highlights the importance of behavioral change for knowledge and attitude to have significant effect to practices against DF. Thus, we recommend two comprehensive health programs, Communication for Behavioral Impact (COMBI) and Health Belief Model (HBM).

A paired comparison of tuberculin skin test results in children with clinical manifestations of tuberculosis using 2 TU and 5 TU tuberculins

Rationale: A paired comparison of reactivity to purified protein derivative 2 TU PPD RT-23 and 5 TU PPD-S in children with clinical manifestations of tuberculosis was done to assess if 2 TU PPD RT-23 can be used instead of 5 TU PPD-S in routine Mantoux testing in the country. Objective: To determine the correlation of skin test reactivity to 2 TU PPD RT-23 and 5 TU PPD-S. Study Design: Cross Sectional Study. Methods: Two simultaneous skin tests using 2 TU PPD RT-23 and 5 TU PPD-S were performed. Each dose was randomly assigned in a blinded manner to the right or left forearm and read after 72 hours. Correlation between the size of induration obtained with 2 TU PPD RT-23 and with 5 TU PPD-S was done, as well as, correlation between tuberculin reactivity and age, gender, nutritional status, presence of BCG vaccination, exposure, and clinical manifestations. A p-value <0.05 was considered statistically significant. Results: Sixty five patients were included in the study. The overall mean difference in paired reaction sizes for the two doses was-1.02 + 2.8 mm (range of -11 to 3 mm). Using the present guidelines in the country to determine a positive tuberculin skin test, 27 (41.5 %) patients were positive when tested with 2 TU PPD RT-23 and 33 (50.8 %) patients were positive when tested with 5 TU PPD. The mean PPD size with 2 TU was 4.7 mm + 6.1 mm compared to 5.8 mm + 6.1 mm with 5 TU. PPD skin test reactivity with the two reagents was highly correlated (intraclass correlation 0.88; 95% CI 0.83-0.94). There was no significant association between age, gender, nutritional status, presence of BCG vaccination, TB exposure, and clinical manifestations to tuberculin reactivity. Conclusion: Tuberculin skin test reactivity among children, who were with clinical manifestations of tuberculosis and tested with 2 TU PPD RT-23 and 5 TU PPD-S, were found to be comparable. Age, gender, nutritional status, presence of BCG vaccination, TB exposure, and clinical manifestations were not factors influencing the size of the PPD reaction. 2 TU PPD RT-23 can be used instead of 5 TU PPD-S in routine Mantoux testing.

Constrictive pericarditis versus restrictive cardiomyopathy: the role of Doppler echocardiography in differential diagnosis.

Doppler ultrasound recordings of velocities of flow across the mitral and tricuspid valves and in the hepatic veins, and their variation with respiration, were recorded in seven patients with constrictive pericarditis and in six patients with restrictive cardiomyopathy. Deceleration of mitral and tricuspid flow was also evaluated during apnea. Color flow Doppler was performed in order to evaluate mitral and tricuspid regurgitation. Eight healthy adults served as controls. The patients with constrictive pericarditis showed higher peak diastolic velocities of mitral flow, as well as marked increase of velocity of flow at the onset of expiration and decrease at the onset of inspiration. Reciprocal respiratory variation of the velocities were also observed across the tricuspid valve. The patients with restrictive cardiomyopathy showed moderate or severe mitral and tricuspid regurgitation. They also showed shorter deceleration of flow across the mitral and tricuspid valves during apnea. The pattern of flow in the hepatic veins showed reversal during systole with accentuated reversion during inspiration. These results suggest that patient with constrictive pericarditis and restrictive cardiomyopathy can be differentiated by comparing Doppler echocardiographic data, along with changes induced by respiration.

[Analysis of the performance of several technicians in the echographic evaluation of the uterus]. / Analisi del comportamento di diversi operatori nella valutazione ecografica dell'utero.

The reproducibility degree in echographic biometry of uterus, studied by three diameters (longitudinal, transverse and antero-posterior), is mainly influenced by the anatomic characteristics of the structures. Starting from this observation a study about if and how much the understanding difficulties can influence differently the evaluation by four operators has been performed. Data, obtained from ten women, have been elaborated for appraising precision and accuracy of the four operators. Results show an almost univocal interpretation of the images and also that the data inhomogeneity in the less reproducible diameters valuation is caused by real difficulty in the interpretation of the pictures.