PREPARATION AND CHARACTERIZATION OF COVALENTLY LINKED ENZYME-ANTIBODY CONJUGATES AND THEIR USE IN MEASURING MINUTE QUANTITIES OF PROTEIN ANTIGENS

The discovery and characterization of oncofetal proteins have led to significant advances in early cancer diagnosis and therapeutic monitoring of patients undergoing cancer chemotherapy. These tumor-associated antigens are presently measured by sensitive, specific immunoassay techniques based on the detection of minute amounts of labeled antigen or antibody incorporated into immune complexes, which must be isolated from free antigen and antibody.^ Since there are several disadvantages with using radioisotopes, the most common immunolabel, one major objective was to prepare covalently coupled enzyme-antibody conjugates and evaluate their use as a practical alternative to radiolabeled immune reagents. An improved technique for the production of enzyme-antibody conjugates was developed that involves oxidizing the carbohydrate moieties on a glycoprotein enzyme, then introducing antibody in the presence of polyethylene glycol (PEG). Covalent enzyme-antibody conjugates involving alkaline phosphatase and amyloglucosidase were produced and characterized.^ In order to increase the sensitivity of detecting the amyloglucosidase-antibody conjugate, an enzyme cycling assay was developed that measures glucose, the product of maltose cleavage by amyloglucosidase, in the picomole range. The increased sensitivity obtained by combined usage of the amyloglucosidase-antibody conjugate and enzyme cycling assay was then compared to that of conventional enzyme immunoassay (EIA).^ For immune complex isolation, polystyrene tubes and protein A-bearing Staphylococcus aureus were evaluated as solid phase matrices, upon which antibodies can be immobilized. A sandwich-type EIA, using antibody-coated S. aureus, was developed that measures human albumin (HSA) in the nanogram range. The assay, using an alkaline phosphatase-anti-HSA conjugate, was applied to the determination of HSA in human urine and evaluated extensively for its clinical applicability.^ Finally, in view of the clinical significance of alpha-fetoprotein (AFP) as an oncofetal antigen and the difficulty with its purification for use as an immunogen and assay standard, a chemical purification protocol was developed that resulted in a high yield of immunochemically pure AFP. ^

Practical scores for the detection of malnutrition

Malnutrition occurs in 30 - 60 % of hospitalized medical or surgical patients, as well as out-patients. Serious consequences at various levels were observed. Malnutrition influences negatively the quality of life, the immune system, muscle strength and worsens the prognosis of the patient. Interventions for a rapid and simple identification and effective treatment of this condition are essential and cost saving. Screening tools for the identification of patients at nutritional risk are very useful in daily practice. The systematic identification of patients with potential or apparent malnutrition is very important allowing an effective nutritional treatment at an early time. The medical team in charge should perform the nutritional risk screening and the following assessment to recognize the nutritional problems and to solve them in an interdisciplinary and -professional team.

A practical approach to designing syndromic surveillance systems for livestock and poultry

The field of animal syndromic surveillance (SyS) is growing, with many systems being developed worldwide. Now is an appropriate time to share ideas and lessons learned from early SyS design and implementation. Based on our practical experience in animal health SyS, with additions from the public health and animal health SyS literature, we put forward for discussion a 6-step approach to designing SyS systems for livestock and poultry. The first step is to formalise policy and surveillance goals which are considerate of stakeholder expectations and reflect priority issues (1). Next, it is important to find consensus on national priority diseases and identify current surveillance gaps. The geographic, demographic, and temporal coverage of the system must be carefully assessed (2). A minimum dataset for SyS that includes the essential data to achieve all surveillance objectives while minimizing the amount of data collected should be defined. One can then compile an inventory of the data sources available and evaluate each using the criteria developed (3). A list of syndromes should then be produced for all data sources. Cases can be classified into syndrome classes and the data can be converted into time series (4). Based on the characteristics of the syndrome-time series, the length of historic data available and the type of outbreaks the system must detect, different aberration detection algorithms can be tested (5). Finally, it is essential to develop a minimally acceptable response protocol for each statistical signal produced (6). Important outcomes of this pre-operational phase should be building of a national network of experts and collective action and evaluation plans. While some of the more applied steps (4 and 5) are currently receiving consideration, more emphasis should be put on earlier conceptual steps by decision makers and surveillance developers (1-3).

The applicability of using different energy levels in CT imaging for differentiation or identification of dental restorative materials.

PURPOSE The goal of this study was to investigate whether different computed tomography (CT) energy levels could supply additional information for the differentiation of dental materials for forensic investigations. METHODS Nine different commonly used restorative dental materials were investigated in this study. A total of 75 human third molars were filled with the restorative dental materials and then scanned using the forensic reference phantom in singlesource mode. The mean Hounsfield unit values and standard deviations (SDs) of each material were calculated at 120, 80 and 140 kVp. RESULTS Most of the dental materials could be differentiated at 120 kVp. We found that greater X-ray density of a material resulted in higher SDs and that the material volume could influence the measurements. CONCLUSION Differentiation of dental materials in CT was possible in many cases using single-energy CT scans at 120 kVp. Because of the number of dental restorative materials available and scanner and scan parameter dependence, as well as the CT imaging artifacts, the identification (in contrast to differentiation) was problematic.

Can 3D ultrasound identify trochlea dysplasia in newborns? Evaluation and applicability of a technique.

Femoro-patellar dysplasia is considered as a significant risk factor of patellar instability. Different studies suggest that the shape of the trochlea is already developed in early childhood. Therefore early identification of a dysplastic configuration might be relevant information for the treating physician. An easy applicable routine screening of the trochlea is yet not available. The purpose of this study was to establish and evaluate a screening method for femoro-patellar dysplasia using 3D ultrasound. From 2012 to 2013 we prospectively imaged 160 consecutive femoro-patellar joints in 80 newborns from the 36th to 61st gestational week that underwent a routine hip sonography (Graf). All ultrasounds were performed by a pediatric radiologist with only minimal additional time to the routine hip ultrasound. In 30° flexion of the knee, axial, coronal, and sagittal reformats were used to standardize a reconstructed axial plane through the femoral condyle and the mid-patella. The sulcus angle, the lateral-to-medial facet ratio of the trochlea and the shape of the patella (Wiberg Classification) were evaluated. In all examinations reconstruction of the standardized axial plane was achieved, the mean trochlea angle was 149.1° (SD 4.9°), the lateral-to-medial facet ratio of the trochlea ratio was 1.3 (SD 0.22), and a Wiberg type I patella was found in 95% of the newborn. No statistical difference was detected between boys and girls. Using standardized reconstructions of the axial plane allows measurements to be made with lower operator dependency and higher accuracy in a short time. Therefore 3D ultrasound is an easy applicable and powerful tool to identify trochlea dysplasia in newborns and might be used for screening for trochlea dysplasia.

Predicting the onset of psychosis in patients at clinical high risk: practical guide to probabilistic prognostic reasoning

Prediction of psychosis in patients at clinical high risk (CHR) has become a mainstream focus of clinical and research interest worldwide. When using CHR instruments for clinical purposes, the predicted outcome is but only a probability; and, consequently, any therapeutic action following the assessment is based on probabilistic prognostic reasoning. Yet, probabilistic reasoning makes considerable demands on the clinicians. We provide here a scholarly practical guide summarising the key concepts to support clinicians with probabilistic prognostic reasoning in the CHR state. We review risk or cumulative incidence of psychosis in, person-time rate of psychosis, Kaplan-Meier estimates of psychosis risk, measures of prognostic accuracy, sensitivity and specificity in receiver operator characteristic curves, positive and negative predictive values, Bayes’ theorem, likelihood ratios, potentials and limits of real-life applications of prognostic probabilistic reasoning in the CHR state. Understanding basic measures used for prognostic probabilistic reasoning is a prerequisite for successfully implementing the early detection and prevention of psychosis in clinical practice. Future refinement of these measures for CHR patients may actually influence risk management, especially as regards initiating or withholding treatment.

Practical approach to the use of daylight photodynamic therapy with topical methyl aminolevulinate for actinic keratosis: a European consensus

INTRODUCTION Daylight-mediated photodynamic therapy has been shown to be an effective therapy for actinic keratoses (AKs) and a simple and tolerable treatment procedure in three randomized Scandinavian studies and two recent Phase III randomized controlled studies in Australia and Europe. OBJECTIVES To establish consensus recommendations for the use of daylight photodynamic therapy (DL-PDT) using topical methyl aminolaevulinate (MAL) in European patients with AKs. METHODS The DL-PDT consensus recommendations were developed on behalf of the European Society for Photodynamic Therapy in Dermatology and comprised of 10 dermatologists from different European countries with experience in how to treat AK patients with PDT. Consensus was developed based on literature review and experience of the experts in the treatment of AK using DL-PDT. RESULTS The recommendations arising from this panel of experts provide general guidance on the use of DL-PDT as a dermatological procedure with specific guidance regarding patient selection, therapeutic indications, when to treat, pre-treatment skin preparation, MAL application and daylight exposure for patients with AK in different countries of Europe. CONCLUSIONS This consensus recommendation provides a framework for physicians to perform DL-PDT with MAL cream while ensuring efficiency and safety in the treatment of patients with AK in different European countries.