Pragmatic approach to nutrition in the ICU: Expert opinion regarding which calorie protein target.

BACKGROUND & AIMS: Since the publications of the ESPEN guidelines on enteral and parenteral nutrition in ICU, numerous studies have added information to assist the nutritional management of critically ill patients regarding the recognition of the right population to feed, the energy-protein targeting, the route and the timing to start. METHODS: We reviewed and discussed the literature related to nutrition in the ICU from 2006 until October 2013. RESULTS: To identify safe, minimal and maximal amounts for the different nutrients and at the different stages of the acute illness is necessary. These amounts might be specific for different phases in the time course of the patient's illness. The best approach is to target the energy goal defined by indirect calorimetry. High protein intake (1.5 g/kg/d) is recommended during the early phase of the ICU stay, regardless of the simultaneous calorie intake. This recommendation can reduce catabolism. Later on, high protein intake remains recommended, likely combined with a sufficient amount of energy to avoid proteolysis. CONCLUSIONS: Pragmatic recommendations are proposed to practically optimize nutritional therapy based on recent publications. However, on some issues, there is insufficient evidence to make expert recommendations.

Rapid diagnostic tests for non-malarial febrile illness in the tropics.

The recent roll-out of rapid diagnostic tests (RDTs) for malaria has highlighted the decreasing proportion of malaria-attributable illness in endemic areas. Unfortunately, once malaria is excluded, there are few accessible diagnostic tools to guide the management of severe febrile illnesses in low resource settings. This review summarizes the current state of RDT development for several key infections, including dengue fever, enteric fever, leptospirosis, brucellosis, visceral leishmaniasis and human African trypanosomiasis, and highlights many remaining gaps. Most RDTs for non-malarial tropical infections currently rely on the detection of host antibodies against a single infectious agent. The sensitivity and specificity of host-antibody detection tests are both inherently limited. Moreover, prolonged antibody responses to many infections preclude the use of most serological RDTs for monitoring response to treatment and/or for diagnosing relapse. Considering these limitations, there is a pressing need for sensitive pathogen-detection-based RDTs, as have been successfully developed for malaria and dengue. Ultimately, integration of RDTs into a validated syndromic approach to tropical fevers is urgently needed. Related research priorities are to define the evolving epidemiology of fever in the tropics, and to determine how combinations of RDTs could be best used to improve the management of severe and treatable infections requiring specific therapy.

A low stringency polymerase chain reaction approach to the identification of Biomphalaria glabrata and B. tenagophila, intermediate snail hosts of Schistosoma mansoni in Brazil

The low stringency-polymerase chain reaction (LS-PCR) with a pair of specific primers for the amplification of the 18S rRNA gene was evaluated as a means of differentiating between the two Schistosoma mansoni intermediate host species in Brazil: Biomphalaria glabrata and B. tenagophila. Individual snails obtained from different states of Brazil were used and the amplification patterns obtained showed a high degree of genetic variability in these species. Nevertheless, 4 and 3 clearly defined specific diagnostic bands was observed in individuals from B. glabrata and B. tenagophila respectively. The detection of snail specific diagnostic bands suggests the possibility of reliable species differentiation at the DNA level using LS-PCR.

High throughput sequencing and proteomics to identify immunogenic proteins of a new pathogen: the dirty genome approach.

With the availability of new generation sequencing technologies, bacterial genome projects have undergone a major boost. Still, chromosome completion needs a costly and time-consuming gap closure, especially when containing highly repetitive elements. However, incomplete genome data may be sufficiently informative to derive the pursued information. For emerging pathogens, i.e. newly identified pathogens, lack of release of genome data during gap closure stage is clearly medically counterproductive. We thus investigated the feasibility of a dirty genome approach, i.e. the release of unfinished genome sequences to develop serological diagnostic tools. We showed that almost the whole genome sequence of the emerging pathogen Parachlamydia acanthamoebae was retrieved even with relatively short reads from Genome Sequencer 20 and Solexa. The bacterial proteome was analyzed to select immunogenic proteins, which were then expressed and used to elaborate the first steps of an ELISA. This work constitutes the proof of principle for a dirty genome approach, i.e. the use of unfinished genome sequences of pathogenic bacteria, coupled with proteomics to rapidly identify new immunogenic proteins useful to develop in the future specific diagnostic tests such as ELISA, immunohistochemistry and direct antigen detection. Although applied here to an emerging pathogen, this combined dirty genome sequencing/proteomic approach may be used for any pathogen for which better diagnostics are needed. These genome sequences may also be very useful to develop DNA based diagnostic tests. All these diagnostic tools will allow further evaluations of the pathogenic potential of this obligate intracellular bacterium.

The transventricular-transseptal access to the aortic root: a new route for extrapleural trans-catheter aortic stent-valve implantation.

Objective: The aim of this study was to investigate the feasibility of transventricular-transseptal approach (TVSA) for extrapleural transcatheter aortic valved stent implantation via a subxyphoidian access. Methods: In five porcine experiments (52.3 +/- 10.9 kg) the right ventricle was exposed via subxyphoidian access. Under the guidance of intracardiac echocardiography (ICE) and fluoroscopy, the transseptal access from right ventricle to left ventricle was created progressively by puncture and dilation with dilators (8F-26F). Valved stents built in-house from commercial tanned pericardium and self-expandable Nitinol stents were loaded into a cartridge. A delivery sheath was then introduced from the right ventricle into the left ventricle and then into the ascending aorta. The cartridge was connected and the valved stent was deployed in the aortic position. Then, the ventricular septal access was sealed with an Amplatzer septal occluder device and the right ventricular access was closed by tying prepared purse-string suture directly. Thirty minutes after the whole procedure, the animals were sacrificed for macroscopic evaluation of the position of valved stent and septal closure device. Result: Procedural success of TVSA was 100% at the first attempt. Mean procedure time was 49 +/- 4 min. Progressive dilatation of the transseptal access resulted in a measurable ventricular septal defect (VSD) after dilator sizes 18F and more. All valved stents were delivered at the target site over the native aortic valve with good acute valve function and no paravalvular leaks. During the procedure, premature beats (5/5) and supraventriclar tachycardias (5/5) were observed, but no atrial-ventricular block (0/5) occurred. Heart rate before (after) was 90 +/- 3 beats min(-1) (100 +/- 2 beats min(-1): p < 0.05), whereas blood pressure was 60 + 1 mm Hg (55 + 2 mm Hg (p < 0.05)). Total blood loss was 280 + 10 ml. The Amplatzer septal occluder devices were fully deployed and the ventricular septal accesses were sealed successfully, without detectable residual shunt. Conclusion: Trans-catheter implantation of aortic valved stent via extrapleural transventricular-transseptal access is technically feasible and has the potential for a simplified procedure under local anaesthesia. (C) 2010 European Association for Cardio-Thoracic Surgery. Published by Elsevier B. V. All rights reserved.

A combined approach for the assessment of cell viability and cell functionality of human fibrochondrocytes for use in tissue engineering.

Temporo-mandibular joint disc disorders are highly prevalent in adult populations. Autologous chondrocyte implantation is a well-established method for the treatment of several chondral defects. However, very few studies have been carried out using human fibrous chondrocytes from the temporo-mandibular joint (TMJ). One of the main drawbacks associated to chondrocyte cell culture is the possibility that chondrocyte cells kept in culture tend to de-differentiate and to lose cell viability under in in-vitro conditions. In this work, we have isolated human temporo-mandibular joint fibrochondrocytes (TMJF) from human disc and we have used a highly-sensitive technique to determine cell viability, cell proliferation and gene expression of nine consecutive cell passages to determine the most appropriate cell passage for use in tissue engineering and future clinical use. Our results revealed that the most potentially viable and functional cell passages were P5-P6, in which an adequate equilibrium between cell viability and the capability to synthesize all major extracellular matrix components exists. The combined action of pro-apoptotic (TRAF5, PHLDA1) and anti-apoptotic genes (SON, HTT, FAIM2) may explain the differential cell viability levels that we found in this study. These results suggest that TMJF should be used at P5-P6 for cell therapy protocols.

Application of Molecular Diagnostic Techniques for Viral Testing.

Nucleic acid amplification techniques are commonly used currently to diagnose viral diseases and manage patients with this kind of illnesses. These techniques have had a rapid but unconventional route of development during the last 30 years, with the discovery and introduction of several assays in clinical diagnosis. The increase in the number of commercially available methods has facilitated the use of this technology in the majority of laboratories worldwide. This technology has reduced the use of some other techniques such as viral culture based methods and serological assays in the clinical virology laboratory. Moreover, nucleic acid amplification techniques are now the methods of reference and also the most useful assays for the diagnosis in several diseases. The introduction of these techniques and their automation provides new opportunities for the clinical laboratory to affect patient care. The main objectives in performing nucleic acid tests in this field are to provide timely results useful for high-quality patient care at a reasonable cost, because rapid results are associated with improvements in patients care. The use of amplification techniques such as polymerase chain reaction, real-time polymerase chain reaction or nucleic acid sequence-based amplification for virus detection, genotyping and quantification have some advantages like high sensitivity and reproducibility, as well as a broad dynamic range. This review is an up-to-date of the main nucleic acid techniques and their clinical applications, and special challenges and opportunities that these techniques currently provide for the clinical virology laboratory.

Perioperative anemia management in colorectal cancer patients: A pragmatic approach.

Anemia, usually due to iron deficiency, is highly prevalent among patients with colorectal cancer. Inflammatory cytokines lead to iron restricted erythropoiesis further decreasing iron availability and impairing iron utilization. Preoperative anemia predicts for decreased survival. Allogeneic blood transfusion is widely used to correct anemia and is associated with poorer surgical outcomes, increased post-operative nosocomial infections, longer hospital stays, increased rates of cancer recurrence and perioperative venous thromboembolism. Infections are more likely to occur in those with low preoperative serum ferritin level compared to those with normal levels. A multidisciplinary, multimodal, individualized strategy, collectively termed Patient Blood Management, minimizes or eliminates allogeneic blood transfusion. This includes restrictive transfusion policy, thromboprophylaxis and anemia management to improve outcomes. Normalization of preoperative hemoglobin levels is a World Health Organization recommendation. Iron repletion should be routinely ordered when indicated. Oral iron is poorly tolerated with low adherence based on published evidence. Intravenous iron is safe and effective but is frequently avoided due to misinformation and misinterpretation concerning the incidence and clinical nature of minor infusion reactions. Serious adverse events with intravenous iron are extremely rare. Newer formulations allow complete replacement dosing in 15-60 min markedly facilitating care. Erythropoiesis stimulating agents may improve response rates. A multidisciplinary, multimodal, individualized strategy, collectively termed Patient Blood Management used to minimize or eliminate allogeneic blood transfusion is indicated to improve outcomes.

Identification of factors associated with diagnostic error in primary care.

BACKGROUND Missed, delayed or incorrect diagnoses are considered to be diagnostic errors. The aim of this paper is to describe the methodology of a study to analyse cognitive aspects of the process by which primary care (PC) physicians diagnose dyspnoea. It examines the possible links between the use of heuristics, suboptimal cognitive acts and diagnostic errors, using Reason's taxonomy of human error (slips, lapses, mistakes and violations). The influence of situational factors (professional experience, perceived overwork and fatigue) is also analysed. METHODS Cohort study of new episodes of dyspnoea in patients receiving care from family physicians and residents at PC centres in Granada (Spain). With an initial expected diagnostic error rate of 20%, and a sampling error of 3%, 384 episodes of dyspnoea are calculated to be required. In addition to filling out the electronic medical record of the patients attended, each physician fills out 2 specially designed questionnaires about the diagnostic process performed in each case of dyspnoea. The first questionnaire includes questions on the physician's initial diagnostic impression, the 3 most likely diagnoses (in order of likelihood), and the diagnosis reached after the initial medical history and physical examination. It also includes items on the physicians' perceived overwork and fatigue during patient care. The second questionnaire records the confirmed diagnosis once it is reached. The complete diagnostic process is peer-reviewed to identify and classify the diagnostic errors. The possible use of heuristics of representativeness, availability, and anchoring and adjustment in each diagnostic process is also analysed. Each audit is reviewed with the physician responsible for the diagnostic process. Finally, logistic regression models are used to determine if there are differences in the diagnostic error variables based on the heuristics identified. DISCUSSION This work sets out a new approach to studying the diagnostic decision-making process in PC, taking advantage of new technologies which allow immediate recording of the decision-making process.

Colorectal Cancer Classification and Cell Heterogeneity: A Systems Oncology Approach.

Colorectal cancer is a heterogeneous disease that manifests through diverse clinical scenarios. During many years, our knowledge about the variability of colorectal tumors was limited to the histopathological analysis from which generic classifications associated with different clinical expectations are derived. However, currently we are beginning to understand that under the intense pathological and clinical variability of these tumors there underlies strong genetic and biological heterogeneity. Thus, with the increasing available information of inter-tumor and intra-tumor heterogeneity, the classical pathological approach is being displaced in favor of novel molecular classifications. In the present article, we summarize the most relevant proposals of molecular classifications obtained from the analysis of colorectal tumors using powerful high throughput techniques and devices. We also discuss the role that cancer systems biology may play in the integration and interpretation of the high amount of data generated and the challenges to be addressed in the future development of precision oncology. In addition, we review the current state of implementation of these novel tools in the pathological laboratory and in clinical practice.

An Interval Intelligent-based Approach for Fault Detection and Modelling

Not considered in the analytical model of the plant, uncertainties always dramatically decrease the performance of the fault detection task in the practice. To cope better with this prevalent problem, in this paper we develop a methodology using Modal Interval Analysis which takes into account those uncertainties in the plant model. A fault detection method is developed based on this model which is quite robust to uncertainty and results in no false alarm. As soon as a fault is detected, an ANFIS model is trained in online to capture the major behavior of the occurred fault which can be used for fault accommodation. The simulation results understandably demonstrate the capability of the proposed method for accomplishing both tasks appropriately

Le trouble de l'adaptation, un diagnostic anodin ? : une étude comparative avec le trouble dépressif majeur

Résumé Le trouble de l'adaptation est un diagnostic très fréquent, mais étonnamment peu étudié et controversé. Il est, selon les études, considéré comme une forme mineure d'un trouble psychiatrique spécifique, comme une fragilité psychologique révélée par un événement stressant pour le sujet ou encore comme une forme précoce annonçant un trouble psychiatrique majeur. Ces trois points de vue ramènent en fait tous à la question de fond concernant son étiologie. L'objectif de cette étude est de montrer si le trouble de l'adaptation est un diagnostic clairement différencié dont l'existence est justifiée. Afin de tenter de répondre à cette question, il nous est apparu intéressant de comparer cette catégorie diagnostique à une autre catégorie diagnostique psychiatrique importante, le trouble dépressif majeur. Dans cette étude rétrospective nous avons sélectionné tous les patients avec un diagnostic de trouble de l'adaptation ou un trouble dépressif majeur parmi les patients hospitalisés à l'hôpital psychiatrique de Malévoz en Valais en 1993 (580). Elle est basée sur des diagnostics cliniques. Nous avons comparé leurs données socio-démographiques (âge, sexe, nationalité, état civil, activité professionnelle), leurs antécédents psychiatriques (hospitalisations antérieures, suivi psychiatrique ambulatoire, antécédents de tentamen), leurs hospitalisations ultérieures dans les 5 ans, leur hospitalisation actuelle (durée, tentamens, comorbidité) et les traitements médicamenteux prescrits (leur nombre et leur classe). Notre étude met en évidence certaines distinctions entre le trouble de l'adaptation et le trouble dépressif majeur: les patients souffrant de trouble de l'adaptation diffèrent des troubles dépressifs majeurs par le fait qu'ils sont plus fréquemment des hommes, célibataires et plus jeunes que ceux souffrant de trouble dépressif majeur; leur durée d'hospitalisation est plus courte, leur évolution entre les hospitalisations est meilleure et ils reçoivent moins de psychotropes. Nous ne pouvons cependant pas conclure à une distinction claire de ces deux catégories diagnostiques, ni que le trouble de l'adaptation n'est pas simplement lié à une moindre gravité. Nos résultats confirment par contre que ce diagnostic n'est pas non plus un diagnostic anodin (nombre élevé d'antécédents psychiatriques, de tentamens, d'hospitalisations psychiatriques ultérieures, importance des comorbidités de même que la lourdeur des traitements psychotropes prescrits (notamment la fréquence des neuroleptiques). A notre avis, les trois hypothèses étiologiques (forme mineure, trouble précoce ou fragilité psychologique spécifique révélée par un événement stressant) qui ont été évoquées peuvent être considérées comme plausibles suivant le point de vue que l'on choisit. Le diagnostic de trouble de l'adaptation révèle une des limitations de l'approche du DSM-Ill-R qui se veut athéorique. Le fait que dans sa définition même, le DSM-111-R évoque "qu'il faut souvent se référer au seul jugement clinique" le montre bien, un tel diagnostic renvoie inévitablement à une référence psychopathologique. Nous pensons qu'il est illusoire de vouloir se passer d'une telle référence qui elle seule permet d'appréhender justement la portée symbolique d'un événement donné pour un individu. Summary In this retrospective study we selected all the patients with a diagnosis of adjustment disorder (77) or major depressive disorder (125) among the patients hospitalised in the psychiatric hospital of Malevoz in Valais during the year 1993 (580). It is based on clinical diagnosis. Their social and demographic characteristics (age, sex, nationality, marital status, professional activity), their past psychiatric history (earlier psychiatric hospitalisations, ambulatory treatment and attempted suicide), their hospitalisations during the next 5 years, their index hospitalisation (length, attempted suicide, comorbidity) and their drug treatment (number and class of prescribed drugs) were compared. This survey confirms certain differences be-tween adjustment disorder and major depression disorder: patients suffering from adjustment disorder were more often men, not married, younger than those suffering from major depression; their hospitalisations were shorter with a better evolution between hospitalisations and they received less medication. However, the study does not allow to clearly distinguish between the two diagnoses or to conclude that adjustment disorder is not only a minor form of a specific psychiatric disorder. Yet it confirms that adjustment disorder is not a light diagnosis (importance of the psychiatric past, high number of past attempted suicides, rehospitalisations, number of comorbid disorders and weight of the prescribed psychotropic treatments among which neuroleptics were frequent). The three aetiological hypotheses that have been proposed (minor form of a specific disorder, specific psychological vulnerability revealed by a stress factor or precursor manifestation of a major psychiatric disorder) can still be considered as plausible. The diagnosis of adjustment disorder points to methodological limitations of the atheoretical approach of the DSM-III-R. The fact that, in its DSM-III-R definition, it is stated that the diagnosis of adjustment disorder has often to be based only on clinical judgment shows very well that such a diagnosis inevitably refers to a psychopathological theory. Indeed, the authors consider an approach without such a reference as difficult, a reference which remains the only way to appreciate accurately the symbolic weight of a given event for an individual person.

Simultaneous detection of aneuploidies for chromosomes 4, 6, 10 and 17 by automated four colour I-FISH in high hyperdiploid acute lymphoblastic leukemia : diagnostic assessment, clonal heterogeneity and chromosomal instability in adults

SummarySimultaneous detection of aneuploidies for chromosomes 4, 6,10 and 17 by automated four color l-FISH in high hyperdiploid acute lymphoblastic leukemia: diagnostic assessment, clonal heterogeneity and chromosomal instability in adultsAnna Talamo BlandinService de Génétique Médicale, Unité de Cytogénétique du Cancer, CHUVAcute lymphoblastic leukemia (ALL) is a malignant hemopathy characterized by the accumulation of the immature lymphoid cells in the bone marrow and, most often, in the peripheral blood. ALL is a heterogeneous disease with distinct biological and prognostic entities. At diagnosis, cytogenetic and molecular findings constitute important and independent prognostic factors. High hyperdiploidy with 51-67 chromosomes (HeH), one of the largest cytogenetic subsets of ALL, in childhood particularly, is generally associated with a relatively favorable outcome. Chromosome gain is nonrandom, extracopies of some chromosome occurring more frequently than those of others. Concurrent presence of trisomy for chromosomes 4, 10 and 17 confers an especially good prognosis. The first aim of our work was to develop an automated four color interphase fluorescence in situ hybridization (l-FISH) methodology and to assess its ability to detect concurrent aneuploidies 4, 6, 10 and 17 in 10 ALL patients. Various combinations of aneuploidies were identified. All clones detected by conventional cytogenetics were also observed by l-FISH. However, in all patients, l-FISH revealed numerous additional abnormal clones, leading to a high level of clonal heterogeneity. Our second aim has been to investigate the nature and origin of this clonal heterogeneity and to test for the presence of chromosome instability (CIN) in HeH ALL at initial presentation. Ten HeH ALL and 10 non-HeH ALL patients were analysed by four colour l-FISH and numerical CIN values were determined for all four chromosomes together and for each chromosome and patient group, an original approach in ALL. CIN values in HeH ALL proved to be much higher than#iose in non-HeH ALL, suggesting that numerical CIN may be at the origin of the high level of clonal heterogeneity revealed by l-FISH. Our third aim has been to study the evolution of these cytogenetic features during the course of the disease in 10 HeH ALL patients. Clonal heterogeneity was also observed again during disease progression, particularly at relapse. Clones detected at initial presentation generally reappeared in relapse, in most cases with newly generated ones. A significant correlation between the number of abnormal clones and CIN suggested that the higher the instability, the larger the number of abnormal clones. Whereas clonal heterogeneity and its evolution most probably result from underlying chromosome instability, operating processes remain conjectural.RésuméLa leucémie lymphoblastique aiguë (LLA) est une hémopathie maligne qui résulte de l'accumulationde cellules lymphoïdes immatures dans la moelle osseuse, et, le plus souvent, dans le sangpériphérique également. La LLA est une affection hétérogène au sein de laquelle se distinguentplusieurs entités biologiques et pronostiques. Les données cytogénétiques et moléculaires font partieintégrante du diagnostic et jouent un rôle essentiel dans l'évaluation du pronostic. L'hyperdiploïdieélevée à 51-­67 chromosomes (HeH), relativement fréquente, en particulier chez l'enfant, s'associe àun pronostic favorable. Le gain de chromosomes ne relève pas du hasard, certains chromosomesétant plus fréquemment impliqués que d'autres. La présence simultanée des trisomies 4, 6, et 17s'associe à un pronostic particulièrement bon. Le premier but du travail a été de développer uneméthode d'analyse automatique par hybridation in situ fluorescente interphasique (I-­FISH) à 4couleurs et de tester sa capacité à identifier la présence simultanée d'aneuploïdies 4, 6, 10 et 17 dans10 cas de LLA. Différentes combinaisons d'aneuploïdies ont été identifiées. Tous les clones détectéspar cytogénétique conventionnelle l'ont été par I-­FISH. Or, chez tous les patients, l'I-­FISH a révélé denombreux clones anormaux additionnels générant un degré élevé d'hétérogénéité clonale. Notredeuxième but a été d'investiguer la nature et l'origine de cette hétérogénéité et de tester la présenced'instabilité chromosomique (CIN) chez les patients avec une LLA HeH en presentation initiale. DixLLA HeH et 10 LLA non-­HeH ont été analysées par I-­FISH et les valeurs de CIN numérique ont étédéterminées pour les 4 chromosomes ensemble et pour chaque chromosome et groupe de patients,approche originale dans la LLA. Ces valeurs étant beaucoup plus élevées dans la LLA HeH que dansla LLA non-­HeH, elles favorisent l'hypothèse selon laquelle la CIN serait à l'origine de l'hétérogénéitéclonale révélée par I-­FISH. Le troisième but de notre travail a été d'étudier l'évolution de cescaractéristiques cytogénétiques au cours de la maladie dans 10 cas de LLA HeH. L'hétérogénéitéclonale a été retrouvée lors de la progression de la maladie, en particulier en rechute, où les clonesanormaux détectés en présentation initiale réapparaissent, généralement accompagnés de clonesnouveaux. La corrélation existant entre nombre de clones anormaux et valeurs de CIN suggère queplus l'instabilité est élevée, plus le nombre de clones anormaux est grand. Bien que l'hétérogénéitéclonale et son évolution résultent très probablement de l'instabilité chromosomique, les processus àl'oeuvre ne sont pas entièrement élucidés.

Radioimmuno positron emission tomography with monoclonal antibodies: a new approach to quantifying in vivo tumour concentration and biodistribution for radioimmunotherapy.

Radioimmunodetection of tumours with monoclonal antibodies is becoming an established procedure. Positron emission tomography (PET) shows better resolution than normal gamma camera single photon emission tomography and can provide more precise quantitative data. Thus, in the present study, these powerful methods have been combined to perform radioimmuno PET (RI-PET). Monoclonal antibodies directed against carcinoembryonic antigen (CEA) an IgG, its F(ab')2 and a mouse-human chimeric IgG derived from it were labelled with 124I, a positron-emitting radionuclide with a convenient physical half-life of four days. Mice, xenografted with a CEA-producing human colon carcinoma, were injected with the 124I-MAb and the tumours were visualized using PET. The concentrations of 124I in tumour and normal tissue were determined by both PET and direct radioactivity counting of the dissected animals, with very good agreement. To allow PET quantification, a procedure was established to account for the presence of radioactivity during the absorption correction measurement (transmission scan). Comparison of PET and tissue counting indicates that this novel combination of radioimmunolocalization and PET (RI-PET) will provide, in addition to more precise diagnosis, more accurate radiation dosimetry for radioimmunotherapy.

Improved HIV-1 RNA quantitation by COBAS AmpliPrep/COBAS TaqMan HIV-1 Test, v2.0 using a novel dual-target approach.

BACKGROUND: HIV-1 RNA viral load is a key parameter for reliable treatment monitoring of HIV-1 infection. Accurate HIV-1 RNA quantitation can be impaired by primer and probe sequence polymorphisms as a result of tremendous genetic diversity and ongoing evolution of HIV-1. A novel dual HIV-1 target amplification approach was realized in the quantitative COBAS AmpliPrep/COBAS TaqMan HIV-1 Test, v2.0 (HIV-1 TaqMan test v2.0) to cope with the high genetic diversity of the virus. OBJECTIVES AND STUDY DESIGN: The performance of the new assay was evaluated for sensitivity, dynamic range, precision, subtype inclusivity, diagnostic and analytical specificity, interfering substances, and correlation with the COBAS AmpliPrep/COBAS TaqMan HIV-1 (HIV-1 TaqMan test v1.0) predecessor test in patients specimens. RESULTS: The new assay demonstrated a sensitivity of 20 copies/mL, a linear measuring range of 20-10,000,000 copies/mL, with a lower limit of quantitation of 20 copies/mL. HIV-1 Group M subtypes and HIV-1 Group O were quantified within +/-0.3 log(10) of the assigned titers. Specificity was 100% in 660 tested specimens, no cross reactivity was found for 15 pathogens nor any interference for endogenous substances or 29 drugs. Good comparability with the predecessor assay was demonstrated in 82 positive patient samples. In selected clinical samples 35/66 specimens were found underquantitated in the predecessor assay; all were quantitated correctly in the new assay. CONCLUSIONS: The dual-target approach for the HIV-1 TaqMan test v2.0 enables superior HIV-1 Group M subtype coverage including HIV-1 Group O detection. Correct quantitation of specimens underquantitated in the HIV-1 TaqMan test v1.0 test was demonstrated.