CHD4 is recruited by GATA4 and NKX2-5 to repress noncardiac gene programs in the developing heart.

The nucleosome remodeling and deacetylase (NuRD) complex is one of the central chromatin remodeling complexes that mediates gene repression. NuRD is essential for numerous developmental events, including heart development. Clinical and genetic studies have provided direct evidence for the role of chromodomain helicase DNA-binding protein 4 (CHD4), the catalytic component of NuRD, in congenital heart disease (CHD), including atrial and ventricular septal defects. Furthermore, it has been demonstrated that CHD4 is essential for mammalian cardiomyocyte formation and function. A key unresolved question is how CHD4/NuRD is localized to specific cardiac target genes, as neither CHD4 nor NuRD can directly bind DNA. Here, we coupled a bioinformatics-based approach with mass spectrometry analyses to demonstrate that CHD4 interacts with the core cardiac transcription factors GATA4, NKX2-5, and TBX5 during embryonic heart development. Using transcriptomics and genome-wide occupancy data, we characterized the genomic landscape of GATA4, NKX2-5, and TBX5 repression and defined the direct cardiac gene targets of the GATA4-CHD4, NKX2-5-CHD4, and TBX5-CHD4 complexes. These data were used to identify putative cis-regulatory elements controlled by these complexes. We genetically interrogated two of these silencers in vivo: Acta1 and Myh11 We show that deletion of these silencers leads to inappropriate skeletal and smooth muscle gene misexpression, respectively, in the embryonic heart. These results delineate how CHD4/NuRD is localized to specific cardiac loci and explicates how mutations in the broadly expressed CHD4 protein lead to cardiac-specific disease states.

Missense Mutation in Human CHD4 Causes Ventricular Noncompaction by Repressing ADAMTS1.

BACKGROUND: Left ventricular noncompaction (LVNC) is a prevalent cardiomyopathy associated with excessive trabeculation and thin compact myocardium. Patients with LVNC are vulnerable to cardiac dysfunction and at high risk of sudden death. Although sporadic and inherited mutations in cardiac genes are implicated in LVNC, understanding of the mechanisms responsible for human LVNC is limited. METHODS: We screened the complete exome sequence database of the Pediatrics Cardiac Genomics Consortium and identified a cohort with a de novo CHD4 (chromodomain helicase DNA-binding protein 4) proband, CHD4M202I, with congenital heart defects. We engineered a humanized mouse model of CHD4M202I (mouse CHD4M195I). Histological analysis, immunohistochemistry, flow cytometry, transmission electron microscopy, and echocardiography were used to analyze cardiac anatomy and function. Ex vivo culture, immunopurification coupled with mass spectrometry, transcriptional profiling, and chromatin immunoprecipitation were performed to deduce the mechanism of CHD4M195I-mediated ventricular wall defects. RESULTS: CHD4M195I/M195I mice developed biventricular hypertrabeculation and noncompaction and died at birth. Proliferation of cardiomyocytes was significantly increased in CHD4M195I hearts, and the excessive trabeculation was associated with accumulation of ECM (extracellular matrix) proteins and a reduction of ADAMTS1 (ADAM metallopeptidase with thrombospondin type 1 motif 1), an ECM protease. We rescued the hyperproliferation and hypertrabeculation defects in CHD4M195I hearts by administration of ADAMTS1. Mechanistically, the CHD4M195I protein showed augmented affinity to endocardial BRG1 (SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily A, member 4). This enhanced affinity resulted in the failure of derepression of Adamts1 transcription such that ADAMTS1-mediated trabeculation termination was impaired. CONCLUSIONS: Our study reveals how a single mutation in the chromatin remodeler CHD4, in mice or humans, modulates ventricular chamber maturation and that cardiac defects associated with the missense mutation CHD4M195I can be attenuated by the administration of ADAMTS1.

Patterns of admixture and introgression in a mosaic Mytilus galloprovincialis and Mytilus edulis hybrid zone in SW England.

The study of hybrid zones offers important insights into speciation. Earlier studies on hybrid populations of the marine mussel species Mytilus edulis and Mytilus galloprovincialis in SW England provided evidence of admixture but were constrained by the limited number of molecular markers available. We use 57 ancestry-informative SNPs, most of which have been mapped genetically, to provide evidence of distinctive differences between admixed populations in SW England and asymmetrical introgression from M. edulis to M. galloprovincialis. We combine the genetic study with analysis of phenotypic traits of potential ecological and adaptive significance. We demonstrate that hybrid individuals have brown mantle edges unlike the white or purple in the parental species, suggesting allelic or non-allelic genomic interactions. We report differences in gonad development stage between the species consistent with a prezygotic barrier between the species. By incorporating results from publications dating back to 1980, we confirm the long-term stability of the hybrid zone despite higher viability of M. galloprovincialis. This stability coincides with a dramatic change in temperature of UK coastal waters and suggests that these hybrid populations might be resisting the effects of global warming. However, a single SNP locus associated with the Notch transmembrane signalling protein shows a markedly different pattern of variation to the others and might be associated with adaptation of M. galloprovincialis to colder northern temperatures.

Positional influence of center of masticatory forces on occlusal contact forces using a digital occlusal analyzer.

STATEMENT OF PROBLEM: Digital occlusal analyzers allow the recording of dental contact forces. Some authors assume a unique location for the center of contact forces at the position of maximum intercuspation, while others indicate variations in dental contact forces when recorded at different times of the day. Which approach is more appropriate is unclear. PURPOSE: The purpose of this in vitro study was to analyze whether a change in the balance of masticatory forces influences the location of the center of contact forces and its magnitude. MATERIAL AND METHODS: Three different dental casts, selected under dental criteria, were mounted in maximum intercuspation on a semiadjustable articulator equipped with a pattern indicating 9 different force application points (intersection point between 3 longitudinal rows and 3 transverse columns). A force of constant magnitude (169 N) was applied 10 times at each of the application points, and occlusal forces were recorded with a digital occlusal analyzer. Then, two variables were studied: the location of the center of contact forces and its magnitude. Each force application position (9 positions × 3 dental casts=27 in total) was repeated 10 times, and measured data were statistically analyzed with 2-way repeated measures ANOVA (α=.05) test. RESULTS: The repeatability of the method indicated that the coefficient of variation mean was 0.37% in the location of the center of contact forces and that its magnitude was 3.70%. The 2-way repeated measures ANOVA test revealed statistically significant variations in the location of the center of contact forces and its magnitude, revealing that longitudinal changes of the application point of masticatory forces affected the magnitude of contact forces and that longitudinal and transverse changes of the application point of masticatory forces affected the location of the center of contact forces. CONCLUSIONS: The location of the center of contact force and its magnitude provided by a digital occlusal analyzer at the position of maximum intercuspation are not necessarily unique to each articulated dental cast. Even if the intensity of the masticatory force remains unchanged, changes in its lateral or longitudinal balance also influence the result of the occlusion forces.

Two branchial pathological conditions, nuclear hypertrophy and abnormal epithelial proliferation, in Mya arenaria from northwest Russian coasts.

Histopathological analysis of soft-shell clams Mya arenaria collected from 2 northwest Russian locations disclosed high prevalence of 2 pathological gill conditions. One involved the occurrence of more or less extended gill areas in which the branchial filaments showed hyperchromatic (basophilic) epithelium with some hypertrophied nuclei, which were considered presumptive signs of viral infection. Another pathological condition involved abnormal proliferation of the branchial epithelium, which lost the main differential features of the normal branchial epithelium (ciliated and simple cell layer structure), becoming non-ciliated, pseudostratified or stratified hyperchromatic epithelium with abundant mitotic figures and frequent apoptotic cells. The most complex cases involved loss of the normal branchial filament architecture, which was replaced with tumour-like growths consisting of branching, convoluted epithelial projections with a connective stroma. Images suggesting migration (invasion) of cells from the abnormally proliferating epithelium to the subjacent connective tissue, which would involve malignancy, were observed in one individual. The occurrence of both pathological conditions in clams from both locations and their co-occurrence in one clam suggest the possibility of a common, possibly viral, aetiology. Furthermore, the high prevalence of the abnormal proliferative disorder in non-polluted areas suggests an infectious aetiology. Additional studies are needed to assess a viral aetiology for the nuclear hypertrophy and/or the abnormal epithelial proliferation as well as the malignancy of the latter condition.

Integrating Sensor Models in Deep Learning Boosts Performance: Application to Monocular Depth Estimation in Warehouse Automation.

Deep learning is the mainstream paradigm in computer vision and machine learning, but performance is usually not as good as expected when used for applications in robot vision. The problem is that robot sensing is inherently active, and often, relevant data is scarce for many application domains. This calls for novel deep learning approaches that can offer a good performance at a lower data consumption cost. We address here monocular depth estimation in warehouse automation with new methods and three different deep architectures. Our results suggest that the incorporation of sensor models and prior knowledge relative to robotic active vision, can consistently improve the results and learning performance from fewer than usual training samples, as compared to standard data-driven deep learning.

The UJI Aerial Librarian Robot: A Quadcopter for Visual Library Inventory and Book Localisation.

Over time, the field of robotics has provided solutions to automate routine tasks in different scenarios. In particular, libraries are awakening great interest in automated tasks since they are semi-structured environments where machines coexist with humans and several repetitive operations could be automatically performed. In addition, multirotor aerial vehicles have become very popular in many applications over the past decade, however autonomous flight in confined spaces still presents a number of challenges and the use of small drones has not been reported as an automated inventory device within libraries. This paper presents the UJI aerial librarian robot that leverages computer vision techniques to autonomously self-localize and navigate in a library for automated inventory and book localization. A control strategy to navigate along the library bookcases is presented by using visual markers for self-localization during a visual inspection of bookshelves. An image-based book recognition technique is described that combines computer vision techniques to detect the tags on the book spines, followed by an optical character recognizer (OCR) to convert the book code on the tags into text. These data can be used for library inventory. Misplaced books can be automatically detected, and a particular book can be located within the library. Our quadrotor robot was tested in a real library with promising results. The problems encountered and limitation of the system are discussed, along with its relation to similar applications, such as automated inventory in warehouses.

The change of teeth color, whiteness variations and its psychosocial and self-perception effects when using low vs. high concentration bleaching gels: a one-year follow-up.

BACKGROUND: Dental bleaching in traditional concentrations generates greater sensitivity. In this respect, new systems of lower concentration of hydrogen peroxide for tooth bleaching appeared, with color stability unknown over time. The aim of this study was to compare the change and stability of color with low-concentration (6%) hydrogen peroxide gel in an in-office bleaching setting relative to conventional 37.5% gel, including their effects on psychosocial and esthetic self-perception, after 1 year. METHODS: Patients (n = 25) were assessed at 12 months post bleaching treatment (whitening with 6% chemo-activated alkaline formula gel versus 37.5% traditional concentration gel). Color changes were measured objectively using total variation in color (ΔE), and subjectively using Vita Classical and Vita Bleached scale (ΔSGU) by calibrated evaluators (Kappa = 0.85). The Psychosocial Impact of Dental Aesthetics Questionnaire (PIDAQ) and Oral Health Impact Profile (OHIP-14) aesthetic questionnaires were used to measure the self-perception and the psychosocial impact of the bleaching protocols. RESULTS: The effect (ΔE) of 37.5% HP (8.37 ± 2.73) was significantly better than that of 6% HP (5.27 ± 2.53) in terms of color rebound after 1 year of follow-up. There were significant differences in psychosocial impact and esthetic self-perception measurements prior to bleaching versus one-year post-whitening time points; positive effects were maintained. CONCLUSIONS: Low concentration (6%) achieved effective bleaching with good stability after 1 year, accompanied by a positive psychosocial impact and enhanced self-perception at follow-up. TRIAL REGISTRATION: NCT03217994 (before enrollment of the first participant). Data register: July 14, 2017.

Vision for Robust Robot Manipulation.

Advances in Robotics are leading to a new generation of assistant robots working in ordinary, domestic settings. This evolution raises new challenges in the tasks to be accomplished by the robots. This is the case for object manipulation where the detect-approach-grasp loop requires a robust recovery stage, especially when the held object slides. Several proprioceptive sensors have been developed in the last decades, such as tactile sensors or contact switches, that can be used for that purpose; nevertheless, their implementation may considerably restrict the gripper's flexibility and functionality, increasing their cost and complexity. Alternatively, vision can be used since it is an undoubtedly rich source of information, and in particular, depth vision sensors. We present an approach based on depth cameras to robustly evaluate the manipulation success, continuously reporting about any object loss and, consequently, allowing it to robustly recover from this situation. For that, a Lab-colour segmentation allows the robot to identify potential robot manipulators in the image. Then, the depth information is used to detect any edge resulting from two-object contact. The combination of those techniques allows the robot to accurately detect the presence or absence of contact points between the robot manipulator and a held object. An experimental evaluation in realistic indoor environments supports our approach.

Shotgun Proteomics Analysis Discards Alkali Labile Phosphate as a Reliable Method To Assess Vitellogenin Levels in Mytilus galloprovincialis.

Vitellogenin, the egg yolk precursor, is a well-known biomarker of endocrine disruption in oviparous vertebrates. In invertebrates, such as bivalves, it has been used in the last 10 years for the same purpose, despite the limited knowledge of invertebrate endocrinology. In bivalves, vitellogenin levels are usually estimated using an indirect technique, alkali labile phosphate (ALP), that assumes that vitellogenin is the most abundant phosphorylated protein in the analyzed tissue. In this study, we applied shotgun proteomics for the identification and quantification of vitellogenin in marine mussel gonads and compared the results with those obtained with the ALP method. The proteomic analysis revealed that vitellogenin is only detected in female gonads with expression levels that are rather variable among female mussels at different stages of gonad development. ALP analysis, on the contrary, detected similar amounts of phosphorylated proteins regardless of sex or gonad development stage. These results show evidence that the ALP method is not providing reliable information about Vtg levels, at least in marine mussel gonads. ALP is not a good proxy to assess Vtg levels in marine mussels, and careful verification of the adequacy of the procedure should be done before ALP is further assumed as a proxy of Vtg in other bivalve mollusks.

Transmission electron microscopy artifacts in characterization of the nanomaterial-cell interactions.

We investigated transmission electron microscopy artifacts obtained using standard sample preparation protocols applied to the investigation of Escherichia coli cells exposed to common nanomaterials, such as TiO2, Ag, ZnO, and MgO. While the common protocols for some nanomaterials result only in known issues of nanomaterial-independent generation of anomalous deposits due to fixation and staining, for others, there are reactions between the nanomaterial and chemicals used for post-fixation or staining. Only in the case of TiO2 do we observe only the known issues of nanomaterial-independent generation of anomalous deposits due to exceptional chemical stability of this material. For the other three nanomaterials, different artifacts are observed. For each of those, we identify causes of the observed problems and suggest alternative sample preparation protocols to avoid artifacts arising from the sample preparation, which is essential for correct interpretation of the obtained images and drawing correct conclusions on cell-nanomaterial interactions. Finally, we propose modified sample preparation and characterization protocols for comprehensive and conclusive investigations of nanomaterial-cell interactions using electron microscopy and for obtaining clear and unambiguous revelation whether the nanomaterials studied penetrate the cells or accumulate at the cell membranes. In only the case of MgO and ZnO, the unambiguous presence of Zn and Mg could be observed inside the cells.

Subtle tissue and sex-dependent proteome variation in mussel (Mytilus galloprovincialis) populations of the Galician coast (NW Spain) raised in a common environment.

The mussel Mytilus galloprovincialis is one of the most important marine resources for aquaculture in Europe, and Galicia (NW Spain) is the EU's leading region for production. Variation in environmental and ecological factors exists in Northern and Southern estuaries of this region, and natural selection could have modulated genetic variation among populations with adaptation to local conditions as the driving force. Results from a previous genetic study using neutral markers suggested subtle genetic differentiation between mussel populations from both estuarine areas. In this new study, mussel samples from Northern and Southern estuaries were brought into a common environment to test for proteome differences due to genetic and permanent non-genetic effects in populations from both estuarine areas, using both foot and mantle border tissues. Because the sex of the mussels was determined through histological tests, sex-specific effects were also examined. Evidence of subtle differences in the foot proteome, dependent on mussel sex, were detected between populations from both estuaries. These differences were more marked for female samples. No evidence of proteome differences was found for the factors estuaries and sex in mantle border tissue. Candidate proteins with a potential role in local adaptation were identified and point to molecular functions that might be involved in responses to different stressors.

Bibliometric profile of the global scientific research on multiple sclerosis (2003-2012).

BACKGROUND AND OBJECTIVES: The aim of this paper is to analyse the scientific research on multiple sclerosis using a bibliographic analysis of articles published during the period 2003-2012. METHODS: The items under study were obtained from the Science Citation Index-Expanded (SCI-E) database, which was accessed through the Web of Science (WOS) platform. All records with the term 'multiple sclerosis' in the title, plus all articles published in the journals Multiple Sclerosis and Multiple Sclerosis Journal, were analysed. RESULTS: A total of 9778 articles, with 160,966 citations, were retrieved on multiple sclerosis, and the majority of the articles were published in Multiple Sclerosis Journal (n = 1511). The articles were published in journals belonging to 135 different subject areas, with the greatest number of papers falling under the category of clinical neurology. The countries that published the largest numbers of articles were the United States (US) (n = 2786), Italy (n = 1263), the United Kingdom (n = 1147) and Germany (n = 1018). International collaborations produced 20.4% of the papers. CONCLUSIONS: We emphasise the progressive growth of publications worldwide, the publication of articles in a wide variety of journals covering numerous subject areas, and the research leadership of Western countries, most notably European countries, the US and Canada.

Proteomic analysis of eggs from Mytilus edulis females differing in mitochondrial DNA transmission mode.

Many bivalves have an unusual mechanism of mitochondrial DNA (mtDNA) inheritance called doubly uniparental inheritance (DUI) in which distinctly different genomes are inherited through the female (F genome) and male (M genome) lineages. In fertilized eggs that will develop into male embryos, the sperm mitochondria remain in an aggregation, which is believed to be delivered to the primordial germ cells and passed to the next generation through the sperm. In fertilized eggs that will develop into female embryos, the sperm mitochondria are dispersed throughout the developing embryo and make little if any contribution to the next generation. The frequency of embryos with the aggregated or dispersed mitochondrial type varies among females. Previous models of DUI have predicted that maternal nuclear factors cause molecular differences among unfertilized eggs from females producing embryos with predominantly dispersed or aggregated mitochondria. We test this hypothesis using females of each of the two types from a natural population. We have found small, yet detectable, differences of the predicted type at the proteome level. We also provide evidence that eggs of females giving the dispersed pattern have consistently lower expression for different proteasome subunits than eggs of females giving the aggregated pattern. These results, combined with those of an earlier study in which we used hatchery lines of Mytilus, and with a transcriptomic study in a clam that has the DUI system of mtDNA transmission, reinforce the hypothesis that the ubiquitin-proteasome system plays a key role in the mechanism of DUI and sex determination in bivalves. We also report that eggs of females giving the dispersed pattern have higher expression for arginine kinase and enolase, enzymes involved in energy production, whereas ferritin, which is involved in iron homeostasis, has lower expression. We discuss these results in the context of genetic models for DUI and suggest experimental methods for further understanding the role of these proteins in DUI.

Reproductive health patterns: Georgia versus Australia.

The review compares a few reproductive indicators - fertility, abortion and contraception in both a developing (Georgia) and a developed (Australia) country. Fertility, abortion and contraception figures in both countries tend to reflect the attitude and the degree of development of each countries sexual health education and their use of effective contraception. Further research is required to accurately evaluate the need and access of Georgian women to modern methods of family planning and their knowledge of the benefits of modern contraception that can assist to reduce pregnancy termination rate. In Australia better insight is needed on how to facilitate a shift to more efficacious long-term contraceptives across all age groups.

Mechanisms of antibacterial activity of MgO: non-ROS mediated toxicity of MgO nanoparticles towards Escherichia coli.

The toxicity of metal oxide nanomaterials and their antimicrobial activity is attracting increasing attention. Among these materials, MgO is particularly interesting as a low cost, environmentally-friendly material. The toxicity of MgO, similar to other metal oxide nanomaterials, is commonly attributed to the production of reactive oxygen species (ROS). We investigated the toxicity of three different MgO nanoparticle samples, and clearly demonstrated robust toxicity towards Escherichia coli bacterial cells in the absence of ROS production for two MgO nanoparticle samples. Proteomics data also clearly demonstrate the absence of oxidative stress and indicate that the primary mechanism of cell death is related to the cell membrane damage, which does not appear to be due to lipid peroxidation.

An active system for visually-guided reaching in 3D across binocular fixations.

Based on the importance of relative disparity between objects for accurate hand-eye coordination, this paper presents a biological approach inspired by the cortical neural architecture. So, the motor information is coded in egocentric coordinates obtained from the allocentric representation of the space (in terms of disparity) generated from the egocentric representation of the visual information (image coordinates). In that way, the different aspects of the visuomotor coordination are integrated: an active vision system, composed of two vergent cameras; a module for the 2D binocular disparity estimation based on a local estimation of phase differences performed through a bank of Gabor filters; and a robotic actuator to perform the corresponding tasks (visually-guided reaching). The approach's performance is evaluated through experiments on both simulated and real data.

Plasma Exchange in Anti-Signal Recognition Particle Myopathy: A Systematic Review and Combined Analysis of Patient Individual Data.

Anti-signal recognition particle myopathy (anti-SRP myopathy) is a rare subtype of immune-mediated inflammatory myopathy characterized by muscle weakness and anti-SRP autoantibodies. Although plasma exchange (PE) is used in severe cases, its role remains unclear. A systematic review was conducted following PRISMA guidelines, identifying 23 patients with anti-SRP myopathy treated with PE. Data on demographics, clinical features, laboratory findings, treatments, and outcomes were analyzed combining individual patient data if available. Sixteen (69.6%) patients were male, with muscle weakness as the predominant symptom in 100% of cases. After PE, most patients showed improvement in symptoms, and the proportion of patients with muscle weakness was reduced (p = 0.001). Relapse occurred in 17.4% of the cases. The incidence of adverse events was low (8.7%). Despite limitations, including a small sample size and heterogeneous data, our systematic review suggests that PE may be effective in inducing remission and controlling symptoms in anti-SRP myopathy, particularly in severe cases. Since evidence on PE in anti-SRP myopathy is limited, further research, including prospective multicenter studies, is warranted to understand better its efficacy and safety and establish its role in treatment algorithms.

Analysis of prognostic factors associated with longitudinally extensive transverse myelitis.

OBJECTIVE: The aim of this study is to report the clinical profile and outcome of longitudinally extensive transverse myelitis (LETM). METHODS: We prospectively studied adult patients who presented with LETM from January 2008 to December 2011. Information on demographic, clinical course, magnetic resonance imaging (MRI) and outcome was collected. HLA-DRB1 genotype was compared with those of 225 normal controls and patients with MS (228) and neuromyelitis optica (NMO) (22). RESULTS: In total, 23 patients (16 female) with a median age of 44.5 years (range: 20-77 years) were included. Most (74%) had moderate-severe disability at nadir (48% non-ambulatory), normal/non-multiple sclerosis (MS) brain MRI (96%) and a median MRI cord lesion of 5 vertebral segments (range: 3-19). Laboratory analysis showed cerebrospinal fluid pleocytosis (45%), NMO-IgG (9%), antinuclear antibodies (70%), and genotype HLA-DRB1*13 (57%). The frequency of DRB1*13 genotype was higher compared with controls (p=0.002), MS (p=0.001) and NMO (p=0.003) patients. After a median follow-up of 32 months, one patient converted to MS, two had relapsing LETM with NMO-IgG, and 20 remained as idiopathic with recurrences in four (20%). Twelve (52%) patients recovered with minimal disability (Expanded Disability Status Scale (EDSS) ≤2.5) and three (13%) remained wheelchair dependent. Disability at nadir was associated with the final outcome and extension of the spinal cord lesion with risk of recurrence. Recurrence was not associated with worse outcome. CONCLUSIONS: Inflammatory LETM is mostly idiopathic with a good outcome. It includes a relatively homogenous group of patients with an overrepresentation of the HLA-DRB1*13 genotype. EDSS at nadir is a predictor of the final outcome and extension of the myelitis of the recurrence risk.

Multiple hypothesis testing in proteomics: a strategy for experimental work.

In quantitative proteomics work, the differences in expression of many separate proteins are routinely examined to test for significant differences between treatments. This leads to the multiple hypothesis testing problem: when many separate tests are performed many will be significant by chance and be false positive results. Statistical methods such as the false discovery rate method that deal with this problem have been disseminated for more than one decade. However a survey of proteomics journals shows that such tests are not widely implemented in one commonly used technique, quantitative proteomics using two-dimensional electrophoresis. We outline a selection of multiple hypothesis testing methods, including some that are well known and some lesser known, and present a simple strategy for their use by the experimental scientist in quantitative proteomics work generally. The strategy focuses on the desirability of simultaneous use of several different methods, the choice and emphasis dependent on research priorities and the results in hand. This approach is demonstrated using case scenarios with experimental and simulated model data.