Relative Quantification of Proteins in Formalin-Fixed Paraffin-Embedded Breast Cancer Tissue Using Multiplexed Mass Spectrometry Assays.

The identification of clinically relevant biomarkers represents an important challenge in oncology. This problem can be addressed with biomarker discovery and verification studies performed directly in tumor samples using formalin-fixed paraffin-embedded (FFPE) tissues. However, reliably measuring proteins in FFPE samples remains challenging.

Steering Away from Current Amoxicillin Dose Reductions in Hospitalized Patients with Impaired Kidney Function to Avoid Subtherapeutic Drug Exposure.

Current dose reductions recommended for amoxicillin in patients with impaired kidney function could lead to suboptimal treatments. In a prospective, observational study in hospitalized adults with varying kidney function treated with an IV or oral dose of amoxicillin, amoxicillin concentrations were measured in 1−2 samples on the second day of treatment. Pharmacometric modelling and simulations were performed to evaluate the probability of target attainment (PTA) for 40% of the time above MIC following standard (1000 mg q6h), reduced or increased IV dosing strategies.

Therapeutic drug monitoring and clinical outcomes in severely ill patients receiving amoxicillin: a single-centre prospective cohort study.

Therapeutic drug monitoring (TDM) of β-lactam antibiotics is increasingly used to overcome rising antimicrobial resistance and improve antibiotic exposure. However, there is little guidance on target amoxicillin plasma concentrations. We aimed to define these by evaluating associations between amoxicillin concentrations and clinical outcomes.

[Vitamin deficiencies in the adult outpatient setting].

Since their discovery more than a century ago to this day, vitamins went from misunderstood molecules with mysterious properties to fundamental components with undoubted clinical implications. Despite the scientific progresses in the understanding of their physiopathological role, vitamins raise to this day multiple interrogations in clinical practice. This article aims at answering questions that are frequently encountered in the outpatient setting regarding vitamin deficiencies: who to screen ? At what moment ? By which test ? How to interpret the results ? How to supplement ? By answering these questions, we hope to provide the general practitioners with a pragmatic tool to guide them in the management of issues related to vitamins.

Internal calibration as an emerging approach for endogenous analyte quantification: Application to steroids.

The use of mass spectrometry methods with triple quadrupole instruments is well established for quantification. However, the preparation of calibration curves can be time-consuming and prone to analytical errors. In this study, an innovative internal calibration (IC) approach using a one-standard calibration with a stable isotope-labeled (SIL) standard version of the endogenous compound was developed.

Case Report: Low Hematocrit Leading to Tacrolimus Toxicity.

Tacrolimus is a calcineurin inhibitor characterized by a narrow therapeutic index and high intra- and inter-individual pharmacokinetic variability. Therapeutic drug monitoring in whole-blood is the standard monitoring procedure. However, tacrolimus extensively binds to erythrocytes, and tacrolimus whole-blood distribution and whole-blood trough concentrations are strongly affected by hematocrit.

Combining the advantages of multilevel and orthogonal partial least squares data analysis for longitudinal metabolomics: Application to kidney transplantation.

Kidney transplantation is one of the renal replacement options in patients suffering from end-stage renal disease (ESRD). After a transplant, patient follow-up is essential and is mostly based on immunosuppressive drug levels control, creatinine measurement and kidney biopsy in case of a rejection suspicion. The extensive analysis of metabolite levels offered by metabolomics might improve patient monitoring, help in the surveillance of the restoration of a "normal" renal function and possibly also predict rejection.

Clinical method evaluation of hemoglobin S and C identification by top-down selected reaction monitoring and electron transfer dissociation.

Background: Biological diagnosis of hemoglobin disorders is a complex process relying on the combination of several analytical techniques to identify Hb variants in a particular sample. Currently, hematology laboratories usually use high-performance liquid chromatography (HPLC), capillary electrophoresis and gel-based methods to characterize Hb variants. Co-elution and co-migration may represent major issues for precise identification of Hb variants, even for the most common ones such as Hb S and C.

Human Neural Organoids for Studying Brain Cancer and Neurodegenerative Diseases.

The lack of relevant in vitro neural models is an important obstacle on medical progress for neuropathologies. Establishment of relevant cellular models is crucial both to better understand the pathological mechanisms of these diseases and identify new therapeutic targets and strategies. To be pertinent, an in vitro model must reproduce the pathological features of a human disease.

Detection of Unknown Chemical Adduct Modifications on Proteins: From Wet to Dry Laboratory.

The detection and characterization of chemical adducts on proteins is of increasing interest. Here, we described a step-by-step procedure to identify unknown chemical adduct modifications on proteins resulting from the interaction with a given reactive compound. The protocol can be divided into two equally important parts: (1) the wet laboratory work, to produce high quality mass spectrometry (MS) data of in vitro modified proteins and (2) the dry laboratory work, to analyze the generated MS data and provide highly confident qualitative and quantitative results on the chemical composition and amino acid localization of adducts.

Liquid chromatography-high resolution mass spectrometry for broad-spectrum drug screening of dried blood spot as microsampling procedure.

Background: Hyphenation of liquid chromatography (LC) with high-resolution mass spectrometry (HRMS) offers the potential to develop broad-spectrum screening procedures from low volumes of biological matrices. In parallel, dried blood spot (DBS) has become a valuable tool in the bioanalysis landscape to overcome conventional blood collection issues. Herein, we demonstrated the applicability of DBS as micro-sampling procedure for broad-spectrum toxicological screening.

Toward a better understanding of chronic kidney disease with complementary chromatographic methods hyphenated with mass spectrometry for improved polar metabolome coverage.

The prevalence of chronic kidney disease (CKD) is increasing worldwide. New technical approaches are needed to improve early diagnosis, disease understanding and patient monitoring, and to evaluate new therapies. Metabolomics, as a prime candidate in the field of CKD research, aims to comprehensively analyze the metabolic complexity of biological systems.

Development of a Highly Multiplexed SRM Assay for Biomarker Discovery in Formalin-Fixed Paraffin-Embedded Tissues.

The search for novel and clinically relevant biomarkers still represents a major clinical challenge and mass-spectrometry-based technologies are essential tools to help in this process. In this application, we demonstrate how selected reaction monitoring (SRM) can be applied in a highly multiplexed way to analyze formalin-fixed paraffin-embedded (FFPE) tissues. Such an assay can be used to analyze numerous samples for narrowing down a list of potential biomarkers to the most relevant candidates.

Detection of Proteoforms Using Top-Down Mass Spectrometry and Diagnostic Ions.

Characterization of protein structure modifications is an important field in mass spectrometry (MS)-based proteomics. Here, we describe a process to quickly and reliably identify a mass change in a targeted protein sequence by top-down mass spectrometry (TD MS) using electron transfer dissociation (ETD). The step-by-step procedure describes how to develop a TD MS method for data acquisition as well as the data analysis process.

Therapeutic drug monitoring of imipenem and the incidence of toxicity and failure in hospitalized patients: a retrospective cohort study.

Objectives: Therapeutic drug monitoring (TDM) of beta-lactam antibiotics is increasingly employed to ensure adequate antibiotic exposure and slow emergence of resistance. Imipenem's therapeutic range has not been defined; we report plasma concentrations and clinical outcomes of patients receiving imipenem for bacterial infections.

Methods: All hospitalized adult patients undergoing imipenem TDM during therapy for suspected or confirmed bacterial infections between 1 January 2013 and 28 February 2017 were included in this single-centre retrospective cohort.

IL8 and EDEM3 gene expression ratio indicates peripheral blood mononuclear cell (PBMC) quality.

Background: Uncontrolled preanalytical variables can reduce the accuracy and reproducibility of downstream analytical results from peripheral blood mononuclear cells (PBMCs).

Methods: PBMCs were isolated from EDTA and citrate-anticoagulated blood samples, obtained from healthy subjects and patients with inflammatory and infectious conditions. PBMC-derived RNA samples were examined for gene expression changes induced by extended blood pre-centrifugation delays at 4 °C and RT.

Regulatory context and validation of assays for clinical mass spectrometry proteomics (cMSP) methods.

Clinical mass spectrometry proteomics (cMSP) assays are being increasingly used in clinical laboratories for analyzing peptides and proteins. It has therefore become urgent to characterize and validate the methods available for liquid chromatography-tandem mass spectrometry (LC/MS-MS) targeted quantification of peptide and protein biomarkers in biological fluids in the context of in vitro diagnostics. LC-MS/MS for the detection of peptides and proteins is currently the main approach used in the field of cMSP.

CRP-Based Cardiovascular Risk Assessment: New Conventional CRP Assay Fit for Purpose?

Background: Subclinical inflammation was shown to play a role in the context of cardiovascular disorder processes. American College of Cardiology/American Heart Association guidelines on cardiovascular risk assessment in specific clinical contexts recommend the use of C-reactive protein (CRP) measurement with high sensitive (hs)-CRP assays that meet the precision requirements for values <2 mg/L. Until now, only hs-CRP assays reached the required limit of quantification.

IL8 and IL16 levels indicate serum and plasma quality.

Background: Longer pre-centrifugation times alter the quality of serum and plasma samples. Markers for such delays in sample processing and hence for the sample quality, have been identified.

Methods: Twenty cytokines in serum, EDTA plasma and citrate plasma samples were screened for changes in concentration induced by extended blood pre-centrifugation delays at room temperature.