BEAMS Lab at MIT: Status report

The Biological Engineering Accelerator Mass Spectrometry (BEAMS) Lab at the Massachusetts Institute of Technology is a facility dedicated to incorporating AMS into life sciences research. As such, it is focused exclusively on radiocarbon and tritium AMS and makes use of a particularly compact instrument of a size compatible with most laboratory space. Recent developments at the BEAMS Lab were aimed to improve different stages of the measurement process, such as the carbon sample injection interface, the simultaneous detection of tritium and hydrogen and finally, the overall operation of the system. Upgrades and results of those efforts are presented here.United States. National Institutes of Health (grant P30-ES02109)United States. National Institutes of Health (grant R42-CA084688)National Institutes of Health. National Center for Research Resources (grant UL1 RR 025005)GlaxoSmithKlin

Progress with a gas-accepting ion source for Accelerator Mass Spectrometry

Author Posting. © The Author(s), 2010. This is the author's version of the work. It is posted here by permission of Elsevier B.V. for personal use, not for redistribution. The definitive version was published in Nuclear Instruments and Methods in Physics Research Section B: Beam Interactions with Materials and Atoms 269 (2011): 3192–3195, doi:10.1016/j.nimb.2011.04.017.The National Ocean Sciences AMS (NOSAMS) facility at Woods Hole Oceanographic Institution has developed a novel, gas-accepting microwave-plasma ion-source. The source is a key component of a compact Accelerator Mass Spectrometry (AMS) system built for the analysis of 14C in a continuously flowing gas stream. The gas source produces carbon currents from a stream of CO2 with currents typical of a traditional graphite source. Details of the gas source, including ion current achieved, optimal flow rate, efficiency, and memory are presented. Additionally, data obtained from coupling a gas chromatograph to the source to will be shown

Photoactive assemblies of organic compounds and biomolecules: drug-protein supramolecular systems

[EN] The properties of singlet and triplet excited states are strongly medium-dependent. Hence, these species constitute valuable tools as reporters to probe compartmentalised microenvironments, including drug@protein supramolecular systems. In the present review, the attention is focused on the photophysical properties of the probe drugs (rather than those of the protein chromophores) using transport proteins (serum albumins and 1-acid glycoproteins) as hosts. Specifically, fluorescence measurements allow investigating the structural and dynamic properties of biomolecules or their complexes. Thus, the emission quantum yields and the decay kinetics of the drug singlet excited states provide key information to determine important parameters such as the stoichiometry of the complex, the binding constant, the relative degrees of occupancy of the different compartments, etc. Application of the FRET concept allows determining donor-acceptor interchromophoric distances. In addition, anisotropy measurements can be related to the orientation of the drug within the binding sites, where the degrees of freedom for conformational relaxation are restricted. Transient absorption spectroscopy is also a potentially powerful tool to investigate the binding of drugs to proteins, where formation of encapsulated triplet excited states is favoured over other possible processes leading to ionic species (i. e. radical ions), and their photophysical properties are markedly sensitive to the microenvironment experienced within the protein binding sites. Even under aerobic conditions, the triplet lifetimes of protein-complexed drugs are remarkably long, which provides a broad dynamic range for identification of distinct triplet populations or for chiral discrimination. Specific applications of the laser flash photolysis technique include the determination of drug distribution among the bulk solution and the protein binding sites, competition of two types of proteins to bind a 3 drug, occurrence of drug-drug interactions within protein binding sites, enzymatic-like activity of the protein or determination of enantiomeric compositions. The use of proteins as supramolecular hosts modifies the photoreactivity of encapsulated substrates by providing protection against oxygen or other external reagents, by imposing conformational restrictions in the binding pockets, or by influencing the stereochemical outcome. In this review, a selected group of examples is presented including decarboxylation, dehalogenation, nucleophilic addition, dimerisation, oxidation, Norrish type II reaction, photo-Fries rearrangement and 6 electrocyclisationFinancial support from the Spanish Government (CTQ2010-14882, JCI-2011-09926, RyC-2007-00476), from the EU (PCIG12-GA-2012-334257), from the Universitat Politènica de València (SP20120757) and from the Consellería de Educació, Cultura i Esport (PROMETEOII/2013/005, GV/2013/051) is gratefully acknowledged.Vayá Pérez, I.; Lhiaubet-Vallet, VL.; Jiménez Molero, MC.; Miranda Alonso, MÁ. (2014). Photoactive assemblies of organic compounds and biomolecules: drug-protein supramolecular systems. Chemical Society Reviews. 43:4102-4122. https://doi.org/10.1039/C3CS60413FS410241224

Recent developments in protein–ligand affinity mass spectrometry

This review provides an overview of direct and indirect technologies to screen protein–ligand interactions with mass spectrometry. These technologies have as a key feature the selection or affinity purification of ligands in mixtures prior to detection. Specific fields of interest for these technologies are metabolic profiling of bioactive metabolites, natural extract screening, and the screening of libraries for bioactives, such as parallel synthesis libraries and small combichem libraries. The review addresses the principles of each of the methods discussed, with a focus on developments in recent years, and the applicability of the methods to lead generation and development in drug discovery

Bioanalytical Challenges in Support of Complex Modalities of Antibody-Based Therapeutics

Antibody-based therapeutic classes are evolving from monoclonal antibodies to antibody derivatives with complex structures to achieve advanced therapeutic effect. These antibody derivatives may contain multiple functional domains and are often vulnerable to in vivo biotransformation. Understanding the pharmacokinetics of these antibody derivatives requires a sophisticated bioanalytical approach to carefully characterize the whole drug and each functional domain with respect to quantity, functionality enabled by biotransformation, and corresponding immune responses. Ligand binding assays and liquid chromatography-mass spectrometry assays are predominantly used in bioanalytical support of monoclonal antibodies and are continuously used for antibody derivatives such as antibody drug conjugate and bispecific antibodies. However, they become increasingly cumbersome in coping with increased complexity of drug modality and associated biotransformation. In this mini-review, we examined the current pharmacokinetic assays in the literature for antibody drug conjugate and bispecific antibodies, and presented our view of promising bioanalytical technologies to address the distinct bioanalytical needs of complex modalities

Metabolite profiling using a novel QqQ linear ion trap mass spectrometer

A novel hybrid RF/DC quadrupole-linear ion trap mass spectrometer (QqQ linear ion trap) was developed for analizing metabolite profiling. Primary hepatocyte incubations were used to demonstrate the utility of the QqQ linear ion trap mass spectrometer for metabolite profiling. High sensitivity in full scan mode MS, MS 2 and MS 3 modes, was provided by the ion trap scan functions of the QqQ linear ion trap. The flexibility to simultaneously perform quantitative and qualitative analysis was provided by MRM scan function. The experimental results show that seven major metabolites were observed during incubations.link_to_subscribed_fulltex

Dixon's Q-test and Student's t-test to assess analog internal standard response in nonregulated LC-MS/MS bioanalysis

Aim: In bioanalytical assays, analyte response is normalized to an internal standard response. When the internal standard works well, it compensates for processing and detection variability. However, in case the internal standard introduces additional variability, due to addition errors or other issues, scientists need to identify this. Results: A new method, using a Q-test for outliers and a t-test to compare internal standard response from different sample types, is applied to 15 cases. The results show that the Q-test/t-test, which uses confidence level rather than arbitrary cut-points, is more discerning of deviations compared with widely used methods. Conclusion: This work may improve the quality of and rationale for the internal standard response monitoring method

Practical Approaches to Incurred Sample LC-MS/MS Reanalysis: Confirming Unexpected Results

Incurred sample reanalysis (ISR) is an important step in assuring the quality of an LC-MS/MS bioanalytical assay and the integrity of bioanalysis conduct. A conventional ISR involves analysis of at least 20 samples taken from an in vivo study a second time using the method that was described in pre-study validation and employed in generating the initial study sample results. However, this practice is sometimes inadequate to confirm bioanalytical results that are unexpected. The present report discuss several additional exploratory activities that were performed to confirm the unexpected plasma concentration-time results of NVP-1, an investigational drug candidate, observed in the plasma samples collected from patients in a phase II trial. These approaches include (1) LC-MS/MS reanalysis of the study samples after multiple freeze/thaw cycles followed by a short term bench top storage, (2) evaluation of additional MS/MS transitions in LC-MS/MS, (3) employment of different sample preparation procedure in LC-MS/MS and (4) study sample dilution using plasma samples from healthy volunteers. These procedures are practical and can be readily implemented to provide confirmatory LC-MS/MS bioanalysis of other small molecules

Quantitative analysis of clofazimine (Lamprene®), an antileprosy agent, in human dried blood spots using liquid chromatography–tandem mass spectrometry

An LC–MS/MS method was developed and validated for bioanalysis of clofazimine in human dried blood spot (DBS) samples in support of a clinical study on multidrug-resistant tuberculosis in developing countries. The validated assay dynamic range was from 10.0 to 2000 ng/mL using a 1/8 inch DBS punch. The accuracy and precision of the assay were ±11.0% (bias) and ≤13.5% (CV) for the LLOQs (10.0 ng/mL) and ±15% (bias) and ≤15% (CV) for all other QC levels. The assay was proved to be free from the possible impact owing to spot size and storage temperature (e.g. at 60°C, ≤ − 60°C). The validated assay is well suited for the intended clinical study where conventional pharmacokinetic sample collection is not feasible