Mass spectrometry assay as an alternative to the enzyme-linked immunosorbent assay test for biomarker quantitation in ecotoxicology: Application to vitellogenin in Crustacea (Gammarus fossarum)

Vitellogenin (Vg) is a widespread biomarker for measuring exposure to endocrine disruptors. Vg quantification is usually done by using the ELISA test (enzyme-linked immunosorbent assay). Since this test is specific to a target protein, it can rarely be used with other species due to low cross-reactivity across species. Therefore alternative analytical methods have to be considered as the development of a specific and sensitive ELISA test for each new target is time-consuming and may prove unsuccessful. This paper focuses on the development of a quantitative assay by liquid chromatography tandem mass spectrometry (LC-MS/MS) of vitelogenin in an invertebrate (Gammarus fossarum) for which no ELISA kit is available. The linearity of the method was within the concentration range 2.5–25,000 pg/mL and the limit of detection was estimated at 0.75 pg/mL of Vg. This method has been demonstrated to be an alternative to existing immunological methods for quantifying Vg in invertebrates due to its greater sensitivity, specificity and reproducibility (intra- and inter-assay below 15%). This assay was applied for Vg determination in female G. fossarum following exposure to a known endocrine disruptor, methyl farnesoate, in crustaceans. The availability of a quantitative G. fossarum LC-MS/MS assay should open the way for further studies to evaluate xenoestrogen effects in aquatic male G. fossarum.     

Folding of a salivary intrinsically disordered protein upon binding to tannins

We used ion mobility spectrometry to explore conformational adaptability of intrinsically disordered proteins bound to their targets in complex mixtures. We investigated the interactions between a human salivary proline-rich protein IB5 and a model of wine and tea tannin: epigallocatechin gallate (EgCG). Collisional cross sections of naked IB5 and IB5 complexed with N = 1-15 tannins were recorded. The data demonstrate that IB5 undergoes an unfolded to folded structural transition upon binding with EgCG.     

Delayed random walks

A delayed random walk $\text{{S}{}^1{}_{\mathrm{n}}\text{, n ≥ 0}}$ is defined here as a partial sum process of independent random variables in which the first N summands (N optional) are distributed F¹,…,F_N, respectively, while all remaining summands are distributed F₀, where {F_k, _k ≥ 0} is a sequence of proper distribution functions on the real line. Delayed random walks arise naturally in the study of certain generalized single server queues. This paper examines optional times of the process such as $\text{π = inf {n: n ≥ 1 and S}{}^1{}_{\mathrm{n}}\text{≥ 0}}$. Conditions insuring the finiteness of E {π} and E {π²} are obtained, generating functions calculated, and illustrative examples given. The bivariate functions $\text{E{r}{}^{\mathrm{\pi }}\text{explsqbitS}{}^1{}_{\mathrm{\pi }}\text{rsqb}}$ and $\text{E {}\text{∑}\text{n=0}\text{π?1}\text{explsqbitS}{}^1{}_{\mathrm{n}}\text{rsqb}}$ are studied for the case where N ≡ 1.     

Quantum studies of H atom trapping on a graphite surface

The trapping and sticking of H and D atoms on the graphite (0001) surface is examined, over the energy range of 0.1-0.9 eV. For hydrogen to chemisorb onto graphite, the bonding carbon must pucker out of the surface plane by several tenths of an angstrom. A quantum approach in which both the hydrogen and the bonding carbon atoms can move is used to model the trapping, and a potential energy surface based on density functional theory calculations is employed. It is found, for energies not too far above the 0.2 eV barrier to chemisorption that a significant fraction of the incident H or D atoms can trap. The forces on the bonding carbon are large, and it can reconstruct within 50 fs or so. After about 100 fs, most of the trapped H atoms scatter back into the gas phase, but the 5%-10% that remain can have lifetimes on the order of a picosecond or more. Calculations of the resonance eigenstates and lifetimes confirm this. An additional lattice degree of freedom is included quantum mechanically and is shown to significantly increase the amount of H that remains trapped after 1 ps. Further increasing the incident energy destabilizes the trapped state, leading to less H remaining trapped at long times. We estimate that for a full dissipative bath, the sticking probabilities should be on the order of 0.1.     

Improved detection specificity for plasma proteins by targeting cysteine-containing peptides with photo-SRM

Targeted mass spectrometry using selected reaction monitoring (SRM) has emerged as an alternative to immunoassays for protein quantification owing to faster development time and higher multiplexing capability. However, the SRM strategy is faced with the high complexity of peptide mixtures after trypsin digestion of whole plasma or the cellular proteome that most of the time causes contamination, irremediably, by interfering compounds in the transition channels monitored. This problem becomes increasingly acute when the targeted protein is present at a low concentration. In this work, the merit of laser-induced photo-dissociation in the visible region at 473 nm implemented in an hybrid quadrupole linear ion-trap mass spectrometer (photo-SRM) was evaluated for detection specificity of cysteine-containing peptides in a group of plasma proteins after tagging with a dabcyl chromophore. Compared with conventional SRM, photo-SRM chromatograms have improved detection specificity for most of peptides monitored. Comparison of the signals obtained for the best proteotypic peptides in SRM mode and those recorded by photo-SRM of cysteine-containing peptides for the same proteins reveals either increased (up to 10-fold) or similar signal to photo-SRM detection. Finally, photo-SRM has extended response linearity across a calibration plot obtained by diluting human plasma in rat plasma, down to the lowest concentrations. Hence, photo-SRM may advantageously complement conventional SRM in assay of proteins in complex biological matrices.     

Chloro(2,9-dimethyl-1,10-phenanthroline-N,N′)(isoquinoline-1-carboxylato-O,N)copper(II)

The asymmetric unit of the title compound, [CuCl(C₁₀H₆NO₂)(C₁₄H₁₂N₂)], contains two monomeric copper mol­ecules, A and B. Each Cu atom is coordinated to one 2,9-di­methyl-1,10-phenanthroline (neocuproine) ligand via both N atoms, to one iso­quinoline-1-carboxyl­ate anion (IQC⁻) via the N and one O atom, and to one Cl⁻ anion. The environment of the Cu atom is approximately square pyramidal, with the apical position occupied by an N atom of neocuproine. In mol­ecule A, the Cu atom is 0.301 (1) Å above the basal plane; this distance is 0.316 (1) Å in mol­ecule B. The crystal packing is characterized by several hydrogen bonds.