Animal models and dose selectionThis analysis utilized three preclinical models from two species of humanized mice: stem cell hemopoietic/RAG 2-(hu-HSC-Rag, n=36) and bone marrow-liver-thymus (BLT, n=13) and one species of nonhuman primate (NHP): rhesus macaques (n=18).Humanization of the mice was by protocols that have been previously described (Melkus et al. 2006; Berges et al. 2006). The extent of humanization for the mouse models was assessed by quantifying the human T-cell populations using flow cytometry. All the humanized mice were female, while six (33.3%) NHPs were female. Half of the animals were left uninfected while the other half were infected with 200 μL of 2.1 x 10 6 IU/mL of HIVBal D7 intraperitoneally (hu-HSC-RAG) or 200 μL of 90,000 tissue culture infectious units (TCIU) of HIVJRcsf intravenously (BLT) or 10 4.5 median tissue culture infective dose (TCID50) RT-SHIV intravenously (NHPs) (North et al. 2005; North et al. 2010) for four weeks. The hu-HSC-RAG and BLT mice underwent humanization when aged three to six months and were then dosed with various ARV regimens for a period of ten days. Hu-HSC-RAG mice were dosed with EFV 10 mg/kg only (six uninfected and six infected animals) or atazanavir (ATZ) 140 mg/kg only (six uninfected and six infected animals) or a combination of tenofovir (TFV) 208 mg/kg, emtricitabine (FTC) 240 mg/kg, raltegravir (RAL) 56 mg/kg and maraviroc (MVC) 62 mg/kg (six uninfected and six infected animals). BLT mice were dosed with a combination of TFV, FTC, RAL, MVC and ATZ at equivalent doses but not EFV due to toxicity concerns. All drugs were administered by oral gavage. Dosing regimens for each of the animal models are summarized in Supplementary Table 1. Dosage selection for all drugs in the animal

A microchip electrospray ionization source was coupled with high pressure mass spectrometry (HPMS). A continuous atmospheric inlet consisting of a stainless steel capillary and DC ion optics was designed to conduct ions into the mass spectrometer. Infusions of amino acids and peptides were performed and detected with a miniature cylindrical ion trap (mini-CIT)-based mass spectrometer operated at ≥1 Torr with air as the buffer gas. Detection of glycine and thymopentin (separately) demonstrated the mass range of the mini-CIT detector could span from m/z 75 to 681. A microchip capillary electrophoresis (CE) separation with mini-CIT detection was performed, and the results were compared with detection using a commercial instrument (Waters Synapt G2). Comparable separation efficiencies were observed with both mass spectrometers as detectors, with about 6 times better signal-to-noise observed on the Synapt G2. Comparison of mass spectra in the two systems reveals similar features observed, but with wider peak widths in the mini-CIT than on the Synapt G2 as expected due to high-pressure operation.

Here, we assess infrared matrix assisted laser desorption electrospray ionization (IR-MALDESI) mass spectrometry imaging (MSI) analysis of hair as a clinical tool for monitoring patient adherence to the antiretroviral maraviroc (MVC). A custom MATLAB-based algorithm has been developed to streamline data analysis and generate longitudinal profiles of drug incorporation along the length of hair strands. Hair strands from volunteers enrolled in a directly observed therapy study were analyzed by IR-MALDESI MSI and processed using this tool to characterize the profiles of single doses and a daily dose regimen of MVC. Single dose responses were 1.7 [1.1, 2.5] mm (median [range]) wide along the length of the hair and were detected in 8 out of 12 volunteers. Daily dose profiles capturing 28 days of continuous dosing were approximately 5 times the intensity of single dose profiles and 10.5 [7.0, 13] mm wide, corresponding to 1 month of hair growth. MVC ion abundance was observed in all 12 volunteers for the daily dosing period. Daily dosing profiles were consistent with a model of MVC accumulation in hair based on linear superposition of a single dose response, indicating the potential for prediction of daily drug-taking behavior based on deconvolution of a complex longitudinal profile in hair.

HPMS significantly reduces the pumping requirements required for miniature mass spectrometers and the use of ambient air buffer gas further reduces size, weight, and logistics requirements. Mass analysis at high pressures of ambient air is another important step for the development of hand portable mass spectrometers. Copyright © 2016 John Wiley & Sons, Ltd.

Improvements were made to a previously developed platform coupling microchip capillary electrophoresis (CE) with high pressure mass spectrometry (HPMS). The RF drive frequency was increased to over 30 MHz from less than 10 MHz, and the ion trap was scaled down to 100 μm critical dimensions. A stretched length ion trap was used to improve sensitivity, and a tube lens was used to improve ion transmission. Detection of the 20 common amino acids was demonstrated, resulting in an average improvement of signal-tonoise of 28 times and an average improvement in peak width of 2.6 times over those obtained in previous work. Consumption of amino acids by cells in growth media was monitored over time using the improved CE-HPMS platform, and several amino acids were shown to be consumed at different rates, demonstrating the potential for real-time bioreactor monitoring.