S-cysteinylated albumin and methionine-oxidized apolipoprotein A-I (apoA-I) have been posed as candidate markers of diseases associated with oxidative stress. Here, a dilute-and-shoot form of LC-electrospray ionization-MS requiring half a microliter of blood plasma was employed to simultaneously quantify the relative abundance of these oxidized proteoforms in samples stored at ؊80°C, ؊20°C, and room temperature and exposed to multiple freeze-thaw cycles and other adverse conditions in order to assess the possibility that protein oxidation may occur as a result of poor sample storage or handling. Samples from a healthy donor and a participant with poorly controlled type 2 diabetes started at the same low level of protein oxidation and behaved similarly; significant increases in albumin oxidation via S-cysteinylation were found to occur within hours at room temperature and days at ؊20°C. Methionine oxidation of apoA-I took place on a longer time scale, setting in after albumin oxidation reached a plateau. Freeze-thaw cycles had a minimal effect on protein oxidation. In matched collections, protein oxidation in serum was the same as that in plasma. Albumin and apoA-I oxidation were not affected by sample headspace or the degree to which vials were sealed. ApoA-I, however, was unexpectedly found to oxidize faster in samples with lower surface-area-to-volume ratios. An initial survey of samples from patients with inflammatory conditions normally associated with elevated oxidative stress-including acute myocardial infarction and prostate cancer-demonstrated a lack of detectable apoA-I oxidation. Albumin S-cysteinylation in these samples was consistent with known but relatively brief exposures to temperatures above ؊30°C (the freez-

Clinical assessment of myocardial ischemia based on visually-assessed wall motion scoring from echocardiography is semiquantitative, operator dependent, and heavily weighted by operator experience and expertise. Cardiac motion estimation methods such as tissue Doppler imaging, used to assess myocardial muscle velocity, provides quantitative parameters such as the strain-rate and strain derived from Doppler velocity. However, tissue Doppler imaging does not differentiate between active contraction and simple rotation or translation of the heart wall, nor does it differentiate tethering (passively following) tissue from active contraction. In this paper, we present a strain imaging modality called cardiac elastography that provides two-dimensional strain information. A method for obtaining and displaying both directional and magnitude cardiac elastograms and displaying strain over the entire cross-section of the heart is described. Elastograms from a patient with coronary artery disease are compared with those from a healthy volunteer. Though observational, the differences suggest that cardiac elastography may be a useful tool for assessment of myocardial function. The method is two-dimensional, real time and avoids the disadvantage of observer-dependent judgment of myocardial contraction and relaxation estimated from conventional echocardiography.

Exposure of blood plasma/serum (P/S) to thawed conditions (> −30 °C) can produce biomolecular changes that skew measurements of biomarkers within archived patient samples, potentially rendering them unfit for molecular analysis. Because freeze-thaw histories are often poorly documented, objective methods for assessing molecular fitness before analysis are needed. We report a 10-μl, dilute-and-shoot, intact-protein mass spectrometric assay of albumin proteoforms called “ΔS-Cys-Albumin” that quantifies cumulative exposure of archived P/S samples to thawed conditions. The relative abundance of S-cysteinylated (oxidized) albumin in P/S increases inexorably but to a maximum value under 100% when samples are exposed to temperatures > −30 °C. The difference in the relative abundance of S-cysteinylated albumin (S-Cys-Alb) before and after an intentional incubation period that drives this proteoform to its maximum level is denoted as ΔS-Cys-Albumin. ΔS-Cys-Albumin in fully expired samples is zero. The range (mean ± 95% CI) observed for ΔS-Cys-Albumin in fresh cardiac patient P/S (n = 97) was, for plasma 12–29% (20.9 ± 0.75%) and for serum 10–24% (15.5 ± 0.64%). The multireaction rate law that governs S-Cys-Alb formation in P/S was determined and shown to predict the rate of formation of S-Cys-Alb in plasma and serum samples—a step that enables back-calculation of the time at which unknown P/S specimens have been exposed to room temperature. A blind challenge demonstrated that ΔS-Cys-Albumin can detect exposure of groups (n = 6 each) of P/S samples to 23 °C for 2 h, 4 °C for 16 h, or −20 °C for 24 h—and exposure of individual specimens for modestly increased times. An unplanned case study of nominally pristine serum samples collected under NIH-sponsorship demonstrated that empirical evidence is required to ensure accurate knowledge of archived P/S biospecimen storage history.

3D echo imaging of the MV allows direct visualization and planimetry of the ROA in patients with severe MR with good agreement to flow-based proximal convergence measurements.