Capsular contracture (CC) is one of the most common postoperative complications associated with breast implant-associated infections. The mechanisms that lead to CC remain poorly understood. Plasma is an ideal biospecimen for early proteomics biomarker discovery. However, as high-abundance proteins mask signals from low-abundance proteins, identifying novel or specific proteins as biomarkers for a particular disease has been hampered. Here, we employed depletion of high-abundance plasma proteins followed by Tandem Mass Tag (TMT)-based quantitative proteomics to compare 10 healthy control patients against 10 breast implant CC patients. A total of 450 proteins were identified from these samples. Among them, 16 proteins were significantly differentially expressed in which 5 proteins were upregulated and 11 downregulated in breast implant CC patients compared to healthy controls. Gene Ontology enrichment analysis revealed that proteins related to cell, cellular processes and catalytic activity were highest in the cellular component, biological process, and molecular function categories, respectively. Further, pathway analysis revealed that inflammatory responses, focal adhesion, platelet activation, and complement and coagulation cascades were enriched pathways. The differentially abundant proteins from TMT-based quantitative proteomics have the potential to provide important information for future mechanistic studies and in the development of breast implant CC biomarkers.

A novel antipsychotic medication named brexpiprazole (BRX) is currently employed for the treatment of schizophrenia and other psychotic disorders. Because BRX\'s molecular structure includes a benzothiophene ring, it natively fluoresces. To detect BRX with precision and speed, a flow injection-fluorometric method, which is both sensitive and selective, is recommended. The fluorescence detection was conducted at 364 nm following excitation at 326 nm to capture the strong intrinsic fluorescence of BRX. The carrier solution employed was a mixture of phosphate buffer (pH 4, 10 mM) and acetonitrile (50: 50, v/v), with a flow rate of 0.5 mL min- 1. The calibration curve, based on peak areas, exhibited linearity within the concentration range of 20-350 ng mL- 1, with a remarkable correlation coefficient (r2) of 0.9999. The limit of quantitation was 9.7 ng mL- 1, and the limit of detection was found to be 3.2 ng mL- 1. This method was applied to quantify BRX in Neopression® tablets, achieving recovery within an acceptable range without interference from the tablet\'s additives. Additionally, the proposed approach was successfully utilised to quantify the drug in spiked human plasma. The approach underwent validation following ICH requirements.

Valproic acid and phenytoin are two prevalent antiepileptic medications known for their narrow indices and propensity for cardiovascular and respiratory system toxicity. Therefore, therapeutic drug monitoring (TDM) of valproic acid (VAL) and phenytoin (PHE) concentrations in patient plasma is extremely beneficial for improving clinical choices, avoiding adverse reactions, and optimizing treatment for individual patients. In this study, a rapid and sensitive ultra-performance liquid chromatographic tandem mass spectrometer (UPLC-MS/MS) method was developed and validated for the simultaneous quantitative determination of valproic acid (VAL) and phenytoin (PHE) in human plasma. Negative electron spray ionization (ESI-) mode with selective ion recording (SIR) was employed to determine the transitions of m/z 142.98 and m/z 250.93 for VAL and PHE, respectively. The internal standard (IS) betamethasone (BETA) was ionized using positive electron spray ionization (ESI+) and detected by multi-reaction monitoring (MRM) mode to obtain precursor ions and specific fragment ions for quantification, and the MRM transition was chosen to be m/z 393.17 → 355.16. The separation was performed using a Phenomenex Synergi Hydro-RP (4 μm, 250 × 4.6 mm, I.D.) with an isocratic mobile phase consisting of acetonitrile - water (75:25, v/v) at a flow rate of 0.8 mL/min. The column temperature was maintained at 25 °C. The lower limit of quantification of VAL and PHE was 3.6 μg/mL and 0.72 μg/mL, respectively, which resulted in a recovery of more than 85 % for most analytes. According to US-FDA bioanalytical technique validation, the specificity, intra- and inter-day precision and accuracy, matrix effect, carryover, dilution, and stability of all analytes were within acceptable ranges. This analytical method was successful in evaluating the levels of valproic acid and phenytoin in human plasma from epileptic patients.

UNASSIGNED: D-dimer at a low level is important evidence for excluding the onset and progression of thrombosis. It is readily detectable and yields rapid results, although significant variability exists among different detection systems. Our study aims to enhance the consistency across various detection systems.
UNASSIGNED: Twelve detection systems were included in our study. We sought to address this inconsistency by using various calibrators (two supplied by manufacturers and two comprising pooled human plasma diluted with different diluents) to standardize D-dimer measurements. We categorized the data into three groups according to D-dimer concentration levels: low (≤0.5 mg/L), medium (>0.5 mg/L - <3 mg/L), and high (≥3 mg/L). We then analyzed the data focusing on range, consistency, comparability, negative coincidence rate, and false negative rate.
UNASSIGNED: Calibrating with pooled human plasma led to narrower result ranges in the low and medium groups (P < 0.05). In the low group, consistency improved from weak to strong (ICC 0.4-0.7, P﹤0.05), while it remained excellent in the other groups and overall (ICC﹥0.75, P﹤0.05). The percentage of pairwise comparability increased in both the low and high groups. Additionally, there was an increase in the negative coincidence rate.
UNASSIGNED: These findings demonstrate that uniform calibration of D-dimer can significantly enhance the consistency of results across different detection systems.

BACKGROUND: Tau is aberrantly acetylated in various neurodegenerative conditions, including Alzheimer\'s disease, frontotemporal lobar degeneration (FTLD), and traumatic brain injury (TBI). Previously, we reported that reducing acetylated tau by pharmacologically inhibiting p300-mediated tau acetylation at lysine 174 reduces tau pathology and improves cognitive function in animal models.
METHODS: We investigated the therapeutic efficacy of two different antibodies that specifically target acetylated lysine 174 on tau (ac-tauK174). We treated PS19 mice, which harbor the P301S tauopathy mutation that causes FTLD, with anti-ac-tauK174 and measured effects on tau pathology, neurodegeneration, and neurobehavioral outcomes. Furthermore, PS19 mice received treatment post-TBI to evaluate the ability of the immunotherapy to prevent TBI-induced exacerbation of tauopathy phenotypes. Ac-tauK174 measurements in human plasma following TBI were also collected to establish a link between trauma and acetylated tau levels, and single nuclei RNA-sequencing of post-TBI brain tissues from treated mice provided insights into the molecular mechanisms underlying the observed treatment effects.
RESULTS: Anti-ac-tauK174 treatment mitigates neurobehavioral impairment and reduces tau pathology in PS19 mice. Ac-tauK174 increases significantly in human plasma 24 h after TBI, and anti-ac-tauK174 treatment of PS19 mice blocked TBI-induced neurodegeneration and preserved memory functions. Anti-ac-tauK174 treatment rescues alterations of microglial and oligodendrocyte transcriptomic states following TBI in PS19 mice.
CONCLUSIONS: The ability of anti-ac-tauK174 treatment to rescue neurobehavioral impairment, reduce tau pathology, and rescue glial responses demonstrates that targeting tau acetylation at K174 is a promising neuroprotective therapeutic approach to human tauopathies resulting from TBI or genetic disease.

Bacterial contamination is the most prevalent infectious complication of blood transfusion in the developed world. To mitigate this, several ultraviolet light-based pathogen reduction technologies (PRTs), some of which require photo-chemicals, have been developed to minimize infection transmission. Relative to UV light, visible 405-nm light is safer and has shown potential to be developed as a PRT for the in situ treatment of ex vivo human plasma and platelet concentrates, without the need for photo-chemicals. This study investigates the effect of 405-nm light on human plasma, with focus on the compatibility of antimicrobial light doses with essential plasma clotting factors. To determine an effective antimicrobial dose that is compatible with plasma, prebagged human plasma (up to 300 mL) was seeded with common microbial contaminants and treated with increasing doses of 405-nm light (16 mW cm-2; ≤ 403 J cm-2). Post-exposure plasma protein integrity was investigated using an AOPP assay, in vitro coagulation tests, and ELISA-based measurement of fibrinogen and Protein S. Microbial contamination in 300 mL prebagged human plasma was significantly reduced (P ≤ 0.05) after exposure to ≤ 288 J cm-2, with microbial loads reduced by > 96.2%. This dose did not significantly affect the plasma protein quality parameters tested (P > 0.05). Increased doses (≥ 345 J cm-2) resulted in a 4.3% increase in clot times with no statistically significant change in protein activity or levels. Overall, this study has demonstrated that the effective microbicidal 405 light dose shows little to no negative effect on plasma quality.

The technical difficulty of separating extracellular vesicles (EVs) from plasma proteins in human blood presents a significant hurdle in EV research, particularly during nano ultra-high-performance liquid chromatography-tandem mass spectrometric (UHPLC-MS/MS) analysis, where detecting \"vesicular\" proteins among abundant plasma proteins is challenging. Standardisation is a pressing issue in EV research, prompting collaborative global efforts to address it. While the MISEV guidelines offer valuable recommendations, unanswered questions remain, particularly regarding sample storage. We compared size exclusion chromatography (SEC) columns with pore sizes of 35 nm and 70 nm to identify fractions with minimal contaminating proteins and the highest concentration of small EVs (sEVs). Following column selection, we explored potential differences in the quality and quantity of sEVs isolated from platelet-free plasma (PFP) after long-term storage at -80 °C (>2.5 years) compared to freshly drawn blood. Our methodologically rigorous study indicates that prolonged storage, under correct storage and processing conditions, does not compromise sEV quality. Both columns effectively isolated vesicles, with the 70 nm column exhibiting a higher abundance of \"vesicular\" proteins. We propose a relatively rapid and moderately efficient protocol for obtaining a comparatively pure sEV fraction from plasma, facilitating sEV processing in clinical trials.

The simultaneous assay of duloxetine hydrochloride (DLX) and avanafil (AVN) in their pure forms, synthetic mixtures, and spiked human plasma was achieved using a novel, eco-friendly, sensitive, and specific HPTLC methodology that have been established and validated. Measuring the levels of co-administered antidepressants and sexual stimulants in biological fluids is an important step for individuals with depression and sexual problems. Separation was performed successfully using pre-coated silica gel 60-F254 as a stationary phase and a mobile phase composed of methanol, acetone, and 33% ammonia (8:2:0.05, v/v/v). Compact bands were produced by the optimized mobile phase that was chosen for development (Rf values were 0.23 and 0.75 for DLX and AVN, individually) after dual-wavelength detection for DLX and AVN at 232 and 253 nm, respectively. The results of polynomial regression analysis were exceptional (r = 0.9999 for both medicines) over concentration ranges of 5-800 and 10-800ng/spot for DLX and AVN, respectively. The quantitation limits were 4.69 and 9.53 ng/spot (0.31 and 0.94 µg/mL), whereas the detection limits were 1.55 and 3.15 ng/spot (0.63 and 1.91 µg/mL), for DLX and AVN, respectively. The International Council for Harmonization (ICH) criteria served as the basis for validating the established approach. Moreover, the proposed technique was evaluated in terms of greenness using four contemporary ecological metrics: The Analytical Greenness software (AGREE), the Green Analytical Procedure Index (GAPI), Eco-Scale, and the National Environmental Method Index (NEMI). Additionally, the Blue Applicability Grade Index (BAGI), a newly developed tool for evaluating the practicality (blueness) of procedures, was taken into consideration when evaluating the sustainability levels of the established approach.

Two sensitive, straightforward and repeatable chromatographic techniques were developed for the determination of Cytarabine HCl and Dexamethazone in their pure form and spiked human plasma without prior separation. The drugs are used co-administered for the treatment of Leukemia, a certain type of blood cancer. Method (A) is an isocratic chromatographic HPLC method; separation was accomplished on C18 column using the eluting mixture of 6.9 g/L Monobasic Sodium Phosphate pH 3: methanol (70:30, v/v) and detection was at 275 nm. Concentrations were in the range of 0.2-15 μg/mL for both CYT and DEX. Method (B) is a HPTLC method in which separation was attained on HPTLC F254 plates using methanol: ethyl acetate: ammonia, (7.8:2:0.2, by volume) as eluting solvents and detection was at 275 nm. Concentrations were in the range of 0.1-4 μg/band for both CYT and DEX. The parameters for system suitability testing were evaluated to determine the effectiveness of the developed chromatographic procedures in terms of performance. The recently developed techniques were applied for the determination of the drugs under investigation in spiked human plasma. Validation parameters were examined in accordance with US-FDA criteria. All results were found to be within the acceptable ranges. To evaluate the greenness characters of the proposed methods to the environment; three greenness assessment tools including eco-scale assessments (ESA), green analytical procedure index (GAPI), and Analytical Greenness calculator (AGREE) were used. Acceptable and satisfying results that demonstrated the greenness characteristics of the suggested methods were attained.

This study presents an efficient high-performance liquid chromatography with ultraviolet detection (HPLC-UV) method for monitoring valproic acid (VPA) level in human plasma. This method is distinguished by its simplicity, cost-effectiveness, and rapid execution, addressing the limitations associated with other advanced analytical techniques like liquid chromatography-mass spectrometry (LC-MS), gas chromatography-mass spectrometry (GC-MS), and immunoassays, which are generally complex and costly for routine application. A challenge in analyzing VPA is its non-linear protein binding profile and the absence of a chromophore in its structure, making direct detection difficult. To overcome this, the study developed an efficient HPLC-UV for VPA determination in human plasma, utilizing a simplified and rapid microwave-assisted derivatization process. Due to the lack of a chromophore in VPA structure, this work developed a microwave-assisted derivatization of VPA using phenylhydrazine hydrochloride (PH HCl). The process optimization was achieved at 450 W for 50 s, facilitating effective HPLC-UV detection. The derivatized product was characterized using 1H nuclear magnetic resonance (NMR) and Fourier transform infrared spectrometer (FT-IR). The derivative, identified as (Z)-N-phenyl-2-propylpentanehydrazonic acid, demonstrated specificity in plasma analysis with no detectable interference. The method exhibited a linear response for VPA concentrations ranging from 30 to 150 μg/mL, with a correlation coefficient exceeding 0.99. Recovery varied between 86.7% and 107%, with a maximum coefficient of variation (CV) of 10.0%. The findings suggest that the microwave-assisted derivatization technique substantially improves the feasibility and cost-effectiveness of HPLC-UV for the analysis of VPA in plasma. This method provides a viable alternative to conventional HPLC methodologies, offering a balance of efficiency and economic practicality for VPA quantification.