Optimal Protocol Research Articles

Open label clinical trial of orally administered molnupiravir as a first-line treatment for naturally occurring effusive feline infectious peritonitis.

Before the discovery of effective antiviral drugs, feline infectious peritonitis (FIP) was a uniformly fatal disease of cats. Multiple antiviral treatments have been recognized, but optimization of treatment protocols is needed. To evaluate the efficacy of PO molnupiravir (MPV; EIDD-2801) to treat effusive FIP. Ten cats with naturally occurring effusive FIP and 10 historical control cats with effusive FIP treated with PO GS-441524. A single-center, prospective, open-label longitudinal, non-inferiority trial with historical controls. Ten cats with FIP were enrolled and treated with PO MPV (10-15 mg/kg PO q12h) for 84 days. Cats were evaluated at 0, 6, and 16 weeks, and the proportion of cats in clinical remission at 16 weeks was determined. Survival and clinicopathologic features were compared with historical control cats with effusive FIP treated with PO GS-441524. Eight of the 10 cats treated with MPV survived and were in remission at 16 weeks. The 2 non-survivors died in the first 24 hours of treatment. No adverse events that necessitated discontinuation of treatment were observed. Survival of cats treated with PO MPV was non-inferior to historic control cats treated with PO GS-441524 (5/9 [55%] survived), with a difference in survival of 25% (90% confidence interval, -9.3% to 59.3%). Clinicopathologic features associated with FIP normalized during the study period, and no differences in clinicopathologic data at each study time point were observed when comparing cats treated with MPV and GS-441524. Molnupiravir is an effective antiviral treatment for effusive FIP.

Challenges and solutions in FISH for formalin-fixed paraffin-embedded tissue: A scoping review.

Fluorescence in situ hybridization (FISH) has revolutionized molecular cytogenetic analysis since the 1980s, enabling precise localization of DNA sequences in cells and tissues. Despite its relevance, applying FISH to formalin-fixed paraffin-embedded (FFPE) tissue samples encounters significant technical challenges. This review addresses the main issues encountered in this context, such as inadequate fixation, contamination, block and slide age, inadequate pretreatment, and FISH technique. Proposed solutions include optimized pretreatment protocols, monitoring of blockage, careful selection of probes, and thorough analysis of results. Implementing good laboratory practices and quality control strategies are essential to ensure reliable results. Additionally, the use of emerging technologies such as artificial intelligence and digital pathology offers new perspectives for improving the efficiency and accuracy of FISH in FFPE samples. This review highlights the importance of a careful and personalized approach to overcome the technical challenges associated with FISH in FFPE samples, strengthening its role in research and clinical diagnosis. RESEARCH HIGHLIGHTS: Few FISH studies on FFPE: The scarcity of studies specifically addressing FISH applications in FFPE tissues highlights a critical gap in the literature. Troubleshooting FISH in FFPE tissues: Identifying and addressing common challenges in FISH techniques when applied to FFPE samples, such as signal quality and hybridization efficiency. Critical aspects of FISH technique: Discuss the main technical considerations crucial for successful FISH in FFPE tissues, including sample preparation, probe selection, and protocol optimization.

T1234: A distortion-matched structural scan solution to misregistration of high resolution fMRI data.

High-resolution fMRI at 7T is challenged by suboptimal alignment quality between functional data and structural scans. This study aims to develop a rapid acquisition method that provides distortion-matched, artifact-mitigated structural reference data. We introduce an efficient sequence protocol termed T1234, which offers adjustable distortions. This approach involves a T1-weighted 2-inversion 3D-EPI sequence with four spatial encoding directions optimized for high-resolution fMRI. A forward Bloch model was used for T1 quantification and protocol optimization. Twenty participants were scanned at 7T using both structural and functional protocols to evaluate the utility of T1234. Results from two protocols are presented. A fast distortion-free protocol reliably produced whole-brain segmentations at 0.8mm isotropic resolution within 3:00-3:40 minutes. It demonstrates robustness across sessions, participants, and three different 7T SIEMENS scanners. For a protocol with geometric distortions that matched functional data, T1234 facilitates layer-specific fMRI signal analysis with enhanced laminar precision. This structural mapping approach enables precise registration with fMRI data. T1234 has been successfully implemented, validated, and tested, and is now available to users at our center and at over 50 centers worldwide.

An Automated Analysis of Homocoupling Defects Using MALDI-MS and Open-Source Computer Software.

Conjugated organic polymers have substantial potential for multiple applications but their properties are strongly influenced by structural defects such as homocoupling of monomer units and unexpected end-groups. Detecting and/or quantifying these defects requires complex experimental techniques, which hinder the optimization of synthesis protocols and fundamental studies on the influence of structural defects. Mass spectrometry offers a simple way to detect these defects but a manual analysis of many complex spectra is tedious and provides only approximate results. In this work, we develop a computational methodology for analyzing complex mass spectra of organic copolymers. Our method annotates spectra similarly to a human expert and provides quantitative information about the proportions of signal assigned to each ion. Our method is based on the open-source Masserstein algorithm, which we modify to handle large libraries of reference spectra required for annotating complex mass spectra of polymers. We develop a statistical methodology to analyze the quantitative annotations and compare the statistical distributions of structural defects in polymer chains between samples. We apply this methodology to analyze commercial and lab-made samples of a benchmark polymer and show that the samples differ both in the amount and in the types of structural defects.

Evaluating the Knowledge of Radiation Exposure Parameters Among CT Technologists in Saudi Arabia: A Survey-Based Study on Adherence to ALARA Principles and Dose Optimization

Computed tomography is a crucial tool in modern healthcare, offering non-invasive imaging for diagnosis and monitoring. However, its widespread use raises concerns about radiation exposure. Adhering to the ALARA principle is vital to minimize risks while maintaining diagnostic quality. This study evaluates the knowledge of CT technologists in Saudi Arabia regarding radiation exposure parameters. A questionnaire-based survey was conducted, covering demographics, perceptions, and understanding of CT parameters. Statistical analysis revealed gaps in knowledge, particularly regarding dose optimization strategies and diagnostic reference levels. While most participants recognized the importance of adjusting parameters based on patient characteristics and clinical indications, deficiencies were observed in understanding specific parameters' effects on dose and image quality. Furthermore, a notable proportion of participants were unfamiliar with DRLs, indicating a need for improved education in this area. The study emphasizes the importance of interdisciplinary collaboration between radiologists, physicists, and technologists in protocol development and optimization.

Regulatory cell therapy for kidney transplantation and autoimmune kidney diseases.

Regulatory cell therapies, including regulatory T cells and mesenchymal stromal cells, have shown promise in early clinical trials for reducing immunosuppression burden in transplantation. While regulatory cell therapies may also offer potential for treating autoimmune kidney diseases, data remains sparse, limited mainly to preclinical studies. This review synthesises current literature on the application of regulatory cell therapies in these fields, highlighting the safety and efficacy shown in existing clinical trials. We discuss the need for further clinical validation, optimisation of clinical and immune monitoring protocols, and the challenges of manufacturing and quality control under Good Manufacturing Practice conditions, particularly for investigator-led trials. Additionally, we explore the potential for expanding clinical indications and the unique challenges posed in paediatric applications. Future directions include scaling up production, refining protocols to ensure consistent quality across manufacturing sites, and extending applications to other immune-mediated diseases.

Optimization of lipofection protocols for CRISPR/Cas9 delivery in porcine zona pellucida intact oocytes: A study of coincubation duration and reagent efficacy

A priority to facilitate the application of lipofection to generate genetically modified porcine embryos and animals will be the use of zona pellucida (ZP)-intact oocytes and zygotes. Recently, our group produced genetically modified embryos by lipofection of ZP-intact oocytes during in vitro fertilization (IVF). This study investigates the effect of two commercial lipofection reagents, Lipofectamine 3000 and Lipofectamine CRISPRMAX, on embryo development and mutation efficiency in ZP-intact porcine oocytes. We compared these reagents with the electroporation method and a control group using two sgRNAs targeting the CAPN3 and CD163 genes. The detrimental effects on cleavage rates were observed in both lipofection treatments compared to the control and electroporated groups. However, blastocyst rates were higher in the Lipofectamine 3000 group than in the electroporated group for both genes. Mutation parameters varied by target gene, with Lipofectamine 3000 achieving higher mutation rates for CD163, while all groups were similar for the CAPN3 gene. Overall efficiency was similar for both lipofectamines, confirming their feasibility for use. In addition, we evaluated the effect of coincubation time (4, 8, and 24 h) on IVF outcomes, embryo development, and mutation parameters. Results indicated that an 8-h coincubation period optimized fertilization and mutation efficiency without significant toxic effects. This study demonstrates that lipofection with either Lipofectamine 3000 or CRISPRMAX during IVF is an effective method for generating genetically modified porcine embryos without the need for specialized equipment or trained personnel, with efficiencies similar to or greater than electroporation. This study also highlights the importance of optimizing reagent selection and coincubation times. There is no difference between Lipofectamine 3000 and CRISPRMAXTM in terms of embryo development and mutation efficiency, and under our experimental conditions, the optimal coincubation time with lipofectamine is 8 h.

Impact of Uniaxial Static Strain on Myoblast Differentiation in Collagen-Coated PCL Microfilament Scaffolds: Role of Onset Time of Mechanical Stimulation.

Tissue engineering endeavors to create in vitro constructs that replicate the properties of native tissue, such as skeletal muscle. This study investigated the use of mechanical stimulation to promote myogenic differentiation and enhance the functionality of bioengineered tissues. Specifically, it aimed to facilitate the differentiation of myoblasts within a three-dimensional scaffold using a defined pattern of mechanical stimulation. C2C12 cells were cultured on a collagen-coated PCL microfilament scaffold and subjected to 24 h of uniaxial static strain using a biomechanical stimulation system. Two onset times of stimulation, 72 h and 120 h post-seeding, were evaluated. Cell proliferation, myogenic marker expression, and alterations in cell morphology and orientation were assessed. Results indicate that static strain on the scaffold promoted myoblast differentiation, evidenced by morphological and molecular changes. Notably, strain initiated at 72 h induced an early differentiation stage marked by MyoD expression, whereas stimulation beginning at 120 h led to a mid-stage differentiation characterized by the co-expression of MyoD and Myogenin, culminating in myotube formation. These results highlight the critical influence of myoblast maturity at the time of strain application on the differentiation outcome. This study provides insights that could guide the optimization of mechanical stimulation protocols in tissue engineering applications.

Addition of Cryoprotectant DMSO Reduces Damage to Spermatozoa of Yellow Catfish (Pelteobagrus fulvidraco) during Cryopreservation: Ultrastructural Damage, Oxidative Damage and DNA Damage.

Spermatozoa cryopreservation protocols have been established for yellow catfish (Pelteobagrus fulvidraco), but cryopreservation can still cause cellular damage and affect spermatozoa viability and fertility. Therefore, the aim of this paper was to evaluate the effects of adding or not adding cryoprotectants during low-temperature storage on the ultrastructural damage, oxidative damage, and DNA damage of thawed yellow catfish spermatozoa. The mixed semen of three male yellow catfish was divided into a fresh spermatozoa group, a frozen spermatozoa group (DMSO+) with a cryoprotectant (10% DMSO), and a frozen spermatozoa group without a cryoprotectant (DMSO-). Ultrastructural of the spermatozoa after thawing were observed under an electron microscope and the spermatozoa were assayed for SOD, MDA, and T-AOC enzyme activities, as well as for DNA integrity. In terms of movement parameters, compared with DMSO-, the addition of DMSO has significantly improved sperm motility, curve line velocity (VCL), and straight line velocity (VSL). The ultrastructural results showed that although thawed spermatozoa exhibited increased damage than fresh spermatozoa, 10% DMSO effectively reduced the damage to the plasma membrane, mitochondria, and flagellum of spermatozoa by cryopreservation, and most of the spermatozoa were preserved with intact structure. The results of oxidative damage showed that compared with frozen spermatozoa, 10% DMSO significantly increased the activities of SOD and T-AOC enzymes and clearly reduced the activity of the MDA enzyme. The antioxidant capacity of spermatozoa was improved, lipid peroxidation was reduced, and the oxidative damage caused by cryopreservation was mitigated. The DNA integrity of spermatozoa showed that 10% DMSO clearly reduced the DNA fragmentation rate. In conclusion, 10% DMSO can effectively reduce the ultrastructural damage, oxidative damage, and DNA damage of yellow catfish spermatozoa during cryopreservation; it can also further optimize the cryopreservation protocol for yellow catfish spermatozoa. Meanwhile, it also provides a theoretical basis for the future optimization of the cryopreservation protocols.

Time-asymmetric fluctuation theorem and efficient free-energy estimation.

The free-energy difference ΔF between two high-dimensional systems is notoriously difficult to compute but very important for many applications such as drug discovery. We demonstrate that an unconventional definition of work introduced by Vaikuntanathan and Jarzynski (2008) satisfies a microscopic fluctuation theorem that relates path ensembles that are driven by protocols unequal under time reversal. It has been shown before that counterdiabatic protocols-those having additional forcing that enforces the system to remain in instantaneous equilibrium, also known as escorted dynamics or engineered swift equilibration-yield zero-variance work measurements for this definition. We show that this time-asymmetric microscopic fluctuation theorem can be exploited for efficient free-energy estimation by developing a simple (i.e., neural-network free) and efficient adaptive time-asymmetric protocol optimization algorithm that yields ΔF estimates that are orders of magnitude lower in mean squared error than the generic linear interpolation protocol with which it is initialized.

Optimization of a Self-management Program for Chronic Musculoskeletal Pain – Protocol of the Pain Education Program Optimization (PEPO) Randomized Controlled Trial

OBJECTIVES: The primary objective is to compare two modes of delivering an 8-week self-management program for chronic musculoskeletal (MSK) pain: standard self-management versus supported self-management, which includes two physiotherapist-led sessions and one patient-partner-led session. The primary outcome measure is pain-related disability after 10 weeks. Secondary objectives include comparing the effects of both delivery modes on pain-related disability after 26 weeks, as well as on pain severity, kinesiophobia, catastrophizing, pain self-efficacy, anxiety, and depression after 10 and 26 weeks. DESIGN: Randomized controlled trial protocol. METHODS: One hundred eight adults with chronic MSK pain will be recruited and evaluated at baseline and after 10 and 26 weeks. At each evaluation, primary (pain-related disability) and secondary (pain severity, kinesiophobia, catastrophization, pain self-efficacy, anxiety, satisfaction, and depression) outcomes will be assessed. After baseline, participants will be randomly assigned to (1) self-management group and (2) supported self-management group. In addition to the 8-week self-management program, participants in the supported self-management group will have two additional 30-minute counselling sessions with a physiotherapist and one 60-minute group session led by a peer living with chronic MSK pain. Linear mixed models will be used to compare the effects of the two delivery modes of the intervention on the outcomes. JOSPT Open 2024;2(4):282-288. Epub 10 September 2024. doi:10.2519/josptopen.2024.0070

Optimization of an efficient micropropagation protocol for Persicaria glabra by the synergistic effect of ancymidol and benzyladenine

Optimization of an efficient micropropagation protocol for Persicaria glabra by the synergistic effect of ancymidol and benzyladenine

Bio-inspired wireless sensor networks - a protocol for an enhanced hybrid energy optimization routing

Recently, there has been a focus on the significance of swarm intelligence-inspired routing algorithms for achieving optimum solutions in biologically inspired wireless sensor networks (WSNs). These protocols depict a network of wireless mobile nodes forming an infrastructure that is agile, dynamic, and independent of a central administrative facility. Among the challenges faced by bio-inspired WSNs, mobility awareness and excessive energy consumption (EC) stand out as significant hurdles, particularly in dynamic models with intermittent connections. This project seeks to tackle these obstacles by deploying the hybrid energy efficiency (HEED) approach to distributed clustering for network system cluster formation, along with fusion routing protocol of particle swarm optimization (PSO) and PIO to select cluster-heads and optimize solutions in bio-inspired WSNs. The success of the suggested approach is assessed using a variety of criteria, such as energy usage, rate of packet delivery, EC, and routing overhead and network lifetime. The methods like ad hoc on-demand distance vector's (AODV) and ant colony optimization (ACO) methods are employed in the testing and validation. In comparison to the reactive AODV routing protocol and ACO, the suggested routing protocol (HPSOPIO) reduces energy usage and increases network lifespan.

An In-depth Analysis of the Adverse Effects of Ionizing Radiation Exposure on Cardiac Catheterization Staffs.

Diagnostic and interventional angiograms are instrumental in the multidisciplinary approach to CAD management, enabling accurate diagnosis and effective targeted treatments that significantly enhance patient care and cardiovascular outcomes. However, cath lab staff, including interventional cardiologists, is consistently exposed to ionizing radiation, which poses inherent health risks. Radiation exposure in the cath lab primarily results from the use of fluoroscopy and cineangiography during diagnostic and interventional procedures. Understanding these risks and implementing effective radiation protection measurements are imperative to ensure the well-being of healthcare professionals while delivering high-quality cardiac care. Prolonged and repeated exposure can lead to both deterministic and stochastic effects. Deterministic effects, such as skin erythema and tissue damage, are more likely to occur at high radiation doses. Interventional cardiologists and staff may experience these effects when safety measures are not rigorously followed. In fact, while ionizing radiation is essential in the practice of radiation cardiology ward, cath lab staff faces inherent risks from radiation exposure. Stochastic effects, on the other hand, are characterized by a probabilistic relationship between radiation exposure and the likelihood of harm. These effects include the increased risk of cancer, particularly for those with long-term exposure. Interventional cardiologists, due to their frequent presence in the cath lab, face a higher lifetime cumulative radiation dose, potentially elevating their cancer risk. Protective measures, including the use of lead aprons, thyroid shields, and radiation monitoring devices, play a crucial role in reducing radiation exposure for cath lab personnel. Adherence to strict dose optimization protocols, such as minimizing fluoroscopy time and maximizing distance from the radiation source, is also essential in mitigating these risks. Ongoing research and advancements in radiation safety technology are essential in further for minimizing the adverse effects of ionizing radiation in the cath lab.

A Guide to Environmental DNA Extractions for Non‐Molecular Trained Biologists, Ecologists, and Conservation Scientists

ABSTRACTEcologists, biologists, and conservation scientists are increasingly interested in the use of environmental DNA (eDNA) data for research and potentially decision‐making. While commercial DNA extraction kits are typically user‐friendly and accessible, they may fail to deliver the desired results with inherently complex eDNA samples, necessitating protocol optimization or educated selection of alternative approaches. To this end, knowledge of the basic steps and principles of DNA extractions is essential, but traditional education tracks in ecology, conservation, and environmental management typically do not include in‐depth training in molecular methods. The primary objective of this paper is to enable scientists with an ecological background and limited molecular training to understand the four key steps of eDNA isolations, and to use this expertise to their advantage. We describe the purpose of commonly used reagents and chemicals, point out alternatives for each key step, explain the impact of certain choices regarding isolation approaches on DNA integrity and purity, and highlight the possibility of a tailor‐made “mix and match” approach. We anticipate that this paper will enable field ecologists to develop a deeper understanding of the mechanisms and chemistry underlying eDNA extractions, thus allowing them to make informed decisions regarding the best eDNA extraction method for their research goals. Our intention is not to provide comprehensive, step‐by‐step protocols, but to offer guiding principles while highlighting alternative solutions. Finally, we hope that this paper will act as a useful resource to support knowledge transfer and teaching.

Advancing Fruit Preservation: Ecofriendly Treatments for Controlling Fruit Softening

Textural softening is a major factor that limits the storage potential of fruit. Fresh produce markets incur severe financial losses due to excessive fruit softening. The application of preservation strategies aimed at mitigating fruit softening is crucial for optimising the marketability of fruit. Proposed preservation strategies include ecofriendly treatments, namely, hexanal, edible coatings, heat treatments, ozone and UV-C irradiation. These treatments optimise firmness retention by targeting the factors that affect fruit softening, such as ethylene, respiration rates, enzymes and pathogens. This review discusses the mechanisms by which ecofriendly treatments inhibit fruit softening, providing insights into their effect on ethylene biosynthesis, cell wall metabolism and disease resistance. Although ecofriendly treatments offer a promising and sustainable approach for delaying fruit softening, the optimisation of treatment application protocols is needed to improve their efficacy in retaining fruit firmness. Studies reporting on the molecular mechanisms by which ecofriendly treatments inhibit fruit softening are limited. Future studies should prioritise proteomic and transcriptome analyses to advance our understanding of the underlying molecular mechanisms by which ecofriendly treatments delay the fruit-softening process.

Survey of CT radiation doses and iodinated contrast medium administration: an international multicentric study.

To assess the relationship between intravenous iodinated contrast media (ICM) administration usage and radiation doses for contrast-enhanced (CE) CT of head, chest, and abdomen-pelvis (AP) in international, multicenter settings. Our international (n = 16 countries), multicenter (n = 43 sites), and cross-sectional (ConRad) study had two parts. Part 1: Redcap survey with questions on information related to CT and ICM manufacturer/brand and respective protocols. Part 2: Information on 3,258 patients (18-96 years; M:F 1654:1604) who underwent CECT for a routine head (n = 456), chest (n = 528), AP (n = 599), head CT angiography (n = 539), pulmonary embolism (n = 599), and liver CT examinations (n = 537) at 43 sites across five continents. The following information was recorded: hospital name, patient age, gender, body mass index [BMI], clinical indications, scan parameters (number of scan phases, kV), IV-contrast information (concentration, volume, flow rate, and delay), and dose indices (CTDIvol and DLP). Most routine chest (58.4%) and AP (68.7%) CECT exams were performed with 2-4 scan phases with fixed scan delay (chest 71.4%; AP 79.8%, liver CECT 50.7%) following ICM administration. Most sites did not change kV across different patients and scan phases; most CECT protocols were performed at 120-140 kV (83%, 1979/2685). There were no significant differences between radiation doses for non-contrast (CTDIvol 24 [16-30] mGy; DLP 633 [414-702] mGy·cm) and post-contrast phases (22 [19-27] mGy; 648 [392-694] mGy·cm) (p = 0.142). Sites that used bolus tracking for chest and AP CECT had lower CTDIvol than sites with fixed scan delays (p < 0.001). There was no correlation between BMI and CTDIvol (r2 ≤ - 0.1 to 0.1, p = 0.931). Our study demonstrates up to ten-fold variability in ICM injection protocols and radiation doses across different CT protocols. The study emphasizes the need for optimizing CT scanning and contrast protocols to reduce unnecessary contrast and radiation exposure to patients. The wide variability and lack of standardization of ICM media and radiation doses in CT protocols suggest the need for education and optimization of contrast usage and scan factors for optimizing image quality in CECT. There is a lack of patient-centric CT protocol optimization taking into consideration mainly patients' size. There is a lack of correlation between ICM volume and CT radiation dose across CT protocol. A ten-fold variation in iodine-load for the same CT protocol in sites suggests a lack of standardization.

Human relevance of in vivo and in vitro skin irritation tests for hazard classification of pesticides

Background: Test methods to inform hazard characterization and labeling of pesticides to protect human health are typically conducted using laboratory animals, and for skin irritation/corrosion the rabbit Draize test is currently required by many regulatory agencies. Although the Draize test is generally regarded to provide protective classifications for human health, new approach methodologies (NAMs) have been developed that offer more human relevant models that circumvent the uncertainty associated with species differences that exist between rabbits and humans. Despite wide applicability and use of these test methods across a broad range of chemicals, they have not been widely adopted for testing pesticides and pesticidal formulations. One of the barriers to adoption of these methods in this sector is low concordance with results from the Draize rabbit test, particularly for chemicals within the mild to moderate irritation spectrum. Methods: This review compares and contrasts the extent to which available models used in skin irritation testing mimic the anatomy and physiology of human skin, and how each aligns with the known key events leading to chemically-induced adverse skin irritation and corrosion. Doing so fully characterizes the human relevance of each method. Results: As alternatives to the rabbit Draize test, several protocols using ex vivo, in chemico, and in vitro skin models are available as internationally harmonized test guidelines. These methods rely on a variety of models of human skin, including excised rodent skin, synthetic biochemical models of barrier function, cell culture systems, and reconstructed human tissue models. We find these models exhibit biological and mechanistic relevance aligned with human skin irritation responses. Further, recent retrospective analyses have shown that the reproducibility of the Draize test is less than 50% for mild and moderate responses, with many of the replicate predictions spanning more than one category (e.g., a moderate response reported in one study followed by a non-irritant response reported in another study). Conclusions: Based on this comparative evaluation, we recommend top-down and bottom-up testing strategies that use the most human relevant in vitro test methods for skin irritation and corrosion classification of pesticides and pesticide formulations. To further discriminate among mild and non-irritant formulations, optimization of a cytokine release protocol and subsequent analyses of reference formulation test results is recommended.

Radioimmunotheragnosis in Cancer Research.

The combination of immunoPET-where an antibody (Ab) is labeled with an isotope for PET imaging-and radioimmunotherapy (RIT), using the same antibody with a therapeutic isotope, offers significant advantages in cancer management. ImmunoPET allows non-invasive imaging of antigen expression, which aids in patient selection for subsequent radioimmunotherapy. It also facilitates the assessment of tumor response to therapy, allowing for treatment adjustments if necessary. In addition, immunoPET provides critical pharmacokinetic data, including antibody biodistribution and clearance rates, which are essential for dosimetry calculations and treatment protocol optimization. There are still challenges to overcome. Identifying appropriate target antigens that are selectively expressed on cancer cells while minimally expressed on normal tissues remains a major hurdle to reduce off-target toxicity. In addition, it is critical to optimize the pharmacokinetics of radiolabeled antibodies to maximize tumor uptake and minimize normal tissue uptake, particularly in vital organs such as the liver and kidney. This approach offers the potential for targeted and personalized cancer therapy with reduced systemic toxicity by exploiting the specificity of monoclonal antibodies and the cytotoxic effects of radiation. However, further research is needed to address remaining challenges and to optimize these technologies for clinical use.

Deep Plasma Proteomics with Data-Independent Acquisition: Clinical Study Protocol Optimization with a COVID-19 Cohort.

Plasma proteomics is a precious tool in human disease research but requires extensive sample preparation in order to perform in-depth analysis and biomarker discovery using traditional data-dependent acquisition (DDA). Here, we highlight the efficacy of combining moderate plasma prefractionation and data-independent acquisition (DIA) to significantly improve proteome coverage and depth while remaining cost-efficient. Using human plasma collected from a 20-patient COVID-19 cohort, our method utilizes commonly available solutions for depletion, sample preparation, and fractionation, followed by 3 liquid chromatography-mass spectrometry/MS (LC-MS/MS) injections for a 360 min total DIA run time. We detect 1321 proteins on average per patient and 2031 unique proteins across the cohort. Differential analysis further demonstrates the applicability of this method for plasma proteomic research and clinical biomarker identification, identifying hundreds of differentially abundant proteins at biological concentrations as low as 47 ng/L in human plasma. Data are available via ProteomeXchange with the identifier PXD047901. In summary, this study introduces a streamlined, cost-effective approach to deep plasma proteome analysis, expanding its utility beyond classical research environments and enabling larger-scale multiomics investigations in clinical settings. Our comparative analysis revealed that fractionation, whether the samples were pooled or separate postfractionation, significantly improved the number of proteins quantified. This underscores the value of fractionation in enhancing the depth of plasma proteome analysis, thereby offering a more comprehensive landscape for biomarker discovery in diseases such as COVID-19.