Unprocessed Plasma Samples Research Articles

Electrochemical sensing of doxorubicin hydrochloride under sodium alginate antifouling conditions using silver nanoparticles modified glassy carbon electrodes

Doxorubicin (DOX) is a highly effective anticancer drug with a narrow therapeutic window; thus, sensitive and timely detection of DOX is crucial. Using electrodeposition of silver nanoparticles (AgNPs) and electropolymerization of alginate (Alg) layers on the surface of a glassy carbon electrode, a novel electrochemical probe was constructed (GCE). The fabricated AgNPs/poly-Alg-modified GCE probe was utilized for the quantification of DOX in unprocessed human plasma samples. For the electrodeposition of AgNPs and electropolymerization of alginate (Alg) layers on the surface of GCE, cyclic voltammetry (CV) was used in the potential ranges of −2.0 to 2.0 V and −0.6 to 0.2 V, respectively. The electrochemical activity of DOX exhibited two oxidation processes at the optimum pH value of 5.5 on the surface of the modified GCE. The DPV spectra of poly(Alg)/AgNPs modified GCE probe toward consecutive concentrations of DOX in plasma samples demonstrated wide dynamic ranges of 15 ng/mL-0.1 μg/mL and 0.1–5.0 μg/mL, with a low limit of quantification (LLOQ) of 15 ng/mL. The validation results indicated that the fabricated electrochemical probe might serve as a highly sensitive and selective assay for the quantification of DOX in patient samples. As an outstanding feature, the developed probe could detect DOX in unprocessed plasma samples and cell lysates without the requirement for pretreatment.

A ratiometric electrochemical probe for the quantification of apixaban in unprocessed plasma samples using carbon aerogel/BFO modified glassy carbon electrodes.

A novel electrochemical probe was established for the quantification of apixaban (APX) in unprocessed plasma samples. Efficiently oxidized graphene oxide aerogels (EEGO-AGs) and nano-sized Bi2Fe4O9 (BFO) particles were electrodeposited on the surface of a glassy carbon electrode (GCE). In this work, a ratiometric electrochemical method was introduced for APX detection to enhance the specificity of the probe in plasma samples. The fabricated ratiometric probe was employed for the indirect detection determination of APX using K3[Fe(CN)6]/K4[Fe(CN)6] as the redox pair. The differential pulse voltammetry technique was used to record the current alteration of the BFO/EEGO-AG-functionalized GCE probe at various APX concentrations. The probe response was proportional to the APX concentrations from 10 ng mL-1 to 10 μg mL-1 with a low limit of quantification (LLOQ) of 10 ng mL-1. After validation, this method was successfully utilized for the determination of APX in patients' plasma samples who have taken APX regularly. The fabricated chemosensor detected APX concentrations in unprocessed plasma samples with high selectivity, resulting from the physical filtering antifouling activity of aerogels.

Electrochemiluminescent detection of epilepsy biomarker miR-134 using a metal complex light switch

The detection of a key biomarker in epilepsy, miR-134, using an environmentally sensitive electrochemiluminescent luminophore, [Ru(DPPZ)2 PIC]2+, is reported, DPPZ is dipyrido[3,2-a:2′,3′-c]phenazine) and PIC is (2,2′-bipyridyl)-2(4-carboxy phenyl) imidazo [4,5][1,10] phenanthroline. A thiolated capture strand is first labelled with [Ru(DPPZ)2 PIC]2+ and then adsorbed onto a gold electrode. No significant electrochemiluminescence, ECL, is observed for immobilised Ru-labelled capture strands which is consistent with the light-switch dye being exposed to the aqueous solution. In sharp contrast, binding of the target turns on ECL. The ECL intensity, IECL, depends on the number of adenine “spacer” bases between the end of the capture sequence and the dye. The ECL intensity for the optimised system increases linearly with increasing miR-134 concentration from 100 nM to approximately 20 μM. Single and double base mismatches produce IECL that are only approximately 30% and 8% respectively of that observed for the fully complementary target reflecting differences in their association constants. Significantly, the presence of BSA protein causes IECL to increase by less 5% in either the single or duplex circumstances. Finally, the ability of the sensor to quantify miR-134 in unprocessed plasma samples from healthy volunteers and people with epilepsy is reported.

Development of LC-MS/MS method and application to bioequivalence study of a light sensitive drug montelukast

Objective The aim of the study was to develop a simple, highthroughput and sensitive LC-MS/MS method and apply to a bioequivalence study of montelukast, a light sensitive drug. Method The effects of organic modifiers in mobile phase, protein precipitation agent to plasma sample ratio, and light on montelukast stability in unprocessed and processed human plasma, were evaluated. Validation was conducted in accordance with European Medicines Agency Guideline on bioanalytical method validation. Results No interference peak was observed when acetonitrile was used as an organic modifier. Acetonitrile to plasma ratio of 4:1 produced clean plasma sample. Approximately 3 % of cis isomer was detected in unprocessed plasma samples while 21 % of cis isomer was detected in processed plasma samples after exposing to fluorescent light for 24h. The standard calibration curve was linear over 3.00–1200.00 ng/mL. All method validation parameters were within the acceptance criteria. Conclusion The validated method was successfully applied to a bioequivalence study of two montelukast formulations involving 24 healthy Malaysian volunteers. The light stability of a light sensitive drug in unprocessed and processed human plasma samples should be studied prior to pharmacokinetic/bioequivalence studies. Measures could then be taken to protect the analyte in human plasma from light degradation.

Direct Enhanced Detection of Multiple Circulating Tumor DNA Variants in Unprocessed Plasma by Magnetic-Assisted Bioelectrocatalytic Cycling.

The detection of single-nucleotide variants (SNVs) in circulating tumor DNA (ctDNA) in liquid biopsies has increasingly been shown to exhibit unique benefits for early detection or minimal residual disease monitoring in cancer. Yet, current clinically validated assays for ctDNA SNV detection are challenged by (i) time-consuming and laborious spin column-based ctDNA purification protocols, (ii) limited detection specificity to discriminate between mutated SNVs from large excess of closely similar wild-type sequences, and (iii) insufficient detection sensitivity required for trace ctDNA target analysis in blood. Herein, a ctDNA assay is demonstrated to tackle these triple key issues by fusing magnetics for quick ctDNA enrichment directly from unprocessed blood, selected bioenzyme activities for rapid discrimination, and molecular amplification of target SNVs, and designed magnetic-assisted bioelectrocatalytic cycling of DNA-intercalating and freely diffusing redox probes for electrochemical signal intensification. The described ctDNA SNV assay enables the detection of clinically relevant ctDNA SNVs in melanoma (BRAFV600E, KITL576P, and NRASQ61K) from unprocessed plasma samples with unprecedented 0.005% detection sensitivity, ultrabroad dynamic range over four orders of magnitude, and excellent single-base specificity.

Efficient Capture and Isolation of Tumor-Related Circulating Cell-Free DNA from Cancer Patients Using Electroactive Conducting Polymer Nanowire Platforms.

Circulating cell-free DNA (cfDNA) is currently recognized as a key non-invasive biomarker for cancer diagnosis and progression and therapeutic efficacy monitoring. Because cfDNA has been detected in patients with diverse types of cancers, the use of efficient strategies to isolate cfDNA not only provides valuable insights into tumour biology, but also offers the potential for developing new cancer-specific targets. However, the challenges associated with conventional cfDNA extraction methods prevent their further clinical applications. Here, we developed a nanostructured conductive polymer platform for the efficient capture and release of circulating cfDNA and demonstrated its potential clinical utility using unprocessed plasma samples from patients with breast and lung cancers. Our results confirmed that the platform's enhanced efficiency allows tumor-specific circulating cfDNA to be recovered at high yield and purity.

Investigating the Use of Protein Saver Cards for Storage and Subsequent Detection of Bovine Anti-Brucella abortus Smooth Lipopolysaccharide Antibodies and Gamma Interferon

Brucella abortus, a smooth strain of the genus Brucella, is the causative agent of bovine brucellosis. To support the ongoing development of diagnostic tests for bovine brucellosis, the use of Protein Saver cards (Whatman) for bovine blood serum and plasma sample collection has been evaluated. These cards offer significant logistical and safety alternatives to transporting and storing liquid samples and may aid in diagnostic programs and validation studies. To evaluate the utility of these cards, 204 bovine blood serum samples from Brucella-infected and noninfected animals were stored on and eluted from the Protein Saver cards. Anti-Brucella smooth lipopolysaccharide (sLPS) antibody titers for the serum eluates were compared to those of the unprocessed original serum samples by indirect enzyme-linked immunosorbent assay (ELISA). The results showed a highly significant correlation between titers from the serum eluates and the unprocessed sera. Therefore, under these circumstances, serum eluates and unprocessed serum samples may be used interchangeably. Blood plasma from 113 mitogen-stimulated whole-blood samples was added to and eluted from the Protein Saver cards. The gamma interferon (IFN-γ) titers in the plasma eluates were compared to those of the unprocessed plasma samples obtained by IFN-γ ELISA. The results showed a significant correlation between the plasma eluates and the unprocessed plasma samples. To derive a signal in the plasma eluate, it was necessary to develop a novel and highly sensitive ELISA for the detection of IFN-γ. The serum samples stored on cards at room temperature over a 10-day period showed little variation in antibody titers. However, the plasma eluates showed a progressive loss of IFN-γ recovery over 10 days when stored at room temperature.

The effect of pulse high-density plasma on superconducting properties of Bi-2223 multilayered tape

The effect of pulse high-power nonlinear shock waves generated at the Plasma Focus installation, which was created according to the nuclear fusion program, on some characteristics of multilayered high-temperature superconducting (HTSC) Bi-2223 tape is demonstrated. The tape is produced using the technique of HTSC powder rolling in a silver coating. A more than a twofold increase in the critical current in zero magnetic fields and external magnetic fields from 0.5 to 6 T at a temperature of 77 K is shown to take place under the action of shock waves at room temperature. The volt-ampere characteristics (VACs) of HTSC tape samples exposed to the effect of shock waves are found to drastically differ from the characteristics of the initial tapes. The shock waves suppress the “gear”-type VAC character of unprocessed plasma samples, their jump amplitudes increasing with an increase in the applied magnetic field intensity.

Internal-surface reversed-phase chromatography for plasma metabolite analysis of radiopharmaceuticals

The use of internal-surface reversed-phase (ISRP) chromatography of unprocessed plasma samples was investigated as an alternative method of quantitation of the arterial plasma metabolite time course of [ 18F] N-methylspiperone. The ISRP method was directly compared to standard solid phase extraction/HPLC (SPE/HPLC) methods currently in wide use. Results indicate that: (1) the ISRP method is rapid and minimizes sample preparation; (2) recovery of radioactivity from the ISRP column is > 90%; (3) no radioactivity remains associated with chromatographically size excluded proteins and (4) the quantitative results are well correlated with conventional SPE/HPLC methods.