Analysis Of Human Blood Serum Research Articles

Novel Siloxane Derivatives as Membrane Precursors for Lactate Oxidase Immobilization.

We report new enzyme-containing siloxane membranes for biosensor elaboration. Lactate oxidase immobilization from water-organic mixtures with a high concentration of organic solvent (90%) leads to advanced lactate biosensors. The use of the new alkoxysilane monomers-(3-aminopropyl)trimethoxysilane (APTMS) and trimethoxy[3-(methylamino)propyl]silane (MAPS)-as the base for enzyme-containing membrane construction resulted in a biosensor with up to a two times higher sensitivity (0.5 A·M-1·cm-2) compared to the biosensor based on (3-aminopropyl)triethoxysilane (APTES) we reported previously. The validity of the elaborated lactate biosensor for blood serum analysis was shown using standard human serum samples. The developed lactate biosensors were validated through analysis of human blood serum.

Application of 96-well plate SPE method for analysis of persistent organic pollutants in low volume blood serum samples

Though many persistent organic pollutants (POPs) are closely regulated the human population is still exposed to these ubiquitous chemicals from the environment and diet. Safe management and human biomonitoring of POPs is necessary to understand the risk of exposure. Within human biomonitoring the mass of sample is often limited, therefore robust methods using smaller sample amounts are necessary. This study developed a 96-well plate solid phase extraction (SPE) method for determination of selected POPs: polychlorinated biphenyls (PCBs), organochlorine pesticides (OCPs), polybrominated diphenyl ethers (PBDEs), hexabromocyclododecane (HBCD) and non-persistent novel flame retardants (NFRs) in low volume blood serum. Non-destructive clean-up coupling Oasis HLB extraction plate with Phree phospholipid removal plate was employed. Extraction efficiency was determined at low and high concentrations in certified reference materials NIST SRM 1957 and 1958, respectively. Target compounds deviated from certified values on average by 15% and 21% for SRM 1957 and SRM 1958, respectively. Observed limit of detections (LODs) ranged from 0.36 pg/mL (PCB 180) to 66.07 pg/mL (δ-HCH). The applicability for real samples is demonstrated on 48 samples from pregnant women enrolled in the pilot phase of the CELSPAC: TNG study. In total, 30 target compounds were detected in at least one sample. The method developed here provides a fast and reliable analysis of human blood serum with possibility to introduce automation for the sample preparation procedure.

Estimation of Calcium and Iron Levels in Gingival Crevicular Fluid and Serum in Periodontal Health and Disease

Introduction: Gingival Crevicular Fluid (GCF) has been referred to as a promising medium for detection of markers for periodontal disease activity. Analysis of GCF shows minute changes in biomarker levels well before the onset of clinical signs and symptoms; which helps to even predict a person’s predisposition towards periodontal disease occurrence. The elemental analysis of human blood serum is noteworthy in routine clinical practice as well as in medical research. Aim: This study was done to determine the changes in calcium and iron levels in GCF and serum in human subjects with normal periodontal health and those with disease. Materials and Methods: This was a cross-sectional study conducted from March 2019 to December 2019. Eight study subjects (4 healthy subjects and 4 periodontitis cases) were selected from the patients reporting to the Department of Periodontics at Tagore Dental College and Hospital, Chennai. The subjects were chosen based on inclusion and exclusion criteria and all patients were subjected to a clinical examination wherein the Probing Depth (PD) and Clinical Attachment Level (CAL) were recorded by a single examiner using William’s Periodontal probe. The GCF samples were collected by Capillary Tubing method. Blood was collected by venipuncture and centrifuged to provide serum samples. Dual viewing Inductively Coupled Plasma Optical Emission Spectrometry (ICP-OES) was used to estimate Calcium and Iron in GCF and serum. SPSS version 21.0 was used for statistical analysis. Mann Whitney U test was used for comparing the groups. p-value less than 0.05 was considered statistically significant. Results: Serum iron levels were significantly less in periodontitis patients than healthy subjects (p-value 0.043). GCF iron level (p-value 0.386), GCF calcium level (p-value 0.149) and serum calcium level (p-value 0.564) did not show any major variation among subjects with normal periodontal health and those with disease. Conclusion: The findings of this study showed that iron and calcium are present in GCF and serum samples of healthy persons and patients with chronic periodontitis which can be detected using ICP-OES. A significant difference in serum iron levels between health and disease could indicate a patient’s predisposition towards developing periodontitis. Calcium levels in GCF and serum do not point towards periodontal disease activity.

Field-resolved infrared spectroscopy of biological systems

The proper functioning of living systems and physiological phenotypes depends on molecular composition. Yet simultaneousquantitative detection of a wide variety of moleculesremains a challenge1-8. Here we show how broadband optical coherence opens up opportunities for fingerprinting complex molecular ensembles in their natural environment. Vibrationally excited molecules emit a coherent electric field following few-cycle infrared laser excitation9-12, and this field is specific to the sample's molecular composition. Employing electro-optic sampling10,12-15, we directly measure this global molecular fingerprint down to field strengths 107 times weaker than that of the excitation. This enables transillumination of intact living systems with thicknesses of the order of 0.1 millimetres, permitting broadband infrared spectroscopic probing of human cells and plant leaves. In a proof-of-concept analysis of human blood serum, temporal isolation of the infrared electric-field fingerprint from its excitation along with its sampling with attosecond timing precision results in detection sensitivity of submicrograms per millilitre of blood serum and a detectable dynamic range of molecular concentration exceeding 105. This technique promises improved molecular sensitivity and molecular coverage for probing complex, real-world biological and medical settings.

Elemental Analysis of Human Blood Serum by Microwave Plasma\u2014Investigation of the Matrix Effects Caused by Sodium Using Model Solutions

Human blood is a complex sample matrix when elemental analysis is considered. In this study, the effects of Na, a natural component of serum samples, was investigated in the quantitative determination of Ca, K, Mg, Cu, Zn, and Fe by microwave plasma atomic emission spectrometry. The robustness of the microwave plasma was tested by evaluating MgII 280.271 nm/MgI 285.213 nm by varying two adjustable operating parameters, the read time, and the nebulizer pressure. The read time has no influence on the robustness while the MgII/MgI ratio decreased when the nebulizer pressure was increased during the analysis. The threshold concentrations of the interfering Na were determined at the analytical lines used for the measurement of other elements. The matrix effect of the commercially available microwave plasma was studied by a series of model experiments with human blood. The increasing concentration of Na in the matrix within the normal ranges reported for blood serum increased the intensities of the measured atomic lines. According to a factorial design—where two applied factors were the concentration of Na matrix and the measured elements as well as their levels were considered as factorial points—it was found that the Na concentration in a serum sample after acid digestion and 10 times dilution affected the intensity values of the measured elements. For Ca, Cu, and Fe, a statistically significant effect was observed, while for Zn, Mg, and K, an interaction effect was also found. However, after calculating the percentage errors caused by the shift, the relative difference was observed to be quite small (< 10%).

Phospholipid-protein balance in affective disorders: Analysis of human blood serum using Raman and FTIR spectroscopy. A pilot study

Raman and FTIR (Fourier Transform Infra Red) spectroscopies provide information on the chemical structure of compounds through identification and analysis of functional groups. In the present study, both spectroscopic techniques were used for investigating the phospholipid – protein balance in blood serum of depressed subjects (major depressive disorder and bipolar disorder type I or II) taking also into account their age and gender. The obtained results were compared with those of healthy subjects. The Raman and FTIR (using ATR (Attenuated Total Reflectance) technique), spectra show that a correlation between the level of phospholipids and proteins exists. Indeed, in depressed subjects the quantity of phospholipids and proteins is lower, compared to healthy ones. The second derivative of FTIR spectra shows that phospholipids directly affect the structure of proteins and their functions. In all male depressed subjects a higher amount of phospholipids and proteins compared to female depressed subjects was measured, offering them faster recovery perspectives. Spectroscopy results show that the phospholipids’ and proteins’ levels are lower in depressed subjects from 41 to 65 compared to the age group between 20 and 40, independently from the gender. Consequently, this study shows that Raman and infrared spectroscopies might be applied as a diagnostic tool to evaluate the balance between phospholipids and proteins in blood serum as a potential biomarker in depressive disorders.

Porous membrane strip microsampling: a dried biofluid collection format and application for quantitative enzyme immunoassay

Applicability of a new and simple membrane-strip microsampling format for the analysis of human blood serum in a strip-dried form for the presence of a range of model low and high molecular weight analytes by ELISA was demonstrated.

Integration of a highly ordered gold nanowires array with glucose oxidase for ultra-sensitive glucose detection

A highly sensitive amperometric nanobiosensor has been developed by integration of glucose oxidase (GOx) with a gold nanowires array (AuNWA) by cross-linking with a mixture of glutaraldehyde (GLA) and bovine serum albumin (BSA). An initial investigation of the morphology of the synthesized AuNWA by field emission scanning electron microscopy (FESEM) and field emission transmission electron microscopy (FETEM) revealed that the nanowires array was highly ordered with rough surface, and the electrochemical features of the AuNWA with/without modification were also investigated. The integrated AuNWA–BSA–GLA–GOx nanobiosensor with Nafion membrane gave a very high sensitivity of 298.2μAcm−2mM−1 for amperometric detection of glucose, while also achieving a low detection limit of 0.1μM, and a wide linear range of 5–6000μM. Furthermore, the nanobiosensor exhibited excellent anti-interference ability towards uric acid (UA) and ascorbic acid (AA) with the aid of Nafion membrane, and the results obtained for the analysis of human blood serum indicated that the device is capable of glucose detection in real samples.

Voltammetric microsensor using PEDOT-modified gold electrode for the simultaneous assay of ascorbic and uric acids

A voltammetric microsensor has been developed for the simultaneous assay of ascorbic (AA) and uric (UA) acids in aqueous solution. The electrode surface has been modified by means of electropolymerized conductive poly(3,4-ethylenedioxythiophene) PEDOT organic films. The electrocatalytic activity of the interface was dependent on the electropolymerization parameters inducing change in the structure and the morphology of the resulting polymer. The PEDOT thickness was optimized in order to maximize the peak potential separation between both acids oxidation to more than 400 mV. By using differential pulse voltammetry (DPV), the sensitivity of the microsensor was 0.87 μA μM −1 cm −2 and 4.05 μA μM −1 cm −2 for AA and UA respectively. The later was sensible to the presence of AA in the mixture, making evidence of the catalytic mechanism of UA regeneration. The calibration curves were linear in the concentration range 5.0–300 μmol L −1 for AA and 2.0–600 μmol L −1 for UA. The detection limits were 2.5 μmol L −1 and 1.5 μmol L −1 respectively. The sensor response was unmodified in the presence of the major electroactive biomarkers. The application of the PEDOT modified microsensor to the analysis of human blood serum was evaluated.

In-depth Exploration of Cerebrospinal Fluid by Combining Peptide Ligand Library Treatment and Label-free Protein Quantification

Cerebrospinal fluid (CSF) is the biological fluid in closest contact with the brain and thus contains proteins of neural cell origin. Hence, CSF is a biochemical window into the brain and is particularly attractive for the search for biomarkers of neurological diseases. However, as in the case of other biological fluids, one of the main analytical challenges in proteomic characterization of the CSF is the very wide concentration range of proteins, largely exceeding the dynamic range of current analytical approaches. Here, we used the combinatorial peptide ligand library technology (ProteoMiner) to reduce the dynamic range of protein concentration in CSF and unmask previously undetected proteins by nano-LC-MS/MS analysis on an LTQ-Orbitrap mass spectrometer. This method was first applied on a large pool of CSF from different sources with the aim to better characterize the protein content of this fluid, especially for the low abundance components. We were able to identify 1212 proteins in CSF, and among these, 745 were only detected after peptide library treatment. However, additional difficulties for clinical studies of CSF are the low protein concentration of this fluid and the low volumes typically obtained after lumbar puncture, precluding the conventional use of ProteoMiner with large volume columns for treatment of patient samples. The method has thus been optimized to be compatible with low volume samples. We could show that the treatment is still efficient with this miniaturized protocol and that the dynamic range of protein concentration is actually reduced even with small amounts of beads, leading to an increase of more than 100% of the number of identified proteins in one LC-MS/MS run. Moreover, using a dedicated bioinformatics analytical work flow, we found that the method is reproducible and applicable for label-free quantification of series of samples processed in parallel.

Extensive Analysis of the Cytoplasmic Proteome of Human Erythrocytes Using the Peptide Ligand Library Technology and Advanced Mass Spectrometry

The erythrocyte cytoplasmic proteome is composed of 98% hemoglobin; the remaining 2% is largely unexplored. Here we used a combinatorial library of hexameric peptides as a capturing agent to lower the signal of hemoglobin and amplify the signal of low to very low abundance proteins in the cytoplasm of human red blood cells (RBCs). Two types of hexapeptide library beads have been adopted: amino-terminal hexapeptide beads and beads in which the peptides have been further derivatized by carboxylation. The amplification of the signal of low abundance and suppression of the signal of high abundance species were fully demonstrated by two-dimensional gel maps and nano-LC-MSMS analysis. The effect of this new methodology on quantitative information also was explored. Moreover using this approach on an LTQ-Orbitrap mass spectrometer, we could identify with high confidence as many as 1578 proteins in the cytoplasmic fraction of a highly purified preparation of RBCs, allowing a deep exploration of the classical RBC pathways as well as the identification of unexpected minor proteins. In addition, we were able to detect the presence of eight different hemoglobin chains including embryonic and newly discovered globin chains. Thus, this extensive study provides a huge data set of proteins that are present in the RBC cytoplasm that may help to better understand the biology of this simplified cell and may open the way to further studies on blood pathologies using targeted approaches.

High‐throughput solid‐phase permethylation of glycans prior to mass spectrometry

Permethylation of glycans prior to their mass spectrometric determination has now become a time-honored methodology in glycoconjugate analysis due to the advantage of a simultaneous analysis of neutral and acidic glycans as well as enhanced sensitivity and easier tandem mass spectrometry interpretation. While the different solvent extraction-based versions of this method often suffice in different structural studies, they are generally less satisfactory in the quantitative determinations aiming at minor quantities of the analyzed materials. To overcome these difficulties, we recently introduced a solid-phase capillary permethylation technique (Kang et al., Rapid Commun. Mass Spectrom. 2005; 19: 3421) for microscale determination. Here, we describe a very useful high-throughput extension of the solid-phase methodology utilizing spin columns packed with sodium hydroxide beads. This procedure has been thoroughly optimized to match the analytical performance parameters of the previously used capillary technique. As demonstrated with a high-precision glycomic profiling analysis of human blood serum, this methodological improvement offers simplicity and reproducibility, allowing the complete permethylation of 12-18 samples in less than 20 min.

Magnetooptical characterization of human blood serum: correlation between neoplasmic changes and their biomolecular information carriers

This article reports a method of gathering information on a neoplasmic disease on the basis of direct magnetooptical analysis of human blood serum. In a strong magnetic field B and optical field k, biomolecules carrying the information on the neoplasmic changes in human blood serum generate transitions following from their multipolar optical polarizability. The results of the study introduce magnetooptical characteristics of the neoplasmic changes in information carrier. The proposed magnetooptical technique for detection of neoplasmic changes requires a sample of human blood serum of about 2 mL and the availability of a strong magnetic field (10–30 T).

Analysis of human blood serum using the off-line coupling of capillary isoelectric focusing to matrix-assisted laser desorption/ionization time of flight mass spectrometry

Off-line coupling of capillary IEF (CIEF) with matrix-assisted laser desorption/ionization mass spectrometry was utilized for the analysis of human blood serum. Serum proteins were initially separated by CIEF, and fractions of the isoelectric separation were eluted sequentially to a MALDI-TOF MS sample target. During pressure elution of the CIEF sample, voltage was maintained across the capillary system utilizing a sheath flow arrangement to minimize band broadening induced by the laminar flow field. Both pI and mass information were obtained from the complex biological sample, similar to traditional 2-DE techniques, and the platform was faster (hours versus days), more automatable, and simpler than 2-DE. The volume of raw sample present in the actual analysis was approximately 100 nL, making this technique well suited for very rare specimens. Additionally, the speed and simplicity of the technology make it an attractive technique for performing initial comparative analyses of complex samples.

Fiber-optic evanescent-wave spectroscopy and neural networks: application to chemical blood analysis

An infrared spectral analysis of human blood serum has been carried out with Fourier transform infrared fiber-optic evanescent-wave spectroscopy. The measured spectra were analyzed by neural network analysis to predict concentrations of cholesterol, creatinine, total protein, urea, and uric acid. The predicted concentrations were compared with results from standard chemical analysis of blood and a good correlation was observed.

Analysis of human blood serum and human brain samples by total reflection X-ray fluorescence spectrometry applying Compton peak standardization

The method of using the Compton peak as internal standard in total reflection X-ray fluorescence (TXRF) determination is established for trace element determination of Fe, Cu, Zn, Se and Pt in human serum and of Cu and Zn in homogenized brain samples. A new method of spectrometer sensitivity calibration using spiked matrices with known amounts of trace elements is tested against established methods of matrix matching as well as internal element addition. The analytical results with the proposed procedure are compared to a certified international standard and to values with Atomic Absorption Spectrometry (AAS) obtaining analytical results of comparable accuracy and precision. The method is adequate for routine clinical analysis as it has the advantages of requiring very small amounts of material and simple preparations, which avoids the chemical digestion stage.

Blood diagnostics using fiberoptic evanescent wave spectroscopy and neural networks analysis

A spectral analysis of human blood serum was undertaken by fiberoptic evanescent wave spectroscopy (FEWS) using a Fourier transform infrared (FTIR) spectrometer. The blood serum samples were introduced into a special cell designed for FEWS of liquids, with an IR transmitting silver halide fiber as the sensing element. The spectra were analyzed by models of neural networks (NN), and the concentrations of protein, cholesterol and uric acid in human blood serum were obtained. Our results are in agreement with those obtained by ordinary chemical enzymatic methods and multivariate calibration methods. The estimated prediction errors obtained (in percent of the average value) were 4.7% for total protein, 22% for cholesterol and 35% for uric acid. This method can be used for in-situ real-time blood analysis.

Fiber-optic evanescent-wave spectroscopy for fast multicomponent analysis of human blood

A spectral analysis of human blood serum was undertaken by fiber-optic evanescent-wave spectroscopy (FEWS) by the use of a Fourier-transform infrared spectrometer. A special cell for the FEWS measurements was designed and built that incorporates an IR-transmitting silver halide fiber and a means for introducing the blood-serum sample. Further improvements in analysis were obtained by the adoption of multivariate calibration techniques that are already used in clinical chemistry. The partial least-squares algorithm was used to calculate the concentrations of cholesterol, total protein, urea, and uric acid in human blood serum. The estimated prediction errors obtained (in percent from the average value) were 6% for total protein, 15% for cholesterol, 30% for urea, and 30% for uric acid. These results were compared with another independent prediction method that used a neural-network model. This model yielded estimated prediction errors of 8.8% for total protein, 25% for cholesterol, and 21% for uric acid.

Multielement analysis of human blood serum by neutron activation and controlled potential electrolysis.

The combination of neutron activation analysis and controlled potential electrolysis has proved itself to be a useful multielement method for the determination of 14 elements in a 500-..mu..L sample of human blood serum. After freeze-drying and irradiation, the samples are decomposed and electrolyzed for 3 h. The radioactive species deposited on the mercury cathode allow simultaneous determination of eight elements (Ag, Au, Cd, Co, Fe, Hg, Sb, Zn). Another six elements (Br, Ca, Cs, Na, Rb, Sc) are determined by measuring the activities in the residual solution. Data illustrating the precision of the method are given. The determination limits are of the order 10/sup -1/-10/sup -3/ ..mu..g/L for the nine trace elements investigated.

Trace element analysis of biological materials by thermal and epithermal neutron activation analysis

A scheme for instrumental neutron activation analysis of biological materials is proposed. The scheme is based on a combination of thermal and epithermal activation. The accuracy is evaluated by analyzing 4 standard reference materials. Results from the analysis of human blood serum and plasma are given.