Sample Volume Research Articles

Development of two ultra-sensitive UHPLC-QqQ-MS/MS methods for the simultaneous determination of hydroxyzine and its active metabolite (cetirizine) in human blood: applications to real cases of forensic toxicology.

Both postmortem toxicological and medical-forensic examinations are very important in the case of analyzing various types of chemical substances. Hydroxyzine (HZ) is a first-generation antihistamine drug with a sedative effect that disrupts cognitive function and affects the ability to drive motor vehicles. Enzymatic oxidation of the hydroxy-methyl group to the carboxyl group leads to the formation of its main metabolite-cetirizine (CZ). CZ is the active substance of antiallergic drugs. Because it does not cross the BBB (blood-brain barrier) easily, it is less likely to cause drowsiness or affect memory and impair cognitive function. Therefore, in criminal studies, it is often important what medication had been taken by a person involved, e.g., in a car accident, HZ or CZ. The analysis of both antihistamine drugs is challenging, as usually very low concentrations of the compound of interest need to be determined. Thus, an ultra-sensitive UHPLC-QqQ-MS/MS method was developed for simultaneous determination of HZ and CZ in biological fluid samples. The lower limit of quantification (LOQ) for HZ and CZ was calculated as 0.345 and 0.3696ng/mL, respectively. Together with a reduced sample volume to 200 μL, it makes the developed method suitable for a sensitive multidrug forensic toxicological analysis. Samples were extracted with simple and fast liquid-liquid extraction (ethyl acetate, pH 9). The present method for the determination of HZ and CZ in human blood proved to be simple, fast, selective, and sensitive. The quantification by LC-MS/MS was successfully applied to the samples coming from 28 authentic biological fluids (blood, urine, vitreous humor, bile and stomach content), both antemortem and postmortem. The performed studies confirm that the developed method is characterized by a high extraction efficiency. Its accuracy, reproducibility, simplicity, and selectivity suggest its application in clinical, toxicological, and forensic laboratories.

Development of an Automatic Liquid Dosing System in Microliter Scale

The goal of this study is to design a novel automatic liquid dosing system for liquid sampling at the microliter level. For this purpose, a mechatronics system is designed to position a syringe at the desired position in the workspace and then drive its piston to inject the liquid to be sampled. Then, an application-specific algorithm is developed to be able to prepare samples in 3x3, 4x4, and 5x5 sample container arrays with a desired volume. The performance tests are conducted for preparing samples with up to three different liquids. The repetitive experiments are performed for 50 and 100 µL sampling volumes. The results indicated that it is possible to dose a single liquid with the highest average deviation of 3.9%. Moreover, it is found that it is possible to prepare a sample with a mixture of three liquids by the highest average deviation from the reference value of around 3.4% when the targeting sampling volume is 250 µL for each liquid.

Influence of Supercritical Conditions on Isothermal Adsorption Capacity Calculation of Methane and Model Optimization.

The study of isothermal adsorption models for coal and methane under supercritical conditions helps us to understand the gas content distribution in coal seams and improve gas management in coal mines. Coal samples from the Hebi mine and Longshan coal mine were selected for this research. Using the weight method, isothermal adsorption curves of supercritical methane at temperatures of 308, 313, and 318 K were measured. The adsorption phase density of supercritical methane was obtained by using the intercept method and model fitting, and the absolute adsorption capacity of methane was calculated. Simultaneously, the pore volume and specific surface area of the coal samples were tested, and the isothermal adsorption model for supercritical methane was studied and optimized. The results indicate that using mercury intrusion, low-temperature N2, and low-temperature CO2 adsorption methods for testing coal sample pore structures allows for multiscale characterization of the coal pore structure. Based on the Gibbs excess adsorption theory, the phase density of methane obtained from the intercept method and model fitting effectively represent the isothermal adsorption curve of supercritical methane. By combination of the specific surface area and pore volume distribution of the coal with the absolute adsorption capacity of methane, the number of methane adsorption layers on the coal surface was calculated to be between 1.11 and 1.3 layers. This suggests that the isothermal adsorption model for supercritical methane on coal is not solely micropore filling or single molecular layer adsorption but primarily single molecular layer adsorption with concurrent micropore filling adsorption. Based on this adsorption mechanism, an L-DA isothermal adsorption model for supercritical methane on coal was established. The L-DA isothermal model shows good fitting results and effectively explains the adsorption characteristics of supercritical methane on coal.

Natural deep eutectic solvent-based dispersive liquid-liquid microextraction of pesticides in drinking waters combined with GC-MS/MS detection

The current research aims to develop a new analytical method applying a dispersive liquid-liquid microextraction (DLLME) assisted by vortex and using an environmentally friendly extractant for the preconcentration of organochlorine and organophosphorus pesticides followed by gas chromatography-tandem mass spectrometry (GC-MS/MS) analysis. The extractant (i.e., natural deep eutectic solvent (NADES)) is safe, cheap, biodegradable and can be prepared by simply mixing DL-menthol and decanoic acid (molar ratio 2:1). The main experimental factors affecting the extraction of all analytes evaluated (19 organochlorine and organophosphorus pesticides) have been optimised using a multivariate analysis consisting in two steps: a Plackett-Burman design followed by a central composite design (CCD). Seven experimental factors have been evaluated: (i) sample volume; (ii) NADES volume; (iii) sample pH; (iv) extraction time; (v) centrifugation time; (vi) centrifugation speed; and (vii) ionic strength (NaCl %, w v−1). For the significant variables, the optimum values were 10 mL sample and 45 μL NADES. No pH adjustment as well as addition of NaCl were needed. The other variables were set at 3 min extraction time, 5 min centrifugation time and 900×g centrifugation speed, respectively. Under the optimised extraction conditions, the limit of quantification (LOQ) values ranged between 0.2 and 78 ng L−1 for all analysed pesticides. Furthermore, the proposed analytical method has been successfully applied to drinking water (bottled spring water). The recovery study (n = 3) has been evaluated at 0.1, 1.0 and 5.0 μg L−1 spiking levels, obtaining relative recovery values within the range of 70 % and 117 % and RSD values between 1 % and 20 % for all the analytes studied, except for p,p-DDT (56–77 % in high conductivity water samples).

Efficacy and speed of effect after the first dose of aspirin in children with congenital heart disease.

Many paediatric studies report that patients must be established on aspirin therapy for a minimum of 5 days to achieve adequate response. This is not always practical especially in critical settings. Prospective identification of patients that are unresponsive to aspirin sooner could potentially prevent thrombotic events. The aim of this study was to investigate prospectively if the first dose of aspirin is effective in decreasing platelet aggregation, and thromboxane formation and if this can be measured after 2 hours in paediatric cardiology patients. A secondary aim was to identify a cut-off for a novel marker of aspirin responsiveness the maximum amplitude with arachidonic acid, which could potentially dramatically reduce the blood volume required. Third, we aimed to prospectively identify potentially non-responsive patients by spiking a sample of their blood ex vivo with aspirin. The majority (92.3%) of patients were responsive, when measured 2 hours post first dose of aspirin. Non-response or inadequate response (7.7%) can also be identified at 2 hours after taking the first dose of aspirin. Additionally, we have shown a novel way to reduce blood sample volume requirements by measurement of the maximum amplitude with arachidonic acid as a marker of response, particularly for monitoring. These findings of rapid efficacy in the majority of patients offer assurance in a sound, practical way to attending clinicians, patients, and families.

General population screening for type 1 diabetes using islet autoantibodies at the preschool vaccination visit: a proof-of-concept study (the T1Early study)

ObjectiveType 1 diabetes (T1D) screening programmes testing islet autoantibodies (IAbs) in childhood can reduce life-threatening diabetic ketoacidosis. General population screening is required to detect the majority of children with T1D,...

From Proteomics to the Analysis of Single Protein Molecules.

Limit of detection (LoD) is a term that is used to characterize the sensitivity of an analytical method. The existing limitation of the sensitivity of analysis using modern mass spectrometry methods has been experimentally shown to be a limiting factor in the application of proteomic technologies in medicine. This article proposes a concept of a new technology that will set a new vector of development in the development of systems for solving problems of medical diagnostics and deals with theoretical and practical aspects of creating a new technology for the detection of single biomacromolecules (in particular, proteins) in biological samples. Such technology should be based on the principle of signal registration similar to that used in a Geiger counter (also known as a Geiger-Müller counter or G-M counter), a device that automatically counts the number of ionizing particles that hit it. This counter is free from probabilistic components; it registers a signal if there is at least one target molecule in the analysis chamber. Predictive medical diagnostics require technology based on methods where sensitivity allows for the detection of single marker molecules in a biological sample volume of 1-10 µL, the smallest volume of biomaterial used in laboratory diagnostics. Creation of a detector with a sensitivity of 10-18 M would allow for the detection of one molecule in 1 µL of the sample, which fundamentally makes this approach analogous to a G-M counter for solutions. To date, bioanalytical methods are limited to a sensitivity of 10-12 M (which is approximately 1 million molecules per 1 μL), which is insufficient to capture the early stages of pathological processes.

Development and validation of a bioanalytical method using LC-MS/MS to quantify Zanamivir in human plasma.

This study presents the development and validation of a highly sensitive and accurate LC-MS/MS method for the quantification of Zanamivir in human plasma. Utilizing a mobile phase of MeOH and Ammonium Formate (30:70 v/v) and a protein precipitation extraction technique, the method achieves excellent specificity and reproducibility. The calibration curve demonstrated linearity from 4-80 ng/ml with a correlation coefficient (r²) ≥ 0.9994. The method adheres to FDA guidelines, exhibiting high precision and accuracy with intra-day and inter-day variations of 0.81-1.98%. The developed method, requiring a minimal sample volume and a short run time, ensures robustness and stability across various analytical conditions, making it an ideal tool for Zanamivir plasma concentration studies.

Greener analysis of eleven basic drugs in blood and urine using carbowax 20M based biofluid sampler (BFS) device

For the first time, a novel biofluid sampler (BFS) and sample preparation deviceis applied for the analysis of 11 basic drugs (i.e., pheniramine, chlorpheniramine, fluoxetine, tramadol, amitriptyline, ketamine, diazepam, chlordiazepoxide, clozapine, chlorpromazine, dothiepin) in biological matrices (i.e., blood and urine). BFS utilizes advanced, highly effective sorbents derived from sol-gel sorbent coating technology onto cellulose fabric substrate, improving sample collection and retention. BFS has the capability to retain a biological sample from 10 to 1000 µL without requiring any dilution or pre-treatmentof the sample. The biological samples were pipetted onto the BFS device and dried at room temperature. Subsequently, adsorbed analytes were back-extracted into 1000 µL of methanol without requiring any imposed external diffusion process andthen analyzed by gas chromatography-mass spectrometry (GC-MS). A one-factor-at-a-time (OFAT) screening procedure was used to extensively screen and optimize several parameters, including sample volume, elution time, solvent volume, and solvent type. Under the optimal conditions of the study, the method was found to be linear within the range 0.1–10 µg mL−1 for both blood and urine. Quantification limits were established for blood samples within the range of 0.084 to 0.099 μg mL−1 and for urine samples within the range of 0.060 to 0.078 μg mL−1. The precisions within and between days were less than 7 % and 10 %, respectively. The target analytes showed good recoveries utilizing the recommended protocol, with ranges of 45.1 % − 103.4 %. Furthermore, the methodology has been effectively implemented in forensic toxicology case work. Moreover, the green characteristics and applicability of the suggested methodology was evaluated using softwares i.e., AGREE and BAGI.

Polypyrrole as Adsorbent in Magnetic Solid Phase Extraction for Progesterone Determination from Human Plasma.

A straightforward and effective chromatographic method has been created for the analysis of progesterone from human plasma using a composite based on polypyrrole/magnetic nanoparticles in the sample preparation procedure. The quantification of progesterone is necessary due to its importance in human development and fertility. The employed conditions used acetonitrile:ultrapure water (70:30, v/v) as the mobile phase at 1.0 mL min-1 and an octadecyl silane column (Phenomenex Gemini, 250 mm × 4.6 mm, 5 μm) at a wavelength of 235 nm. The composite and its precursors were synthesized and properly characterized by X-ray diffraction, Fourier transform infrared spectroscopy, scanning electron microscopy/energy dispersive spectroscopy, thermogravimetric analysis, and point of zero charge. The main factors affecting the extraction recovery of progesterone from pool human plasma samples employing magnetic solid phase extraction have been studied, such as sample pH (without adjustment), sample volume (1000 μL), washing solvent (ultrapure water), eluent (acetonitrile), eluent volume (1000 μL), and amount of adsorbent (10 mg). The extraction recoveries ranged from 98% to 102%, and linearity ranged from 5 to 3000 ng mL-1. The correlation coefficient (r) was ≥0.99, and acceptable relative standard deviation (precision), relative error (accuracy), and p-values (robustness) were observed. Lastly, the plasma samples from pregnant women were successfully analyzed by the validated method.

Modelling and impact of tensiometer plate geometry and sample volume on biosurfactant surface activity assessment

Biosurfactants are molecules with hydrophilic and hydrophobic moieties with the capacity to reduce the surface tension of water. Given the limited quantity of biosurfactant extracts in laboratories, it is recommended to use equipment that requires minimal sample quantities for detecting the presence of biosurfactants. In this work, commercial glycolipids biosurfactants (rhamnolipids or sophorolipids) were diluted in water and subjected to different analyses to obtain their minimum surface tension (ST) reduction and their critical micellar concentration (CMC). The independent variables of the study were: the geometry of platinum plate (rectangular or cylindrical), the sample volume (2, 4 and 20 mL) and the container material consisting of either glass or polytetrafluoroethylene (PTFE). The variation of ST with biosurfactant concentration was studied based on the isotherm model proposed by Li & Lu. It was observed that the profile of ST values did not vary so much using the different independent variables described, observing that platinum rectangular plate can be used for volumes of 4 mL biosurfactants instead of cylindrical plate usually recommended for volumes lower than 20 mL, the container material was also not significant based on the Pearson and Spearman statistical treatment. Moreover, well-fitting regression model results were obtained for a non-commercial biosurfactant extract obtained from a residual stream of the dairy industry, predicting values close to the observed data.

Synthesis of magnetic multi-walled carbon nanotubes with layered double hydroxide (M-MWCNTs@MnAl-LDH) nanocomposite as an adsorbent for lead extraction

MnAl layered double hydroxide hybrid with magnetic-multiwalled carbon nanotubes was synthesized by a hydrothermal method and used for the extraction of Pb(II) (lead) from spices and water samples in the dispersive solid phase microextraction (dSPμE) technique using FAAS. The as-prepared adsorbent MMWCNTs@MnAl-LDH was characterized by XRD, FTIR, EDX, and SEM techniques. Various analytical parameters were optimized, including pH 8, adsorbent dosage of 5 mg, HNO3 eluent concentration of 1 mol L−1, eluent volume of 3 mL, eluent time of 60 s, and sample volume of 20 mL, for quantitative lead recoveries, with an LOD of 0.314 μg L−1, an LOQ of 1.048 μg L−1, and PF of 11.53. Under the optimized conditions, the linearity ranges from 0.5 to 500 μg L−1 (R2 = 0.9997). For the validation test of the established dSPμE procedure, Certified reference materials (CRMs) were used, yielding satisfactory recovery results ranging from 97.8 to 102.7 %. The method was applied to determine lead in turmeric, tap water, and industrial water samples.

Order-disorder phase transitions in Fe81Ga19-RE ALLOYS (RE = Dy, Er, Tb, Yb) according to neutron diffraction data

New data on the phase compositions and structural transformations in a number of Fe81Ga19 alloys doped with trace amount (≤ 0.2 at.%) of rare earth elements are presented. The data are obtained in neutron diffraction experiments performed with high resolution and in continuous temperature scanning mode when heated to 900 °C and subsequent cooling. It has been established that structural rearrangements proceed in a generally identical manner both in the original Fe81Ga19 alloy and in its doped analogues. Slow heating and subsequent cooling of the alloys (rate ±2 °C/min) leads to the formation of clusters of the D03 phase with sizes in the range (200–300) Å in the matrix of the disordered A2 phase. The sizes and volume fraction of clusters (~0.3 of the sample volume) weakly depend on the specific composition. The degree of ordering of the clusters atomic structure changes with temperature according to a second-order phase transition and is close to unity at room temperature. The search for structural ordering corresponding to the modified D03 phase, discovered in a number of electron diffraction studies, did not lead to a positive result.

Lab-on-a-chip paper-based electrochemically assisted solid-phase microextraction and ion selective sensor for determination of naproxen in biological fluid samples

BackgroundThe integration of paper-based analytical devices with lab-on-a-chip technology has opened up new possibilities for miniaturized, rapid, low cost and portable diagnostic testing in resource limited settings. This work introduces an integrated lab-on-a-chip platform coupling paper-based analytical devices (LOC-PADs) for dual usage: electrochemically controlled solid-phase microextraction (EC-SPME) and determination of naproxen (NAP) by potentiometric ion-selective electrode (ISE) in biological samples. ResultsConductive papers were successfully fabricated by a chemical procedure. Thereupon, a conducting molecularly imprinted polymer (CMIP) film based on PPy and an anionic model drug (NAP) as a template were applied to the conductive paper substrate via electrochemical methods. A microfluidic chip device was employed to assess the analytical performance of the fabricated paper-based CMIP. The designed chip is divided into two parts with a microfluidic channel between them. The first chamber is an extraction zone for EC-SPME of NAP by using the CMIP paper as a selective sorbent. The second chamber is the detection zone and consists of a paper-based NAP ion selective sensor for potentiometric detection of drug. The analytical process was investigated and optimized for each chamber. Remarkable selectivity towards NAP was achieved in the concentration range from 4.0 × 10−7 to 1.0 × 10−2 mol L−1 with determination coefficient (R2 = 0.99) and the limit of detection (LOD) was 2.0 × 10−7 mol L−1. Anionic Nernstian compliance was gained −58.5 ± 0.5 mV decade−1, and response time in the detection step was 7s for ISE. Satisfactory spiking recovery values within the acceptable range of 90–110 % with RSDs ≤7 % were achieved for the analysis of real samples with fully portable LOC device. SignificanceThe LOC-PADs were favorably used for selective extraction and determination of NAP in serum and saliva samples, respectively. EC-SPME caused sample clean-up, reduced or eliminated different interferences and enhanced selective response of ISE as detection zone of device. Integration of EC-SPME and ISE in a single chip enabled easy and fast determination of trace amounts of NAP in limited real sample volumes, and fully portable advantages.

Statistical analysis of Cu content effects on structural properties in CuZr metallic glasses

This study examines the effects of casting conditions on the structural properties of CuZr metallic glasses (MGs) using molecular dynamics simulations. The influence of Cu content on various structural properties was explored, finding significant power-law relationships that indicate increased Cu promotes the formation of icosahedra-like structures and enhances the population of solid-like polyhedra. In contrast, the clustering coefficient, reflecting solid-like connectivity, showed a linear relationship with Cu content, revealing that while Cu increases solid-like structures, their connectivity does not scale proportionally. No significant correlations were found for sample volume, cooling rate, or temperature within the studied ranges. This study highlights the utility of statistical analysis in elucidating material property relationships, contrasting with the less interpretable nature of machine learning models. The findings provide valuable insights into the role of Cu content in MGs and demonstrate the importance of traditional statistical approaches for material characterization.

Luminescent Terbium Probe for Time‐Resolved FRET and NSET Binding Assays with Quantum Dots and Gold Nanoparticles

Time‐gated or time‐resolved FRET (TR‐FRET) assays are important tools in biosensing, bioimaging, drug screening, and molecular diagnostics. Efficient TR‐FRET assays require stable lanthanide complexes with high absorption cross sections, high quantum yields, and long photoluminescence lifetimes. Owing to their challenging synthesis, such complexes are relatively rare and new components are of potential interest when developing TR‐FRET probes. Here, we evaluate the recently developed Tb complex CoraFluor‐1 concerning its analytical performance in terbium‐to‐quantum dot FRET and terbium‐to‐gold nanoparticle NSET assays using the prototypical biological recognition system of biotin and streptavidin. Biological binding was quantifiable at sub‐picomolar concentrations in small sample volumes, with broad applicability demonstrated across three commercial fluorescence plate readers used for time‐resolved, spectrally‐resolved, and clinical bioanalysis. Overall, CoraFluor‐1 provided excellent analytical performance as both FRET and NSET donor, validating its potential for developing new TR‐FRET probes for biosensing and bioimaging.

Fan assisted extraction and low pressure chromatographic system with a phenyl monolithic column for amperometric determination of volatile α-dicarbonyl compounds in coffee brews

In this work, a low pressure chromatographic system featuring a single 0.5 cm length phenyl monolithic column and an amperometric detector was developed to determine volatile ⍺-dicarbonyl compounds in coffee brews. The analytical strategy relied on the extraction of the volatile ⍺-dicarbonyl compounds and their derivatization with o-phenylenediamine, prior to the determination of the resulting quinoxaline derivative compounds in the designed chromatographic system. The studies of the mobile phase composition and electrochemical conditions are presented. The extraction of the volatile ⍺-dicarbonyl compounds was performed resourcing to a recent approach based on the fan assisted extraction system. This system performs the full evaporation technique and allowed the quantitative recovery (>85 %) of diacetyl and 2,3-pentanedione from the coffee brew based on the following experimental conditions: sample volume 30 µL, extraction period 15 min, acceptor solution (water, acetonitrile, 90:10, v/v) volume 400 µL, and sample temperature 50 °C. The developed method showed no matrix effect for different coffee brews and the quantification of the studied analytes can be performed through the external calibration method. Detection limits of 3.6 × 10−6 mol/L for diacetyl and 12.9 × 10−6 mol/L for 2,3-pentanedione were obtained. Sample analysis rate was of 5 h−1 and the acetonitrile consumption was of ca. 450 µL per chromatographic run.

Selective Gas-Phase Depletion of Chemical Contaminants in Dissolved Organic Matter Increases Compositional Coverage by FT-ICR Mass Spectrometry.

Fourier transform ion cyclotron resonance mass spectrometry of dissolved organic matter (DOM) extracted from environmental samples provides molecular speciation that enables visualization of compositional trends in the fate and cycling of biogenic and anthropogenic organics. Often, chemical contamination is introduced during field sampling (i.e., remote locations, cannot use glass). Further, preconcentration of DOM by solid-phase extraction often results in chemical contamination. When chemical noise is a dominant fraction of the ion signal, mass spectral performance is degraded by reduction of the ion trap analyte accumulation capacity and enhanced ion cloud dephasing during ICR detection. We have developed gas-phase ion depletion of unwanted chemical contaminants during ion injection into the linear RF ion trap of the hybrid linear ion trap 21 T FT-ICR mass spectrometer that improves detection of analytes by removing unwanted chemical noise. We demonstrate improvements in signal-to-noise ratio, dynamic range, and the number of observed analytes in dissolved organic matter samples that results in a 40-100% increase in the number of identified analytes. In many cases, the number of peaks observed per nominal mass more than doubles over select m/z regions. This gas-phase "clean-up" can salvage precious samples challenged by sampling location, sample volume, or collection protocols that cannot be avoided and maximizes the compositional information obtained. Further, this approach is generalizable and extendable to any hybrid linear ion trap instrument platform (e.g., LTQ-Orbitrap or linear ion trap-TOF). We highlight the power of gas-phase depletion with electrospray ionization, but this method is also applicable to other ionization modes.

Development and Application of a Liquid Sampling Device with Timing, Quantitative and Depth Determination

This thesis describes the development process of a liquid sampling device that can be timed, quantitative, and deep. The structural composition, working principle, and innovative features of the liquid sampling device are introduced, and the application prospects are analyzed. The device has the function of controllable and adjustable sampling point depth and sampling volume, and adopts quantitative collection and collection methods without contact with accessories. The device has a silent function in the sample body, which can realize pre-booking of collection time points and multi-point synchronous collection. Wireless remote control automatic mode is adopted for sending and stopping collection instructions, which can be sent one-to-one or one-to-many at the same time. It provides automation links for water quality monitoring projects and meets the requirements of modern scientific and technological development.

A Novel Size-Based Centrifugal Microfluidic Design to Enrich and Magnetically Isolate Circulating Tumor Cells from Blood Cells through Biocompatible Magnetite-Arginine Nanoparticles.

This paper presents a novel centrifugal microfluidic approach (so-called lab-on-a-CD) for magnetic circulating tumor cell (CTC) separation from the other healthy cells according to their physical and acquired chemical properties. This study enhances the efficiency of CTC isolation, crucial for cancer diagnosis, prognosis, and therapy. CTCs are cells that break away from primary tumors and travel through the bloodstream; however, isolating CTCs from blood cells is difficult due to their low numbers and diverse characteristics. The proposed microfluidic device consists of two sections: a passive section that uses inertial force and bifurcation law to sort CTCs into different streamlines based on size and shape and an active section that uses magnetic forces along with Dean drag, inertial, and centrifugal forces to capture magnetized CTCs at the downstream of the microchannel. The authors designed, simulated, fabricated, and tested the device with cultured cancer cells and human cells. We also proposed a cost-effective method to mitigate the surface roughness and smooth surfaces created by micromachines and a unique pulsatile technique for flow control to improve separation efficiency. The possibility of a device with fewer layers to improve the leaks and alignment concerns was also demonstrated. The fabricated device could quickly handle a large volume of samples and achieve a high separation efficiency (93%) of CTCs at an optimal angular velocity. The paper shows the feasibility and potential of the proposed centrifugal microfluidic approach to satisfy the pumping, cell sorting, and separating functions for CTC separation.