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Objectives Our objectives were to determine the separation characteristics and blood product quality of a gravity-driven microfiltration blood separation system (HemoClear, The Netherlands). Background A range of centrifugal blood separation devices, including intraoperative cell salvage devices (cell savers) and apheresis machines, are available to assist in preparing both allogenic and autologous blood products. These devices are expensive to operate and require extensive training. Methods and materials Nine whole blood units were collected under standard conditions and analysed for haematological parameters, thromboelastographic properties, platelet morphology and activation, and red blood cell (RBC) deformability and morphology. Three whole blood units were separated by means of the HemoClear device, into a liquid and cellular component. The cellular component was diluted with SAGM and cold stored for 14 days. To simulate cell salvage six whole blood units were diluted with isotonic saline, followed by multiple HemoClear separation rounds. Results The recovery of both RBCs (100 ± 1.6%) and white blood cells (99 ± 4.5%) after undiluted filtration were very high, while platelet recovery was high (83 ± 3.0%). During the filtration, and cold storage after filtration storage both the non-deformable RBC fraction and the RBC maximum elongation remained stable. Parameters of thromboelastography indicated that platelets remain functional after filtration and after 7 days of cold storage. In the cell salvage simulation the total protein load in the cellular fraction was reduced by 65 ± 4.1% after one washing round and 84 ± 1.9% after two consecutive washing rounds. Conclusion The novel blood filter studied effectively separates whole blood into diluted plasma and platelet-rich RBCs. Moreover, the device effectively washed diluted whole blood, driving over 80% of proteins to the liquid component.

Severe internal trauma results in millions of hospitalizations each year, including thousands of deaths caused by subsequent multiple organ failure. The majority of these deaths occur within the first 24 h, and thus, rapid diagnosis of internal trauma severity is necessary for immediate treatment. For early organ damage identification, diagnosis in point-of-care settings is crucial for rapid triage and treatment. Recent reports suggest that circulating histones may serve as a biomarker for severe organ damage and the risk of multiple organ failure. Here, we report a point-of-care diagnostic system that utilizes the inherent interactions between histones and DNA for the fluorescence-based detection of histones in whole blood. In the assay, histones within the sample are wrapped by DNA, thus preventing an intercalating dye from binding the DNA and fluorescing. To allow for quantitative fluorescent measurements to be made in a point-of-care setting, we integrate a rapid, automated blood separation step into our assay. Furthermore, we eliminate manual reagent additions using a thermally responsive alkane partition (TRAP), thus making the system sample-to-answer. Finally, we demonstrate the assay in a portable fluorescence reader compatible with a point-of-care environment. We report a limit of detection 112 ng/mL in whole blood, suggesting that our device can be used to rapidly diagnose internal trauma severity and the likelihood of multiple organ failure in near-patient settings.

The benefit of autologous platelet‐rich plasma (PRP) treatment is still under discussion. Variations in PRP products, consequence of the lack of a standardised protocol for the multitude of commercially available blood separation systems and the lack of knowledge of the optimal composition of PRP or its suitability for the proposed indication are some of the reasons behind clinical inconsistencies. The impact of inter‐subject variability in PRP has received less attention in comparison. The purpose of this study was to determine the inter‐subject variability, based on age, in the concentrates prepared by a new blood concentration system. Twenty‐six healthy volunteers of both genders (29‐93 years) were enrolled. Whole blood (WB) was collected from each participant to prepare PRP using the Easy PRP kit. Platelets and white blood cells (WBC) from WB and PRP were analysed after split population by age; patients younger than 65 years (n = 13) and patients ≥65 years old (n = 13). Among the demographic characteristics tested, only age was significantly different between the groups. Cell capture efficiency of the system was specific for each type of blood cell and identical for both age groups. Platelets and WBC in PRP were higher than in WB (P

Antibodies are important biomarkers in clinical diagnostics in addition to being increasingly used for therapeutic purposes. Although numerous methods for their detection and quantification exist, they predominantly require benchtop instruments operated by specialists. To enable the detection of antibodies at point-of-care (POC), the development of simple and rapid assay methods independent of laboratory equipment is of high relevance. In this study, we demonstrate microfluidic thread-based analytical devices (μTADs) as a new platform for antibody detection by means of bioluminescence resonance energy-transfer (BRET) switching sensor proteins. The devices consist of vertically assembled layers including a blood separation membrane and a plastic film with a sewn-in cotton thread, onto which the BRET sensor proteins together with the substrate furimazine have been predeposited. In contrast to intensity-based signaling, the BRET mechanism enables time-independent, ratiometric readout of bioluminescence signals with a digital camera in a darkroom or a smartphone camera with a 3D-printed lens adapter. The device design allows spatially separated deposition of multiple bioluminescent proteins on a single sewn thread, enabling quantification of multiple antibodies in 5 μL of whole blood within 5 min. The bioluminescence response is independent of the applied sample volume within the range of 5-15 μL. Therefore, μTADs in combination with BRET-based sensor proteins represent user-friendly analytical tools for POC quantification of antibodies without any laboratory equipment in a finger prick (5 μL) of whole blood.

Background One of the elements that makes up blood safety is automation of processing whole blood into ingredients In 2017, Reveos was installed in Blood Bank in Lodz. By combining the centrifuge function and the press, Reveos provides full automation of the whole blood separation process, minimizing the risk of human error, thus ensuring accuracy and repeatability. The aim is to compare the results of the quality control of ingredients obtained after the blood separation using the Compomat G5, Optipress II and the Reveos system as well as to compare the level of compliance with the recommendations on the quality of blood components contained in the applicable regulations. Materials and methods 150 units were separated into ingredients using above mentioned systems showing choosen parameters. Results Parameters were determined for Compomat, Optipress i Reveos: plasma: volume 260±13ml, 262±14ml i 239±20ml, platelets: 16, 23 i 23x109/l. RBC: volume 286±13ml, 272±13ml i 295±17ml, hemoglobin 55, 51, 58g/unit., hematocrit 59, 58 i 52%, leukocytes 0,6x109/unit., 0,6x109/unit., 0,1x106/unit. Buffy coat volume: 56,6ml i 56,1ml for Compomat i Optipress and interim platelets volume 37,8ml. The platelet content in the above-mentioned components was 0.8, 0.7, 0.66x1011/unit, respectively. Conclusions The comparison of the quality of ingredients after the blood separation showed slight differences in the parameters determined. The main difference concerns the volume of the obtained plasma and red blood cells. Nevertheless, all the ingredients obtained meet rigorous quality control assumptions and separation methods can be successfully used in routine work. The Reveos system allows to obtain qualitatively good ingredients in a one-step process. This reduces staff interference in the whole process, which may result in minimizing errors during processing of the whole blood.

Developing simple, portable, rapid, and easy-to-use diagnostic technologies is essential for point-of-care (POC) blood molecular testing. Integrated microfluidic devices that include the functionalities of blood separation, microfluidic pumping, and molecular detection are desirable for POC testing; however, current technologies still rely on off-chip sample processing or require bulky equipment. We report a fully-integrated microfluidic diagnostic device, i.e., an integrated pneumatic microfluidic circuit (iPC), that can autonomously pump whole blood, continuously sort blood plasma, and readily enable blood plasma proteomic analysis. The iPC contains vacuum pillars as a vacuum source and waste reservoir, as well as microchannels connecting the pillars as a plasma separator or a flow stabilizer. We combined the iPC and a miniaturized fluorescence microscope to create a portable diagnostic platform that enables fluorescence-based biomarker detection. First, we performed systematic parametric studies to establish design rules for determining the transport and distribution of fluid streams in the iPC. We then demonstrated the capability of the iPC-based diagnostic platform by successfully separating blood plasma from microliter quantities of whole blood while simultaneously quantifying thrombin in blood samples using an aptamer beacon within 5 min of sample injection. Our platform holds potential as a rapid, field-deployable, essentially universal diagnostic tool in POC settings.

Separation of blood plasma or serum from blood is essential for accurate analysis. Conventional blood separation requires instrumentation, reagents, and large sample volumes, limiting this process to laboratory environments with trained personnel. Full implementation of effective blood separation and analysis on microliter sample volumes for point of care (POC) diagnostics has proven extremely challenging resulting in a growing market demand, with common challenges such as expensive device fabrication processes or devices being comprised of materials which are not easily disposable. We developed a membrane-based wicking microfluidic device which is made using a simple fabrication process. This device uses a unique 3D flow channel geometry, fabricated in a polycaprolactone-filled glass microfiber membrane, to efficiently separate microliter sample volumes of blood. Colorimetric assay chemistries were integrated to demonstrate utility of these devices in POC diagnostics. The devices are capable of separating both fresh and anticoagulant-treated blood at microscale sample volumes (

This work introduces the procedure of using non-immunoassay distance-based paper analytical devices (dPADs) to accurately measure any traces of the cardiac troponin I (TnI) in whole blood samples without the use of any external blood separation. This enables a rapid clinical diagnosis and the subsequent follow-up in regard to identifying acute myocardial infarction. These dPADs are designed and constructed to accommodate three parts: (1) a blood separation zone that is immobilized with a hemostatic agent, this no longer requires a blood separation membrane for the isolation of the plasma from the blood element, (2) a pretreatment zone, and (3) a detection zone coated with thymol blue. The quantitative TnI level in the whole blood was determined by measuring the blue color length found in the detection zone, which is proportional to the concentration, owing to the dry protein binding principle. Correspondingly, a mere single drop of human whole blood performs adequately within our proposed method. This reduces both the size of the collection process and the sample volumes needed in the respective medical fields. As we cover all of the optimization studies, our dPADs provide an evaluation of the linearity range from 0.025 to 2.5 ng/mL (R2 = 0.9989) of TnI, with a detection limit as low as 0.025 ng/mL by use of an observation just using the naked eye. To validate the clinical utilities of our proposed method, our dPADs were then applied for the detection of TnI in humans using the whole blood sample of 15 volunteers. A great amount of accuracy was required in this assay because there was no significant difference between both methods, with the confidence level being as high as 95%. This technique also showed that the recoveries ranged from 99.40 to 104.27%, with the highest relative standard deviation being at 3.77%. Thus, our proposed dPADs offer more benefits for a rapid TnI determination.

In this paper, we describe two fabrication methods (well array-based and biopsy punching-based) of gel disks to construct a gel-based point-of-care (POC) diagnosis device for direct colorimetric measurement of human whole blood glucose without any extra blood separation step. The gel disks are made of Polyethylene glycol (PEG) diacrylate (PEG-DA) containing immobilized glucose colorimetric assay reagents. The performances of three types of PEG-DA gel (molecular weight: 575, 3,400, and 10,000) based sensors as well as the two fabrication methods were investigated.•The fabricated devices enabled colorimetric whole blood glucose sensing assay without the need for blood cell separation•The biopsy punching-based gel disk fabrication method provided less variation on the fabricated gel disks, Graphical abstract Image, graphical abstract

A microfluidic chip, which can separate and enrich leukocytes from whole blood, is proposed. The chip has 10 switchback curve channels, which are connected by straight channels. The straight channels are designed to permit the inertial migration effect and to concentrate the blood cells, while the curve channels allow the Dean flow to further classify the blood cells based on the cell sizes. Hydrodynamic suction is also utilized to remove smaller blood cells (e.g., red blood cell (RBC)) in the curve channels for higher separation purity. By employing the inertial migration, Dean flow force, and hydrodynamic suction in a continuous flow system, our chip successfully separates large white blood cells (WBCs) from the whole blood with the processing rates as high as 1 &times, 108 cells/sec at a high recovery rate at 93.2% and very few RBCs (~0.1%).

Interest in developing paper-based devices for point-of-care diagnostics in resource-limited settings has risen remarkably in recent decades. In this paper, we demonstrate what we refer to as “High Yield Passive Erythrocyte Removal” (HYPER) technology, which utilizes capillary forces in a unique cross-flow filtration for the separation of whole blood with performance comparable to centrifuges. As we will demonstrate, state-of-the-art passive blood separation methods implemented in paper-based systems exhibit rapid blood cell clogging on the filtration media or serum outlet and yield only about 10%−30% of the total serum present in the sample. Our innovation results from the inclusion of a differentiation pad, which exploits hydrodynamic effects to reduce the formation of a fouling layer on the blood filtration membrane resulting in more than 60% serum yield with undiluted whole blood as direct input. To demonstrate the effectiveness of the HYPER technology we implement it in a lateral flow system and demonstrate the accurate quantification of vitamin A and iron levels in whole blood samples in 15 minutes.

This chapter presents current efforts to design smart materials for blood separation, which do not rely on molecular sieving only, but on mechanisms of interactions between the membrane and the blood component to isolate. Although concepts have been introduced, there is more to do than has ever been done on this topic. This chapter stresses the need for a specific combination of materials to separate the component of interest from the bloodstream without inducing blood coagulation. PEGylated, zwitterionic and pseudo-zwitterionic materials can all improve the hemocompatibility of the membrane design. But to perform a smart separation, a charge bias has to be introduced by incorporating charged polymers, or a stimuli-responsive polymer has to be grafted which interactions with the blood component are tuned by environmental conditions. Attention is also given to methods for preparing supporting layers, poly(vinylidene fluoride)-based or polypropylene-based. Finally, examples of reported smart blood separations are scrutinized, including the separation of proteins from whole blood, the development of leukocyte depletion or platelet concentration filters. We end this chapter with an identification of the current challenges to overcome to expand the development of smart membranes for blood separation.

Evaluation and diagnosis of blood alterations is a common request for clinical laboratories, requiring a complex technological approach and dedication of health resources. In this paper, we present a microfluidic device that owing to a novel combination of hydrodynamic and dielectrophoretic techniques can separate plasma from fresh blood in a microfluidic channel and for the first time allows optical real-time monitoring of the components of plasma without pre- or post-processing. The microchannel is based on a set of dead-end branches at each side and is initially filled using capillary forces with a 2-μL droplet of fresh blood. During this process, stagnation zones are generated at the dead-end branches and some red blood cells (RBCs) are trapped there. An electric field is then applied and dielectrophoretic trapping of RBCs is used to prevent more RBCs entering into the channel, which works like a sieve. Besides, an electroosmotic flow is generated to sweep the rest of the RBCs from the central part of the channel. Consequently, an RBC-free zone of plasma is formed in the middle of the channel, allowing real-time monitoring of the platelet behavior. To study the generation of stagnation zones and to ensure RBC trapping in the initial constrictions, two numerical models were solved. The proposed experimental design separates up to 0.1 μL blood plasma from a 2-μL fresh human blood droplet. In this study, a plasma purity of 99 % was achieved after 7 min, according to the measurements taken by image analysis. Graphical Abstract Schematics of a real-time plasma monitoring system based on a Hydrodynamic and direct-current insulator-based dielectrophoresis microfluidic channel.

Neutrophil dysfunction is strongly linked to type 2 diabetes mellitus (T2DM) pathophysiology, but the prognostic potential of neutrophil biomarkers remains largely unexplored due to arduous leukocyte isolation methods. Herein, a novel integrated microdevice is reported for single-step neutrophil sorting and phenotyping (chemotaxis and formation of neutrophil extracellular traps (NETosis)) using small blood volumes (fingerprick). Untouched neutrophils are purified on-chip from whole blood directly using biomimetic cell margination and affinity-based capture, and are exposed to preloaded chemoattractant or NETosis stimulant to initiate chemotaxis or NETosis, respectively. Device performance is first characterized using healthy and in vitro inflamed blood samples (tumor necrosis factor alpha, high glucose), followed by clinical risk stratification in a cohort of subjects with T2DM. Interestingly, "high-risk" T2DM patients characterized by severe chemotaxis impairment reveal significantly higher C-reactive protein levels and poor lipid metabolism characteristics as compared to "low-risk" subjects, and their neutrophil chemotaxis responses can be mitigated after in vitro metformin treatment. Overall, this unique and user-friendly microfluidics immune health profiling strategy can significantly aid the quantification of chemotaxis and NETosis in clinical settings, and be further translated into a tool for risk stratification and precision medicine methods in subjects with metabolic diseases such as T2DM. MOE (Min. of Education, S’pore) NMRC (Natl Medical Research Council, S’pore) Accepted version

This paper describes a novel microfluidic chip, including implementation of serially connected filters for improvement of collection efficiency in blood plasma separation. We have previously reported on a blood plasma separation device, comprising serially connected filters, and its effect on plasma collection efficiency. This paper describes improvement of the device structure for biological blood separation and integration of the filters into a microfluidic chip. Plasma collection into an outer reservoir enables use of the separated plasma in further processing, although the ultimate objective is on-chip analysis of plasma constituents, this is not currently possible. Thus, a collection mechanism for the serially connected filters was also developed and implemented. The device developed in this study was able to extract approximately 4 μL of plasma from 50 μL of non-diluted whole blood into an outer reservoir without critical hemolysis. The successfully miniaturized filter device was able to collect both plasma and un-separated blood into outer reservoirs for further processing and will be a useful separation platform for μTAS applications.

A novel device of smart pipette has been suggested to extract and deliver plasma from whole blood in a disposable format. By operating an on-chip disposable micropump, approximately 30 μL of plasma was obtained from 100 μL of whole blood within 5 min without any external equipment for point-of-care blood analysis.

Determining cell mechanical properties is increasingly recognized as a marker-free way to characterize and separate biological cells. This emerging realization has led to the development of a plethora of appropriate measurement techniques. Here, we use a fairly novel approach, deterministic lateral displacement (DLD), to separate blood cells based on their mechanical phenotype with high throughput. Human red blood cells were treated chemically to alter their membrane deformability and the effect of this alteration on the hydrodynamic behaviour of the cells in a DLD device was investigated. Cells of defined stiffness (glutaraldehyde cross-linked erythrocytes) were used to test the performance of the DLD device across a range of cell stiffness and applied shear rates. Optical stretching was used as an independent method for quantifying the variation in stiffness of the cells. Lateral displacement of cells flowing within the device, and their subsequent exit position from the device were shown to correlate with cell stiffness. Data showing how the isolation of leucocytes from whole blood varies with applied shear rate are also presented. The ability to sort leucocyte sub-populations (T-lymphocytes and neutrophils), based on a combination of cell size and deformability, demonstrates the potential for using DLD devices to perform continuous fractionation and/or enrichment of leucocyte sub-populations from whole blood.

A novel noninstrumented technology (ChemTrak AccuMeter) for the quantitative measurement of analytes in biological fluids was reported at the 1990 Oak Ridge Conference (Clin Chem 1990;36:1591-7). This instrument-free technology has been adapted for the quantitative measurement of high-density lipoprotein (HDL). An in situ lipoprotein separation method has been developed and incorporated into the test for HDL determination. The sensitivity of the assay system has been adjusted so that HDL is measured over a clinically significant range of 250 to 1000 mg/L. This compares to a range of 1000 to 4000 mg/L for the total cholesterol in the earlier reported assay. Like the AccuMeter total cholesterol test, the Accumeter HDL test system is self-contained and consolidates blood separation, lipoprotein separation, and specimen measurement into a single step. The test procedure is simple, and the results are accurate.

Most clinical chemistry tests are performed on cell-free serum or plasma. Therefore micro assay devices for blood tests require integrated on-chip microfluidics for separation of plasma or serum from blood. This is achieved by a new blood separation technique based on a microchannel bend structure developed within the collaborative microtele-biochip (/spl mu/TBC) project co-funded by the German Ministry for education and research (BMBF). Different prototype polymer chips have been manufactured with an UV-LIGA process and hot embossing technology. The separation efficiency of these chips has been determined with samples of human whole blood as well as diluted blood samples. The results show different separation efficiencies up to 90% for blood cells and plasma depending on microchannel geometry as well as cell concentration. As compared to present microfluidic devices for the separation of blood cells like filters or filtration by diffusion the microchannel bend is an integrated on-chip blood separation method, which combines the advantages of rapid separation times and a simple geometry.

We propose a MEMS device which integrates a dielectrophoretic blood separation technique with a cytometer and pump for use in low cost blood analysis. Our technical analysis focuses on fluid flow, blood modeling for electrical systems and the dielectrophoresis and cytometry methods. Simulation results of the flow, separation and cytometry are presented.

A blood analysis chip including a power-free micro blood separator capable of separating cells-free plasma from whole blood using a specially designed sub-micron structure and electrochemical electrodes for glucose sensing is developed. Preliminary tests show the possibility of the presented chip.

The development of microfluidic platforms broke the barrier for rampant growth of on-chip biomarker identification in the field of disease diagnostics. Unfortunately, the amplification of nucleic acids directly from raw patient blood has yet to find a simple but complete design on a point-of-care microfluidic device. Here we report a technological approach to detect circulating DNA typically found in whole blood of cancer patients through an on-chip blood-plasma separation, filtration, reagent mixing, and PCR amplification. Raw patient sample blood will first be placed through a self-driven, gravitational and drag powered filtration system, where separation times as low as 10 minutes per 5 μL of blood can be achieved. Plasma separation coupled with nitrocellulose filters will lead to a 99% pure plasma product. Patient plasma can then be properly placed into micro-chambers, where they will be exposed to corresponding lyophilized reagents and enzymes. With each well equipped to an individual micro-heater, concentrated thermo-cycling induces DNA amplification by following golden-standard PCR methodology on a minute-scale basis. Amplification of nucleic acids in the steps indicated above present a simple device design for diagnosis of DNA biomarkers from patient to PCR. To thoroughly accomplish all desired tasks in a direct and efficient manner, the components of an on-chip PCR platform will include a simple yet powerful blood separation method, lyophilized reagent storage, and micro-heated thermo-cycling.

In this work, a new lab-on-a-chip for rapid analysis of low volume blood samples was designed, fabricated and demonstrated for integration of serum separation, hematocrit evaluation, and protein quantitation. Blood separation was achieved using microchannel flow-based separation. A novel method for evaluating hematocrit from microfluidic flow-separated blood samples was developed using gray scale analysis of a point-and-shoot digital photograph of separated blood in a micochannel. Protein quantitation was subsequently performed in a high surface area-to-volume ratio microfluidic chemiluminescent immunoassay using cell depleted serum produced by microfluidic flow-based separation of whole blood samples. All three steps were achieved in a single microchannel with separation of blood samples and hematocrit evaluation in less than 1 min, and protein quantitation in 5 min.

A single-stage disposable channel and seal that provides for leukocyte and granulocyte collection by continuous-flow cell separation (CFCS) has been designed by the IBM Corporation. This paper describes (1) the separation characteristics of whole blood as it responds to varying gravitational (G) forces and flow rates through the channel; (2) the mechanism by which the buffy coat accumulates and is extracted; (3) the efficiency of extraction; (4) those donor and procedural variables that contribute to the final yield; (5) posttransfusion increment response in patients; and (6) the functional integrity of the cells collected.