This paper demonstrates the enrichment of reticulocytes by centrifuging whole blood through aqueous multiphase systems (AMPSs)-immiscible phases of solutions of polymers that form step-gradients in density. The interfaces of an AMPS concentrate cells; this concentration facilitates the extraction of blood enriched for reticulocytes. AMPS enrich reticulocytes from blood from both healthy and hemochromatosis donors. Varying the osmolality and density of the phases of AMPS provides different levels of enrichment and yield of reticulocytes. A maximum enrichment of reticulocytemia of 64 6 3% was obtained from donors with hemochromatosis. When used on peripheral blood from normal donors, AMPS can provide a higher yield of enriched reticulocytes and a higher proportion of reticulocytes expressing CD71 than differential centrifugation followed by centrifugation over Percoll. Blood enriched for reticulocytes by AMPS could be useful for research on malaria. Several species of malaria parasites show a preference to invade young erythrocytes and reticulocytes; this preference complicates in vitro cultivation of these species in human blood. Plasmodium knowlesi malaria parasites invade normal human blood enriched for reticulocytes by AMPSs at a rate 2.2 times greater (P < 0.01) than they invade unenriched blood. Parasite invasion in normal blood enriched by AMPS was 1.8 times greater (P < 0.05) than in blood enriched to a similar reticulocytemia by differential centrifugation followed by centrifugation over Percoll. The enrichment of reticulocytes that are invaded by malaria parasites demonstrates that AMPSs can provide a label-free method to enrich cells for biological research. IntroductionThe characterization of different cellular components of human whole blood plays an important role in medical diagnoses and research [1]. This task is often accomplished by isolating or enriching a specific cellular sub-population from the complex mixture of cells found in whole blood. Whole blood is composed of cells-leukocytes, platelets, and erythrocytes-suspended in protein-rich plasma; each type of cell is useful in the evaluation of the health of a patient. These cells can be further subdivided into important components. For example, the isolation of reticulocytes-immature erythrocytes-is important for research in malaria.Plasmodium (P.) vivax, P. ovale, and P. knowlesi-three causative agents of malaria in humans-preferentially invade human reticulocytes [2][3][4]. The study of these species of malaria suffers from the practical difficulty of in vitro cultures in whole human blood. Their cultivation in blood enriched for reticulocytes would provide concentrated host cells in which the parasites proliferate; continuous cultures of these parasites would make their study much more practical [4][5][6][7]. Enrichment is difficult because reticulocytes exist at a low concentration (0.5-2.5% of erythrocytes) in whole blood [8] and have a short half-life in culture ( 30 hr) [9]. Cryopreservation provides a method to store samples that hav...

In this work we describe a novel and simple self-assembly of colloidal silica beads to create nanofluidic junction between two microchannels. The nanoporous membrane was used to induce ion concentration polarization inside the microchannel and this electrokinetic preconcentration system allowed rapid concentration of DNA samples by ∼1700 times and protein samples by ∼100 times within 5 minutes.

One significant challenge in medical diagnostics lies in the development of label-free methods to separate different cells within complex biological samples. Here we demonstrate a generic, low-power ultrasonic separation technique, able to enrich different cell types based upon their physical properties. For malaria, we differentiate between infected and non-infected red blood cells in a fingerprick-sized drop of blood. We are able to achieve an enrichment of circulating cells infected by the ring stage of the parasite over nonparasitized red blood cells by between two and three orders of magnitude in less than 3 seconds (enabling detection at parasitemia levels as low as 0.0005 %). In a second example, we also show that our methods can be used to enrich different cell types, concentrating Trypanosoma in blood at very low levels of infection, on disposable, low-cost chips.

This comprehensive review serves as a guide for developing scalable and robust liquid biopsies on chip for capture, detection, and analysis of circulating tumor cells (CTCs). Liquid biopsy, the detection of biomarkers from body fluids, has proven challenging because of CTC rarity and the heterogeneity of CTCs shed from tumors. The review starts with the underlying biological mechanisms that make liquid biopsy a challenge before moving into an evaluation of current technological progress. Then, a framework for evaluation of the technologies is presented with special attention to throughput, capture rate, and cell viability for analysis. Technologies for CTC capture, detection, and analysis will be evaluated based on these criteria, with a focus on current approaches, limitations and future directions. The paper provides a critical review for microchip developers as well as clinical investigators to build upon the existing progress towards the goal of designing CTC capture, detection, and analysis platforms.

One significant challenge in medical diagnostics lies in the development of label-free methods to separate different cells within complex biological samples. Here we demonstrate a generic, low-power ultrasonic separation technique, able to enrich different cell types based upon their physical properties. For malaria, we differentiate between infected and non-infected red blood cells in a fingerprick-sized drop of blood. We are able to achieve an enrichment of circulating cells infected by the ring stage of the parasite over nonparasitized red blood cells by between two and three orders of magnitude in less than 3 seconds (enabling detection at parasitemia levels as low as 0.0005 %). In a second example, we also show that our methods can be used to enrich different cell types, concentrating Trypanosoma in blood at very low levels of infection, on disposable, low-cost chips.