The application of microbeads to microfluidic systems for enhanced detection and purification of biomolecules.

This paper describes microbead-based microfluidic systems. Several aspects of bead assays in microfluidics make them advantageous for bioassays in simple microchannels, including enhanced surface-to-volume ratio, improved molecular recognition reaction efficiency, and the wide range of surface functionalization available with commercial microbeads. Two-level SU-8 molds are used to fabricate PDMS microchannels that can hydrodynamically trap different types of microbeads, with characteristic dimensions of tens of microns. The use of these microbead-based microfluidic systems in the biosensing of antibodies, toxins and nucleic acids, as well as in antibody purification will be presented and discussed in this paper.

A biomolecular recognition approach for the functionalization of cellulose with gold nanoparticles.

Materials with new and improved functionalities can be obtained by modifying cellulose with gold nanoparticles (AuNPs) via the in situ reduction of a gold precursor or the deposition or covalent immobilization of pre-synthesized AuNPs. Here, we present an alternative biomolecular recognition approach to functionalize cellulose with biotin-AuNPs that relies on a complex of 2 recognition elements: a ZZ-CBM3 fusion that combines a carbohydrate-binding module (CBM) with the ZZ fragment of the staphylococcal protein A and an anti-biotin antibody. Paper and cellulose microparticles with AuNPs immobilized via the ZZ-CBM3:anti-biotin IgG supramolecular complex displayed an intense red color, whereas essentially no color was detected when AuNPs were deposited over the unmodified materials. Scanning electron microscopy analysis revealed a homogeneous distribution of AuNPs when immobilized via ZZ-CBM3:anti-biotin IgG complexes and aggregation of AuNPs when deposited over paper, suggesting that color differences are due to interparticle plasmon coupling effects. The approach could be used to functionalize paper substrates and cellulose nanocrystals with AuNPs. More important, however, is the fact that the occurrence of a biomolecular recognition event between the CBM-immobilized antibody and its specific, AuNP-conjugated antigen is signaled by red color. This opens up the way for the development of simple and straightforward paper/cellulose-based tests where detection of a target analyte can be made by direct use of color signaling.

Skeletal anomalies in reared Senegalese sole Solea senegalensis juveniles: a radiographic approach.

Reared Senegalese sole Solea senegalensis Kaup show a high incidence of vertebral anomalies; however, little is known about its skeletal anomaly profile in the later farming phases. The purpose of this study was to provide a detailed description and quantification of the most common skeletal anomalies in reared Senegalese sole in the juvenile stage by means of computed radiography. A total of 374 Senegalese sole were classified according to the external morphology of the fish as normal or altered and then radiographed in latero-lateral and in dorso-ventral projections. Radiographic evaluation of anomalies focused especially on vertebral body anomalies (VBA) and vertebral column deviations (VCD). The 2 orthogonal projections provided a more complete visualization of the skeleton. Approximately 75% of the individuals showed at least 1 anomaly, while VBA and/or VCD were detected in 48.9% of the specimens. Regarding external morphology, 88% of the fish were categorized as normal, although about 72% of these normal fish displayed abnormalities in radiographies. The most frequent anomalies consisted of deformations of the caudal complex plates (hypurals, parhypural and epural), preurals and caudal vertebrae. Scoliosis was the most prevalent among VCD, affecting the caudal area in almost 15% of the individuals. The anomaly profile at the juvenile stages showed some differences compared to what has been reported previously in earlier stages of development. In light of these results, further investigation into the progression of skeletal anomalies over time and the causative factors at later stages is required.

New insight on vertebral anomalies in cultured Senegalese sole (Solea senegalensis, Kaup) at early stages of development.

Senegalese sole (Solea senegalensis, Kaup) is a promising flatfish species in aquaculture. However, skeletal anomalies are still a great concern in sole farming. Investigation of this issue is crucial to improving larval quality and optimizing production. The aim of this study was to thoroughly assess anomalies in the rachis of reared sole at early developmental stages. Sole (n = 507) were sampled at 31 or 32 days after hatching (dah). The specimens were stained with alcian blue and alizarin red and evaluated for the detection of vertebral deformities. Most fish presented 9:34:3 vertebrae in abdominal, caudal and caudal complex regions, respectively. Remarkably, all specimens showed at least one spinal anomaly. Alterations of neural/haemal elements, as well as deformities of hypurals, parhypural and epural, were recurrent. Vertebral body anomalies and/or vertebral column deviations were identified in 52% of the individuals. Vertebral deformations and fusions were common, especially in caudal complex. 'Minor' anomalies were predominant, and some of the detected disorders might be a result of non-/low-pathological processes. These results contribute a new insight into the main skeletal anomalies affecting cultured sole larvae. Further research is required to determine their impact on fish welfare and external appearances at commercial stages.

Phytophagous insects and natural enemies on Sapindus saponaria L. (Sapindales: Sapindaceae) plants fertilized with or without dehydrated sewage sludge.

Management programs and efficient techniques are necessary to recover degraded ecosystems. The sewage sludge is rich in nitrogen (N) and with the potential to fertilize Sapindus saponaria L. (Sapinales: Sapindaceae), used in the recovery of degraded areas; this can affect the insect fauna. The study's objective was to evaluate, for 24 months, the abundance of chewing insects, dipterans, pollinators, and predators on S. saponaria plants fertilized with or without dehydrated sewage sludge in a degraded area. The experimental design was completely randomized (with the same characteristics) with two treatments (with or without dehydrated sewage sludge) and 24 replicates, each with one plant. The abundance of Anastrepha sp. (Tephritidae), Cerotoma sp. (Chrysomelidae), Curculionidae, Musca domestica L. (Muscidae), Mantis religiosa L. (Mantodea: Mantidae), Oxyopidae, Salticidae, Tettigoniidae (Orthoptera), and Teudis sp. (Anyphaenidae) was higher on fertilized plants. The abundances of Teudis sp. and Tmarus sp. (Thomisidae) and M. religiosa and Teudis sp. were positively correlated with chewing insects and Diptera, respectively. The population increase of insects and spiders on S. saponaria plants fertilized with dehydrated sewage sludge (bigger crowns) has shown to be suitable for recovering degraded areas with a higher number of niches and better food quality, improving the ecological indices of the area.

The Kazal-type inhibitors infestins 1 and 4 differ in specificity but are similar in three-dimensional structure.

Blood coagulation is an important process in haemostasis, and disorders of blood coagulation can lead to an increased risk of haemorrhage and thrombosis. Coagulation is highly conserved in mammals and has been comprehensively studied in humans in the investigation of bleeding or thrombotic diseases. Some substances can act as inhibitors of blood coagulation and may affect one or multiple enzymes throughout the process. A specific thrombin inhibitor called infestin has been isolated from the midgut of the haematophagous insect Triatoma infestans. Infestin is a member of the nonclassical Kazal-type serine protease inhibitors and is composed of four domains, all of which have a short central α-helix and a small antiparallel ß-sheet. Domains 1 and 4 of infestin (infestins 1 and 4) possess specific inhibitory activities. Infestin 1 inhibits thrombin, while infestin 4 is an inhibitor of factor XIIa, plasmin and factor Xa. Here, the structure determination and structural analysis of infestin 1 complexed with trypsin and of infestin 4 alone are reported. Through molecular modelling and docking, it is suggested that the protein-protein binding site is conserved in the infestin 1-thrombin complex compared with other Kazal-type inhibitors. Infestin 4 is able to bind factor XIIa, and the F9N and N11R mutants selected by phage display were shown to be more selective for factor XIIa in comparison to the wild type.

Probing the quality of Ni filled nanoporous alumina templates by magnetic techniques.

Pulsed electrodeposition prepared porous alumina templates with Ni nanowires pore filling ranged from 1 to 100%, depending on the alumina barrier-layer thickness, were probed by continuous wave ferromagnetic resonance at room temperature. For completely filled samples, a single resonance peak was observed in the whole range of angles between the applied magnetic field and normal to the sample plane. Its position was described by Kittel formula that takes into account shape anisotropy of individual Ni wires and dipolar interactions between them. For the samples with lower pore filling the effective anisotropy field decreased and the resonance linewidth in the perpendicular configuration increased. Also a quite intense second peak was observed at lower fields for these samples. These changes are associated with reduction of pore filling percentage that can lead to decrease of dipolar interactions between nanowires and to appearance of magnetic inhomogeneities inside wires.

Sucking insects and their predators on tree canopies of a monocultural stand of Caryocar brasiliense.

Caryocar brasiliense Camb. (Malpighiales: Caryocaraceae) trees are widely distributed throughout the Cerrado ecosystem. The fruits of C. brasiliense trees are used by humans for food and as the main income source in many communities. C. brasiliense conservation is seriously threatened due to habitat loss caused by the land-use change. Sucking insects constitute an important ecological driver that potentially impact C. brasiliense survival in degraded environments. In addition, insects sampling methodologies for application in studies related to the conservation of C. brasiliense are poorly developed. In this study, sucking insects (Hemiptera) and their predators were recorded in three vertical strata of Caryocar brasiliense canopies. The distribution of sucking species showed vertical stratification along the canopy structure of C. brasiliense. The basal part of the canopy had the highest numbers of sucking insects Aphis gossypii (Glover 1877) (Hemiptera: Aphididae) and Bemisia tabaci (Genn. 1889) (Hemiptera: Aleyrodidae), and their predators Chrysoperla sp. (Neuroptera: Chrysopidae), spiders (Araneae), and Zelus armillatus (Lep. & Servi., 1825) (Hemiptera: Reduviidae). Predators' distribution follows the resource availability and preferred C. brasiliense tree parts with a higher abundance of prey.

Behavior and development of Tetranychus ludeni Zacher, 1913 (Acari: Tetranychidae) and physiological stress in genetically modified cotton expressing Cry1F and Cry1Ac proteins.

Genetically modified plants are one of the tactics used in integrated pest management - IPM. There is great concern about the impact of these plants on non-target organisms. On the other hand, there is little information in the literature on the effects of transgenics (Bacillus thuringiensis) Bt on populations of phytophagous mites, and the physiological responses that this attack promotes on plants. The objective of this work was to evaluate the biology of the T. ludeni mite in Bt cotton, expressing the Cry1F and Cry1Ac proteins. To evaluate the behavior of food and oviposition preference of the T. ludeni with Bt cotton and isohybrid. Verify if the physiological stress caused by T. ludeni's attack is differentiated in Bt cotton. The mites were reared in Bt cotton and isohybrid, in a total of 40 replicates in the completely randomized design and the biological cycle was evaluated. The food preference and oviposition analysis were done with 10 replicates, with choice. The physiological stress was evaluated through chlorophyll fluorescence, under greenhouse conditions. The data of the T. ludeni biology were analyzed by Student's t-test, for food and oviposition preference the chi-square test was performed. Regression models were fitted for the fluorescence parameters. The model identity test was used to evaluate the differences between Bt and isohybrid treatments. Cry1F and Cry1Ac proteins have not affected the biology of T. ludeni. The photosynthetic parameters in Bt cotton plants were less influenced by T. ludeni infestation.

Distribution of galling insects and their parasitoids on Caryocar brasiliense tree crowns.

Caryocar brasiliense Camb. (Malpighiales: Caryocaraceae) is widely distributed in the Brazilian savanna and its fruits are used by humans for food, production of cosmetics, lubricants, and in the pharmaceutical industry. This plant is damaged by galling insects. Number of these galling insects and their parasitoids was recorded, in the field (galls) and in the laboratory (adults emerged from the galls), from three C. brasiliense crown heights, during three years. Numbers of adults of Eurytoma sp. (Hymenoptera: Eurytomidae), galling insect (younger attack) and Sycophila sp. (Hymenoptera: Eurytomidae) (a parasitoid of Eurytoma sp.), were greater on the apical parts of C. brasiliense tree crowns. Numbers and groups of Eurytoma sp. globoid galls (older attack) were higher in the median and basal crown parts. The numbers of Eurytoma sp. galls were higher on apical part of C. brasiliense tree crown and also of their parasitoids.

Thrombin inhibitors from different animals.

Venous and arterial thromboembolic diseases are still the most frequent causes of death and disability in high-income countries. Clinical anticoagulants are inhibitors of enzymes involved in the coagulation pathway, such as thrombin and factor X(a). Thrombin is a key enzyme of blood coagulation system, activating the platelets, converting the fibrinogen to the fibrin net, and amplifying its self-generation by the activation of factors V, VIII, and XI. Thrombin has long been a target for the development of oral anticoagulants. Furthermore, selective inhibitors of thrombin represent a new class of antithrombotic agents. For these reasons, a number of specific thrombin inhibitors are under evaluation for possible use as antithrombotic drugs. This paper summarizes old and new interests of specific thrombin inhibitors described in different animals.

Sucking insects and their predators on tree canopies of a monocultural stand of Caryocar brasiliense / Insetos sugadores e seus predadores na copa de Caryocar brasiliense em sistema de monocultura

Abstract Caryocar brasiliense Camb. (Malpighiales: Caryocaraceae) trees are widely distributed throughout the Cerrado ecosystem. The fruits of C. brasiliense trees are used by humans for food and as the main income source in many communities. C. brasiliense conservation is seriously threatened due to habitat loss caused by the land-use change. Sucking insects constitute an important ecological driver that potentially impact C. brasiliense survival in degraded environments. In addition, insects sampling methodologies for application in studies related to the conservation of C. brasiliense are poorly developed. In this study, sucking insects (Hemiptera) and their predators were recorded in three vertical strata of Caryocar brasiliense canopies. The distribution of sucking species showed vertical stratification along the canopy structure of C. brasiliense. The basal part of the canopy had the highest numbers of sucking insects Aphis gossypii (Glover 1877) (Hemiptera: Aphididae) and Bemisia tabaci (Genn. 1889) (Hemiptera: Aleyrodidae), and their predators Chrysoperla sp. (Neuroptera: Chrysopidae), spiders (Araneae), and Zelus armillatus (Lep. & Servi., 1825) (Hemiptera: Reduviidae). Predators' distribution follows the resource availability and preferred C. brasiliense tree parts with a higher abundance of prey.

Resumo Caryocar brasiliense Camb. (Malpighiales: Caryocaraceae) é amplamente distribuído por todo o ecossistema de cerrado. Os frutos de C. brasiliense são utilizados na alimentação humana e constitui uma importante fonte de renda para muitas comunidades. A perda de habitat provocada pelas mudanças de uso da terra coloca em risco a conservação de C. brasiliense. Insetos sugadores constituem um importante fator ecológico que, potencialmente, afeta o fitness de C. brasiliense em ambientes degradados. Além disso, as metodologias de amostragem de insetos para aplicação em estudos relacionados à conservação de C. brasiliense são pouco desenvolvidas. Neste estudo, o número de insetos sugadores (Hemiptera) e seus predadores foram avaliados em três estratos verticais do dossel de C. brasiliense. A distribuição das espécies sugadoras apresentou estratificação vertical ao longo da estrutura do dossel. O estrato basal do dossel apresentou o maior número de insetos sugadores Aphis gossypii (Glover 1877) (Hemiptera: Aphididae) e Bemisia tabaci (Genn. 1889) (Hemiptera: Aleyrodidae), e seus predadores Chrysoperla sp. (Neuroptera: Chrysopidae), aranhas (Araneae) e Zelus armillatus (Lep. & Servi., 1825) (Hemiptera: Reduviidae). Os predadores distribuíram-se de acordo com a disponibilidade de recursos, ocorrendo em maior número nas partes do dossel com maior abundância de suas presas.

Sucking insects and their predators on tree canopies of a monocultural stand of Caryocar brasiliense

Abstract Caryocar brasiliense Camb. (Malpighiales: Caryocaraceae) trees are widely distributed throughout the Cerrado ecosystem. The fruits of C. brasiliense trees are used by humans for food and as the main income source in many communities. C. brasiliense conservation is seriously threatened due to habitat loss caused by the land-use change. Sucking insects constitute an important ecological driver that potentially impact C. brasiliense survival in degraded environments. In addition, insects sampling methodologies for application in studies related to the conservation of C. brasiliense are poorly developed. In this study, sucking insects (Hemiptera) and their predators were recorded in three vertical strata of Caryocar brasiliense canopies. The distribution of sucking species showed vertical stratification along the canopy structure of C. brasiliense. The basal part of the canopy had the highest numbers of sucking insects Aphis gossypii (Glover 1877) (Hemiptera: Aphididae) and Bemisia tabaci (Genn. 1889) (Hemiptera: Aleyrodidae), and their predators Chrysoperla sp. (Neuroptera: Chrysopidae), spiders (Araneae), and Zelus armillatus (Lep. & Servi., 1825) (Hemiptera: Reduviidae). Predators' distribution follows the resource availability and preferred C. brasiliense tree parts with a higher abundance of prey.

Resumo Caryocar brasiliense Camb. (Malpighiales: Caryocaraceae) é amplamente distribuído por todo o ecossistema de cerrado. Os frutos de C. brasiliense são utilizados na alimentação humana e constitui uma importante fonte de renda para muitas comunidades. A perda de habitat provocada pelas mudanças de uso da terra coloca em risco a conservação de C. brasiliense. Insetos sugadores constituem um importante fator ecológico que, potencialmente, afeta o fitness de C. brasiliense em ambientes degradados. Além disso, as metodologias de amostragem de insetos para aplicação em estudos relacionados à conservação de C. brasiliense são pouco desenvolvidas. Neste estudo, o número de insetos sugadores (Hemiptera) e seus predadores foram avaliados em três estratos verticais do dossel de C. brasiliense. A distribuição das espécies sugadoras apresentou estratificação vertical ao longo da estrutura do dossel. O estrato basal do dossel apresentou o maior número de insetos sugadores Aphis gossypii (Glover 1877) (Hemiptera: Aphididae) e Bemisia tabaci (Genn. 1889) (Hemiptera: Aleyrodidae), e seus predadores Chrysoperla sp. (Neuroptera: Chrysopidae), aranhas (Araneae) e Zelus armillatus (Lep. & Servi., 1825) (Hemiptera: Reduviidae). Os predadores distribuíram-se de acordo com a disponibilidade de recursos, ocorrendo em maior número nas partes do dossel com maior abundância de suas presas.

Purification and characterization of patagonfibrase, a metalloproteinase showing alpha-fibrinogenolytic and hemorrhagic activities, from Philodryas patagoniensis snake venom.

Venoms of Colubridae snakes are a rich source of novel compounds, which may have applications in medicine and biochemistry. In the present study, we describe the purification and characterization of a metalloproteinase (patagonfibrase), the first protein to be isolated from Philodryas patagoniensis (Colubridae) snake venom. Patagonfibrase is a single-chain protein, showing a molecular mass of 53,224 Da and an acidic isoelectric point (5.8). It hydrolyzed selectively the Aalpha-chain of fibrinogen and when incubated with fibrinogen or plasma, the thrombin clotting time was prolonged. Prominent hemorrhage developed in mouse skin after intradermal injection of patagonfibrase. When administered into mouse gastrocnemius muscle, it induced local hemorrhage and necrosis, and systemic bleeding in lungs. Patagonfibrase showed proteolytic activity toward azocasein, which was enhanced by Ca(2+) and inhibited by Zn(2+), cysteine, dithiothreitol and Na(2)EDTA. Patagonfibrase impaired platelet aggregation induced by collagen and ADP. Thus, patagonfibrase may play a key role in the pathogenesis of disturbances that occur in P. patagoniensis envenomation, and may be used as a biological tool to explore many facets of hemostasis.

Integrated process for the purification of antibodies combining aqueous two-phase extraction, hydrophobic interaction chromatography and size-exclusion chromatography.

We have evaluated a process incorporating aqueous two-phase extraction, hydrophobic interaction chromatography (HIC) and size-exclusion chromatography (SEC) for the purification of human immunoglobulin G (IgG) from a Chinese hamster ovary (CHO) cell supernatant. These unit operations were chosen not only for allowing the removal of target impurities but also for facilitating the integration of different process units without the need for any conditioning step. Extraction in aqueous two-phase systems (ATPSs), composed of polyethylene glycol (PEG) and sodium citrate, allowed the concentration of the antibodies in the citrate-rich phase and the removal of the most hydrophobic compounds in the PEG-rich phase. An ATPS composed of 10% (w/w) PEG 3350 and 12% (w/w) citrate, at pH 6, allowed the recovery of IgG with a 97% yield, 41% HPLC purity and 72% protein purity. This bottom phase was then directly loaded on a phenyl-Sepharose HIC column. This intermediate purification step allowed the capture of the antibodies using a citrate mobile phase with 99% of the antibody recovered in the elution fractions, with 86% HPLC purity and 91% protein purity. Finally, SEC allowed the final polishing by removing IgG aggregates. HIC-eluted fractions were directly injected in a Superose 6 size-exclusion column affording a 100% pure IgG solution with 90% yield.

A functional and thromboelastometric-based micromethod for assessing crotoxin anticoagulant activity and antiserum relative potency against Crotalus durissus terrificus venom.

The assessment of the capacity of antivenoms to neutralize the lethal activity of snake venoms still relies on traditional rodent in vivo lethality assay. ED50 and LD50 assays require large quantities of venoms and antivenoms, and besides leading to animal suffering. Therefore, in vitro tests should be introduced for assessing antivenom neutralizing capacity in intermediary steps of antivenom production. This task is facilitated when one key lethal toxin is identified. A good example is crotoxin, a ß-neurotoxin phospholipase A2-like toxin that presents anticoagulant activity in vitro and is responsible for the lethality of venoms of Crotalus durissus snakes. By using rotational thromboelastometry, we reported recently one sensitive coagulation assay for assessing relative potency of the anti-bothropic serum in neutralizing procoagulant activity of Bothrops jararaca venom upon recalcified factor-XII-deficient chicken plasma samples (CPS). In this study, we stablished conditions for determining relative potency of four batches of the anti-crotalic serum (ACS) (antagonist) in inactivating crotoxin anticoagulant activity in CPS (target) simultaneously treated with one classical activator of coagulation (agonists). The correlation coefficient (r) between values related the ACS potency in inactivating both in vitro crotoxin anticoagulant activity and the in vivo lethality of whole venom (ED50) was 0.94 (p value < 0.05). In conclusion, slowness in spontaneous thrombin/fibrin generation even after recalcification elicit time lapse sufficient for elaboration of one dose-response curve to pro- or anti-coagulant agonists in CPS. We propose this methodology as an alternative and sensitive assay for assessing antivenom neutralizing ability in plasma of immunized horses as well as for in-process quality control.

Affinity partitioning of human antibodies in aqueous two-phase systems.

The partitioning of human immunoglobulin (IgG) in a polymer-polymer and polymer-salt aqueous two-phase system (ATPS) in the presence of several functionalised polyethylene glycols (PEGs) was studied. As a first approach, the partition studies were performed with pure IgG using systems in which the target protein remained in the bottom phase when the non-functionalised systems were tested. The effect of increasing functionalised PEG concentration and the type of ligand were studied. Afterwards, selectivity studies were performed with the most successful ligands first by using systems containing pure proteins and an artificial mixture of proteins and, subsequently, with systems containing a Chinese hamster ovary (CHO) cells supernatant. The PEG/phosphate ATPS was not suitable for the affinity partitioning of IgG. In the PEG/dextran ATPS, the diglutaric acid functionalised PEGs (PEG-COOH) displayed great affinity to IgG, and all IgG could be recovered in the top phase when 20% (w/w) of PEG 150-COOH and 40% (w/w) PEG 3350-COOH were used. The selectivity of these functionalised PEGs was evaluated using an artificial mixture of proteins, and PEG 3350-COOH did not show affinity to IgG in the presence of typical serum proteins such as human serum albumin and myoglobin, while in systems with PEG 150-COOH, IgG could be recovered with a yield of 91%. The best purification of IgG from the CHO cells supernatant was then achieved in a PEG/dextran ATPS in the presence of PEG 150-COOH with a recovery yield of 93%, a purification factor of 1.9 and a selectivity to IgG of 11. When this functionalised PEG was added to the ATPS, a 60-fold increase in selectivity was observed when compared to the non-functionalised systems.

Determination of partition coefficients of biomolecules in a microfluidic aqueous two phase system platform using fluorescence microscopy.

Aqueous two phase systems (ATPS) offer great potential for selective separation of a wide range of biomolecules by exploring differences in molecular solubility in each of the two immiscible phases. However, ATPS use has been limited due to the difficulty in predicting the behavior of a given biomolecule in the partition environment together with the empirical and time-consuming techniques that are used for the determination of partition and extraction parameters. In this work, a fast and novel technique based on a microfluidic platform and using fluorescence microscopy was developed to determine the partition coefficients of biomolecules in different ATPS. This method consists of using a microfluidic device with a single microchannel and three inlets. In two of the inlets, solutions containing the ATPS forming components were loaded while the third inlet was fed with the FITC tagged biomolecule of interest prepared in milli-Q water. Using fluorescence microscopy, it was possible to follow the location of the FITC-tagged biomolecule and, by simply varying the pumping rates of the solutions, to quickly test a wide variety of ATPS compositions. The ATPS system is allowed 4min for stabilization and fluorescence micrographs are used to determine the partition coefficient.The partition coefficients obtained were shown to be consistent with results from macroscale ATPS partition. This process allows for faster screening of partition coefficients using only a few microliters of material for each ATPS composition and is amenable to automation. The partitioning behavior of several biomolecules with molecular weights (MW) ranging from 5.8 to 150kDa, and isoelectric points (pI) ranging from 4.7 to 6.4 was investigated, as well as the effect of the molecular weight of the polymer ATPS component.