Discrimination of leukemic Jurkat cells from normal lymphocytes via novo label-free cytometry based on fluctuation of image gray values.

We introduce a simple, label-free cytometry technique, based on the spatio-temporal fluctuation analysis of pixel gray levels of a cell image utilizing the Gray Level Information Entropy (GLIE) function. In this study, the difference in GLIE random fluctuations and its biophysical etiology in a comparison cell model of leukemic Jurkat cells and human healthy donor lymphocytes was explored. A combination of common bright field microscopy and a unique imaging dish wherein cells are individually held untethered in a picoliter volume matrix of optical chambers was used. Random GLIE fluctuations were found to be greater in malignant Jurkat cells than in benign lymphocytes, while these fluctuations correlate with intracellular vesicle Mean Square Displacement (MSD) values and are inhibited by myosin-2 and adenosine triphosphate (ATP) inhibitors. These results suggest that the incoherent active forces acting on the cytoskeleton which cause mechanical dissipative fluctuation of the cytoskeletal and related intracellular content are the biophysical cellular mechanism behind the GLIE random fluctuation results. Analysis of the results in Jurkat cells and normal lymphocytes suggests the possible potential of this simple and automated label-free cytometry to identify malignancy, particularly in a diagnostic setup of multiple cell examination.

Ultrasound-guided cannulation of the femoral vein in electrophysiological procedures: a systematic review and meta-analysis.

AIMS: In an effort to minimize periprocedural stroke risk, increasingly, electrophysiological (EP) procedures are being performed on anticoagulation. The decrease in stroke has been accompanied by an increase in potentially devastating vascular access complications. Ultrasound guidance for femoral vein cannulation reduces complications in other applications. The aim of this study is to determine the utility of real-time two-dimensional (2D) ultrasound guidance for femoral vein cannulation in EP. METHODS AND RESULTS: A comprehensive literature search of Medline, Embase, Google Scholar, and the Cochrane Central Register of Controlled Trials was performed. Five years of conference abstracts from the Heart Rhythm Society, European Heart Rhythm Association, and European Cardiac Arrhythmia Society were reviewed. Two independent reviewers identified trials comparing ultrasound-guided with standard cannulation in EP procedures. Data were extracted on study design, study size, operator and patient characteristics, use of anticoagulation, vascular complication rates, first-pass success rate, and inadvertent arterial puncture. Four trials, with a total of 4065 subjects, were included in the review, with 1848 subjects in the ultrasound group and 2217 subjects in the palpation group. Ultrasound guidance for femoral vein cannulation was associated with a 60% reduction of major vascular bleeding (relative risk, 0.40; 95% confidence interval, 0.28-0.91). Additionally, there was a 66% reduction in minor vascular complications (relative risk, 0.34; 95% confidence interval, 0.15-0.78). CONCLUSION: The use of real-time 2D ultrasound guidance for femoral vein cannulation decreases access-related bleeding rates and life-threatening vascular complications.

Analysis of the Spectroscopic Aspects of Cationic Dye Basic Orange 21.

Spectroscopic properties of cationic dye basic orange 21 (BO21) in solutions, in solids, and within leukocytes were examined. Results obtained with solutions indicate that influence of variables such as pH, viscosity, salt composition, and various proteins on the absorption spectrum of BO21 is negligible. However, in the presence of heparin, a blue shift (484-465 nm) is observed, which is attributed to the aggregation of BO21 on the polyanion. Applying density functional theory demonstrates that in aqueous solutions (a) the formation of BO21 oligomers is thermodynamically favorable, they are oriented in an antiparallel dipolar arrangement, and their binding energies are lower than those of parallel dipolar arrangements, (b) association between BO21 aggregates and heparin is highly favorable, and (c) the blue shift is due to the mixing of π → π* transitions caused by BO21 molecule stacking. However, when embedded in basophils, the absorption spectra of intracellular BO21 is extremely red-shifted, with two peaks (at 505 and 550 nm) found to be attributed to BO21 and the BO21-heparin complex, respectively, which are intracellularly hosted in nonaqueous environments. Initial evidence of the ability to differentiate between leukocyte types by BO21 is presented.

Real-time quantification of protein expression and translocation at individual cell resolution using imaging-dish-based live cell array.

Cell populations represent intrinsically heterogeneous systems with a high level of spatiotemporal complexity. Monitoring and understanding cell-to-cell diversity is essential for the research and application of intra- and interpopulation variations. Optical analysis of live cells is challenging since both adherent and nonadherent cells change their spatial location. However, most currently available single-cell techniques do not facilitate treatment and monitoring of the same live cells over time throughout multistep experiments. An imaging-dish-based live cell array (ID-LCA) has been developed and produced for cell handling, culturing, and imaging of numerous live cells. The dish is composed of an array of pico scale cavities-pico wells (PWs) embossed on its glass bottom. Cells are seeded, cultured, treated, and spatiotemporally measured on the ID-LCA, while each cell or small group of cells are locally constrained in the PWs. Finally, predefined cells can be retrieved for further evaluation. Various types of ID-LCAs were used in this proof-of-principle work, to demonstrate on-ID-LCA transfection of fluorescently tagged chimeric proteins, as well as the detection and kinetic analysis of their induced translocation. High variability was evident within cell populations with regard to protein expression levels as well as the extent and dynamics of protein redistribution. The association of these parameters with cell morphology and functional parameters was examined. Both the new methodology and the device facilitate research of the translocation process at individual cell resolution within large populations and thus, can potentially be used in high-throughput fashion.

Spatial-temporal correlations in the speckle pattern for the characterization of cellular motion within a 3D object.

Dynamic light scattering analysis has been demonstrated recently to be a promising tool for the assessment of structural changes taking place inside opaque tissue samples. Specifically, quantification of velocity and direction of cellular motion inside spheroids and organoids has attracted much attention as a potent indicator in personalized therapy research. Here, we propose a method for the quantitative extraction of cellular motion, velocity, and direction, by applying a concept of speckle spatial-temporal correlation dynamics. Numerical simulations and experimental results obtained on phantom and biological spheroids are presented.

Correlative analyses of nitric oxide generation rates and nitric oxide synthase levels in individual cells using a modular cell-retaining device.

Nitric oxide (NO) is recognized as one of the major immune system agents involved in the pathogenesis and control of various diseases that may benefit from novel drug development, by exploiting NO signaling pathways and targets. This calls for detection of both intracellular levels of NO and expression of its synthesizing enzymes (NOS) in individual, intact, living cells. Such measurements are challenging, however, due to short half-life, low and fluctuating concentrations of NO, cellular heterogeneity, and inability to trace the same cells over time. The current study presents a device and methodology for correlative analysis of NO generation rates and NOS levels in the same individual cells, utilizing fluorescent imaging followed by immunohistochemistry (IHC). U937 promonocyte cell populations demonstrated significant heterogeneity in their baseline levels, in NO-generation kinetics, and in their response rates to stimuli. Individual cell analysis exposed cell subgroups which showed enhanced NO production upon stimulation, concomitantly with significant up-regulation of inducible NOS (iNOS) levels. Exogenous NO modulated the expression of iNOS in nondifferentiated cells within 1 h, in a dose-dependent manner, while treatment with lysophosphatidylcholine (LPC) enhanced the expression of iNOS, demonstrating a nondependence on NO production.

Co-culture hydrogel micro-chamber array-based plate for anti-tumor drug development at single-element resolution.

In response to the need for reliable cellular models that reflect complex tumor microenvironmental properties, and enable more precise testing of anti-cancer therapeutics effects on humans, a co-culture platform for in-vitro model that enhances the physiology of breast cancer (BC) microenvironment is presented. A six well imaging plate wherein each macro-well contains several separate compartments was designed. Three-dimensional (3D) cancer spheroids are generated and cultured in the inner compartment which is embossed with an array of nano-liter micro-chambers made of hydrogel. Stromal cells are cultured in the outer chambers. The two cell types are cultured side-by-side, sharing a common space, thus enabling extra-cellular communication via secreted molecules. As proof of concept, a model of BC tumor microenvironment was recapitulated by co-cultivating 3D MCF7 spheroids in the presence of tumor-associated macrophages (TAMs). The presence of TAMs induced an aggressive phenotype by promoting spheroid growth, enhancing survivin expression levels and enabling invasive behavior. Moreover, TAMs influenced the response of BC spheroids to cytotoxic treatment as well as hormonal drug therapy, and enhanced the effects of nitric oxide donor. The platform enables time-lapse imaging and treatment without losing spatial location of the measured spheroids, thereby allowing measurements and analysis at individual-object resolution in an easy and efficient manner.

The individual-cell-based cryo-chip for the cryopreservation, manipulation and observation of spatially identifiable cells. I: methodology.

BACKGROUND: Cryopreservation is the only widely applicable method of storing vital cells for nearly unlimited periods of time. Successful cryopreservation is essential for reproductive medicine, stem cell research, cord blood storage and related biomedical areas. The methods currently used to retrieve a specific cell or a group of individual cells with specific biological properties after cryopreservation are quite complicated and inefficient. RESULTS: The present study suggests a new approach in cryopreservation, utilizing the Individual Cell-based Cryo-Chip (i3C). The i3C is made of materials having appropriate durability for cryopreservation conditions. The core of this approach is an array of picowells, each picowell designed to maintain an individual cell during the severe conditions of the freezing--thawing cycle and accompanying treatments. More than 97% of cells were found to retain their position in the picowells throughout the entire freezing--thawing cycle and medium exchange. Thus the comparison between pre-freezing and post-thawing data can be achieved at an individual cell resolution. The intactness of cells undergoing slow freezing and thawing, while residing in the i3C, was found to be similar to that obtained with micro-vials. However, in a fast freezing protocol, the i3C was found to be far superior. CONCLUSIONS: The results of the present study offer new opportunities for cryopreservation. Using the present methodology, the cryopreservation of individual identifiable cells, and their observation and retrieval, at an individual cell resolution become possible for the first time. This approach facilitates the correlation between cell characteristics before and after the freezing--thawing cycle. Thus, it is expected to significantly enhance current cryopreservation procedures for successful regenerative and reproductive medicine.

The individual-cell-based cryo-chip for the cryopreservation, manipulation and observation of spatially identifiable cells. II: functional activity of cryopreserved cells.

BACKGROUND: The cryopreservation and thawing processes are known to induce many deleterious effects in cells and might be detrimental to several cell types. There is an inherent variability in cellular responses among cell types and within individual cells of a given population with regard to their ability to endure the freezing and thawing process. The aim of this study was to evaluate the fate of cryopreserved cells within an optical cryo apparatus, the individual-cell-based cryo-chip (i3C), by monitoring several basic cellular functional activities at the resolution of individual cells. RESULTS: In the present study, U937 cells underwent the freezing and thawing cycle in the i3C device. Then a panel of vital tests was performed, including the number of dead cells (PI staining), apoptotic rate (Annexin V staining), mitochondrial membrane potential (TMRM staining), cytoplasm membrane integrity and intracellular metabolism (FDA staining), as well as post-thawing cell proliferation assays. Cells that underwent the freezing - thawing cycle in i3C devices exhibited the same functional activity as control cells. Moreover, the combination of the multi-parametric analysis at a single cell resolution and the optical and biological features of the device enable an accurate determination of the functional status of individual cells and subsequent retrieval and utilization of the most valuable cells. CONCLUSIONS: The means and methodologies described here enable the freezing and thawing of spatially identifiable cells, as well as the efficient detection of viable, specific, highly biologically active cells for future applications.

Analysis of Cancer Cell Invasion and Anti-metastatic Drug Screening Using Hydrogel Micro-chamber Array (HMCA)-based Plates.

Cancer metastasis is known to cause 90% of cancer lethality. Metastasis is a multistage process which initiates with the penetration/invasion of tumor cells into neighboring tissue. Thus, invasion is a crucial step in metastasis, making the invasion process research and development of anti-metastatic drugs, highly significant. To address this demand, there is a need to develop 3D in vitro models which imitate the architecture of solid tumors and their microenvironment most closely to in vivo state on one hand, but at the same time be reproducible, robust and suitable for high yield and high content measurements. Currently, most invasion assays lean on sophisticated microfluidic technologies which are adequate for research but not for high volume drug screening. Other assays using plate-based devices with isolated individual spheroids in each well are material consuming and have low sample size per condition. The goal of the current protocol is to provide a simple and reproducible biomimetic 3D cell-based system for the analysis of invasion capacity in large populations of tumor spheroids. We developed a 3D model for invasion assay based on HMCA imaging plate for the research of tumor invasion and anti-metastatic drug discovery. This device enables the production of numerous uniform spheroids per well (high sample size per condition) surrounded by ECM components, while continuously and simultaneously observing and measuring the spheroids at single-element resolution for medium throughput screening of anti-metastatic drugs. This platform is presented here by the production of HeLa and MCF7 spheroids for exemplifying single cell and collective invasion. We compare the influence of the ECM component hyaluronic acid (HA) on the invasive capacity of collagen surrounding HeLa spheroids. Finally, we introduce Fisetin (invasion inhibitor) to HeLa spheroids and nitric oxide (NO) (invasion activator) to MCF7 spheroids. The results are analyzed by in-house software which enables semi-automatic, simple and fast analysis which facilitates multi-parameter examination.

Nitric oxide is cytoprotective to breast cancer spheroids vulnerable to estrogen-induced apoptosis.

Estrogen-induced apoptosis has become a successful treatment for postmenopausal metastatic, estrogen receptor-positive breast cancer. Nitric oxide involvement in the response to this endocrine treatment and its influence upon estrogen receptor-positive breast cancer progression is still unclear. Nitric oxide impact on the MCF7 breast cancer line, before and after estrogen-induced apoptosis, was investigated in 3D culture systems using unique live-cell imaging methodologies. Spheroids were established from MCF7 cells vulnerable to estrogen-induced apoptosis, before and after exposure to estrogen. Spheroids derived from estrogen-treated cells exhibited extensive apoptosis levels with downregulation of estrogen receptor expression, low proliferation rate and reduced metabolic activity, unlike spheroids derived from non-treated cells. In addition to basic phenotypic differences, these two cell cluster types are diverse in their reactions to exogenous nitric oxide. A dual effect of nitric oxide was observed in the breast cancer phenotype sensitive to estrogen-induced apoptosis. Nitric oxide, at the nanomolar level, induced cell proliferation, high metabolic activity, downregulation of estrogen receptor and enhanced collective invasion, contributing to a more aggressive phenotype. Following hormone supplementation, breast cancer 3D clusters were rescued from estrogen-induced apoptosis by these low nitric oxide-donor concentrations, since nitric oxide attenuates cell death levels, upregulates survivin expression and increases metabolic activity. Higher nitric oxide concentrations (100nM) inhibited cell growth, metabolism and promoted apoptosis. These results suggest that nitric oxide, in nanomolar concentrations, may inhibit estrogen-induced apoptosis, playing a major role in hormonal therapy. Inhibiting nitric oxide activity may benefit breast cancer patients and ultimately reduce tumor recurrence.

Predictors of Cardiac Rehabilitation Initiation and Adherence in a Multiracial Urban Population.

BACKGROUND: Lack of initiation and adherence to cardiac rehabilitation (CR) remains a persistent problem. We sought to examine predictors of initiation, adherence, and completion of CR in a unique, minority predominant, urban population. METHODS: We included all patients who were first-time referred to the outpatient CR program at Montefiore Medical Center between 1997 and 2010. The indications for referral included acute myocardial infarction, coronary artery disease, heart failure, stable angina, and valvular heart disease. Adherence was defined as attendance of at least 18 sessions of CR, and completion was defined as attendance of 36 sessions. Multivariable logistic regression was utilized to examine the predictors of initiation, adherence, and completion of CR. RESULTS: A total of 590 patients were included (43.9% white and 56.1% nonwhite patients). Among 400 patients who initiated CR, 229 patients (57.3%) attended at least 18 sessions and 140 patients (35.0%) completed all sessions. Initiation of CR was less likely in patients who were nonwhite (OR = 0.66; 95% CI: 0.44-0.97; P = .04) and those who lacked insurance (OR = 0.54; 95% CI: 0.29-0.83; P = .04). Older age was associated with better adherence (OR = 1.04; 95% CI: 1.02-1.07; P < .001). Requirement of a copayment (OR = 0.57; 95% CI: 0.37-0.87; P = .01) was associated with poor adherence. CONCLUSION: In a multiracial population, nonwhite patients and those who did not have insurance were less likely to initiate CR. Younger age and requirement of copayment were independent predictors for poor adherence. Increasing medical insurance coverage and eliminating copayment may improve the participation and adherence of CR.

In situ label-free static cytometry by monitoring spatiotemporal fluctuations of image gray values.

Spatiotemporal fluctuation of homogeneity and randomness of gray values within an image was explored and utilized as a label-free means for cell examination. This was done by utilizing a user-friendly combination of simple bright field microscope and Cytocapture dish, wherein cells are individually held, each within a picoliter optical chamber, forming an array of cells to be repeatedly measured over time and biomanipulated in situ at single-cell resolution. First, the measured gray level information entropy (GLIE) was used and, based on the fact that living cells are not in a state of thermodynamic equilibrium but rather in a metastable state, two fluctuation-sensitive measures were proposed and examined: ASDE­the spatial average of temporal standard deviation (SD) of GLIE, and AA­the average time autocorrelation of GLIE. System performance was validated on cell-free solutions. This was followed by examining the performance of the measures AGLIE, ASDE, and AA to distinguish among individual live-still, dead and live cells from various cell lines, as well as between cells which were and were not induced to differentiate. Results, which were obtained on four types of cells, indicate advantages of the proposed measures which are believed to be significant additions to the microscope-based probe-free toolbox.

Ultrasound-Guided Catheterization of the Femoral Artery: A Systematic Review and Meta-Analysis of Randomized Controlled Trials.

OBJECTIVES: The goal of this meta-analysis was to determine the utility of real-time two-dimensional (2D) ultrasound guidance for femoral artery catheterization. BACKGROUND: Despite the shift toward establishing vascular access via the radial artery rather than the femoral artery, femoral artery cannulation is still frequent in cardiac catheterization. Since vascular complications related to femoral artery cannulation can be quite devastating, preventing these complications is vital. METHODS: A comprehensive literature search of Medline, Embase, Google Scholar, and the Cochrane Central Register of Controlled Trials was performed. Additionally, five years of conference abstracts from critical care, interventional radiology, vascular surgery, and cardiology were reviewed. Two independent reviewers identified prospective, randomized controlled trials comparing ultrasound guidance with traditional palpation techniques of femoral artery catheterization (with or without fluoroscopy). Data were extracted on study design, study size, operator and patient characteristics, complication rates, first-pass success, procedure time, and number of attempts. RESULTS: Four trials with a total of 1422 subjects were included in the review, with 703 subjects in the palpation group and 719 subjects in the ultrasound-guided group. Compared with traditional methods, ultrasound guidance for femoral artery catheterization was associated with 49% reduction in overall complications, including hematoma and accidental venipuncture (relative risk, 0.51; 95% confidence interval, 0.28-0.91). It was also associated with 42% improvement in the likelihood of first-attempt success (relative risk, 1.42; 95% confidence interval, 1.01-2.00). CONCLUSIONS: The use of real-time 2D ultrasound guidance for femoral artery catheterization decreases life-threatening vascular complications and improves first-pass success rate.

Racial disparities in cardiac rehabilitation initiation and the effect on survival.

OBJECTIVES: To examine predictors of initiation and adherence, identify racial disparities, and compare the survival benefit of cardiac rehabilitation between a white and a unique predominantly non-white minority in an urban environment. DESIGN: A retrospective cohort study. SETTING: The outpatient cardiac rehabilitation program at Montefiore Medical Center, Bronx, New York. PATIENTS: Consecutive patients (n = 822) referred to outpatient cardiac rehabilitation were evaluated. METHODS: Baseline characteristics and outcomes were ascertained from medical records. Multivariate logistic regression was used to examine the association among initiation, age, gender, race, reason for referral, and copayment. Kaplan-Meier analysis was performed to evaluate mortality outcomes. MAIN OUTCOME MEASUREMENTS: Racial disparities in rates of initiation, adherence and completion, and survival benefit associated with cardiac rehabilitation. RESULTS: Among 822 patients referred (51.5% non-white minorities, 61.1% male), 59.4% initiated cardiac rehabilitation. Non-white minorities initiated cardiac rehabilitation less often than did white patients (54.4% versus 65.2%, P = .003). After adjustment, white patients were 77.5% more likely to initiate cardiac rehabilitation (odds ratio 1.78; 95% confidence interval 1.13-2.80). Both white populations and non-white minorities who participated in cardiac rehabilitation had a lower risk of death (P = .0022). CONCLUSIONS: In a predominantly minority population, racial disparities exist among cardiac rehabilitation participants. Minorities were less likely to initiate cardiac rehabilitation. Gender, referral patterns, and the presence of copayment did not influence initiation. Cardiac rehabilitation initiation was associated with decreased mortality.

The use of sequential staining for detection of heterogeneous intracellular response of individual Jurkat cells to lysophosphatidylcholine.

Living cells are known to exhibit great morphological, functional, spatial and temporal heterogeneity. Hence, the study of cells in a bulk, whether this bulk is homogenous or heterogeneous, does not provide sufficiently detailed or interpretable results. An advantageous approach would rather be a comprehensive study of cell biological activity in single isolated living cells. In this study, we present an imaging approach for studying pre-apoptotic and very early apoptotic events, during cell death induced by lysophosphatidylcholine (LPC) at the single cell level. The aim of this study is to investigate intracellular events, such as the mitochondrial membrane potential (MMP) and reactive oxygen species (ROS) formation, before and immediately after LPC introduction to the lymphocytes at the level of individual cells. A new protocol of sequential staining was developed to study the relation between early apoptosis signs (PS externalization), MMP changes and intracellular ROS production rates at an individual Jurkat cell resolution. Simultaneous kinetic assessments of MMP, intracellular ROS levels and phosphatidylserine (PS) externalization were performed at a single cell resolution, using Optical LiveCell™ Array technology and image analysis. The parameters were measured and analyzed both before and during exposure to inducers in a Jurkat cell population, including three groups of single cells: spontaneous apoptotic cells, induced apoptotic cells and fully functional living cells. Exogenous LPC caused a heterogeneous intracellular response among Jurkat cells immediately after its introduction. Subgroups of cells with opposite changes of MMP and different kinetics of ROS increase, were revealed within the whole cell population. The subset of apoptosis-induced Jurkat cells, which became apoptotic within 3h after the LPC introduction, exhibited higher initial MMP compared to fully functional or spontaneous apoptotic cells. LPC-induced apoptosis was accompanied by a concomitant increase in intracellular ROS levels. In the present study, a method is described to assess the intracellular events in cells which were initially different in their physiological status. The individual T lymphocytes (Jurkat cells) in vitro have various susceptibilities to LPC effects at the very early stage of contact with the inducer. The apoptotic effect of LPC in individual Jurkat cells is associated with a relatively higher initial MMP before the introduction of the inducer and with a faster ROS formation within the affected cells. Such divergence may be significant in regulating the balance of lymphocyte subsets in pathological sites, either maintaining or preventing the inflammation components of atherosclerosis. We conclude that the presented approach provides the researcher, not only with the cell retaining methodology, but with opportunities to observe and find the distinctive cell subsets within the whole cell population as well, thus helping to define more exactly the role and importance of such sub-populations in physiological or pathological conditions.

A polymer microstructure array for the formation, culturing, and high throughput drug screening of breast cancer spheroids.

Multicellular spheroid models have been recognized as superior to monolayer cell cultures in antitumor drug screening, but their commercial adaptation in the pharmaceutical industry has been delayed, primarily due to technological limitations. The current study presents a new spheroid culture platform that addresses these technical restrictions. The new culturing device is based on a multiwell plate equipped with a glass bottom patterned with an array of UV adhesive microchambers. Each microchamber is designed to accommodate a single spheroid. The system facilitates the simultaneous creation and culturing of a large number of spheroids, as well as screening their response to antitumor drugs. The volume of the spheroids is easily controlled by seeding density. The location of each spheroid is preserved in the same microchamber throughout its growth, treatment with soluble agents, and imaging. The growth ratio parameter, a non-intrusive size analysis of the same spheroid before and after exposure to drugs, was found to be a sensitive indicator for the reaction of MCF7 breast cancer spheroids to cytotoxic drugs. This feature helps reveal the heterogeneity within the spheroid population during the formation process and their drug response, and provides an opportunity to detect specific, highly active or drug-resistant spheroid sub-groups. The advantages of this spheroid-based system make it an efficient drug-screening tool that may be valuable to related fields of research and clinical applications.

Polymer live-cell array for real-time kinetic imaging of immune cells.

Direct quantitative experimental investigations of the function of lymphocytes and other immune cells are challenging due to the cell mobility and the complexity of intercellular communications. In order to facilitate such investigations, an in vitro system is required that is noninvasive and provides kinetic data on cellular responses to challenges such as drug treatments. The present work reports the development of a disposable, inexpensive polymer-made device, the Polymer Live Cell Array (PLCA), for real-time, kinetic analysis of immune cells. The PLCA proved to be optically and biologically compatible, thus individual immune cells can be observed and treated independently without being tethered. The cells share a common space which facilitates cellular communications via secreted molecules or via direct intercellular interactions. These properties facilitate real-time, non-intrusive, repeated measurements of immune cells under multiple experimental treatments.