Multicenter evaluation of TB-SPRINT 59-Plex Beamedex®: accuracy and cost analysis.

BACKGROUND: Molecular tests can allow the rapid detection of tuberculosis (TB) and multidrug-resistant TB (MDR-TB). TB-SPRINT 59-Plex Beamedex® is a microbead-based assay developed for the simultaneous spoligotyping and detection of MDR-TB. The accuracy and cost evaluation of new assays and technologies are of great importance for their routine use in clinics and in research laboratories. The aim of this study was to evaluate the performance of TB-SPRINT at three laboratory research centers in Brazil and calculate its mean cost (MC) and activity-based costing (ABC). METHODS: TB-SPRINT data were compared with the phenotypic and genotypic profiles obtained using Bactec™ MGIT™ 960 system and Genotype® MTBDRplus, respectively. RESULTS: Compared with MGIT, the accuracies of TB-SPRINT for the detection of rifampicin and isoniazid resistance ranged from 81 to 92% and 91.3 to 93.9%, respectively. Compared with MTBDRplus, the accuracies of TB-SPRINT for rifampicin and isoniazid were 99 and 94.2%, respectively. Moreover, the MC and ABC of TB-SPRINT were USD 127.78 and USD 109.94, respectively. CONCLUSION: TB-SPRINT showed good results for isoniazid and rifampicin resistance detection, but still needs improvement to achieve In Vitro Diagnostics standards.

Comparative evaluation of the cost and efficiency of four types of sexing methods for the production of dairy female calves.

This study was conducted to evaluate and compare the economic benefits of different embryo sexing methods, based on the cost per female dairy calf produced. Female calves were produced from four kinds of female embryos: (1) those collected from superstimulated donors at 7-8 days after artificial insemination (AI) with X-sorted semen; (2) those sex-determined by loop-mediated isothermal amplification assay of a biopsy sample of embryos collected from superstimulated donors after AI with conventional unsorted semen; (3) those obtained by invitro embryo production (IVEP), using X-sorted semen and in vitro-matured oocytes collected from donors by ovum pick-up (OPU); and (4) those obtained by IVEP, using X-sorted semen and oocytes collected by OPU after dominant follicle ablation and follicle growth stimulation of the donors. The respective productivities of female calves per technical service and the total production cost per female calf of each sexing method were compared. The production cost per female calf (66,537 JPY), as calculated from the number of female calves per service (1.30), pregnancy rate of transfer (42.9%), rate of female calves obtained (92.9%), and total cost of the method (56,643 JPY plus embryo transfer fee), was less for IVEP with X-sorted semen and follicular growth-stimulated (FGS) oocytes than for the other groups (P < 0.05). The results demonstrate that embryo production with X-sorted semen and FGS oocytes provides a more efficient method for producing female calves than the other embryo sexing methods.

Multicenter validation of a simplified method for paroxysmal nocturnal hemoglobinuria screening.

BACKGROUND: Paroxysmal nocturnal hemoglobinuria (PNH) diagnostic guidelines recommend single-tube five- to six-color or two-tube four-color assays. PNH clones are detectable in only a fraction of patients at risk, and screening for new PNH cases can be complex and expensive. In this multicenter study, we have validated a simplified, one-tube two-color FLAER-based assay suitable for PNH screening. METHODS: Six laboratories received samples containing spiked PNH leukocyte clones to be analyzed in parallel with a common six-color cocktail (FLAER/CD24/CD45/CD64/CD15/CD14) and a simplified two-color mixture (FLAER/CD15), a shared calibration procedure, and a common analysis protocol. Replicate precision and sensitivity tests were performed on PNH patients, from undiluted to 1:10 000. Specificity tests were performed on normal donors to identify the possible sources of artifacts. RESULTS: The performance comparison between six-color and two-color assays showed an excellent agreement for granulocyte PNH clones. Dilution experiments showed an accurate detectability down to 0.01% sensitivity level for granulocyte PNH clones and to 1% for monocytes. Specificity experiments disclosed that basophils and platelets can contaminate the monocyte gate and generate false PNH events. CONCLUSIONS: A simplified two-color (FLAER/CD15) PNH screening test has been validated in a highly standardized multicenter study and proved feasible and effective in ongoing regional programs. Precision, sensitivity, and specificity of the simplified test for granulocytes were comparable to the more complex and expensive six-color assay and applicable for screening also in peripheral laboratories. The diagnostic confirmation of PNH should be always performed by a reference center using the established technique on all cell lineages.

Cost-effectiveness of a new system in ruling out negative urine cultures on the day of administration.

Urine samples account for a significant part of the workload in clinical microbiology laboratories. However, the culture process is time-consuming and a large proportion is reported as negative. To reduce unnecessary culture procedures and speed up the reporting of negative results, a reliable screening method is needed. For this purpose, urine samples submitted to our clinical microbiology laboratory were simultaneously screened by a flow cytometry method (Sysmex UF-1000i, Japan). During screening, the evaluation of various combinations of leucocytes and bacteria cut-offs demonstrated that cut-offs of 30 and 50/µL, respectively, were the best threshold values to reach a 100% negative predictive value (NPV) with a culture reduction rate of 44.8% in adults and 61.9% in children between the ages of 6 and 17 years. With the culture reduction rates mentioned above, the screening method has provided at least 24% savings in expenditures of the routine clinical microbiology laboratory. Since we did not reach such an NPV with any combinations of screening parameters in children younger than 5 years of age, we recommend cultivation of all urine samples in those patients without a screening step. In conclusion, Sysmex UF-1000i as a screening method was capable of improving the efficiency of the routine microbiology laboratory by providing negative results in a few minutes in children greater than 6 years of age and in adults.

High-throughput miniaturized microfluidic microscopy with radially parallelized channel geometry.

In this article, we present a novel approach to throughput enhancement in miniaturized microfluidic microscopy systems. Using the presented approach, we demonstrate an inexpensive yet high-throughput analytical instrument. Using the high-throughput analytical instrument, we have been able to achieve about 125,880 cells per minute (more than one hundred and twenty five thousand cells per minute), even while employing cost-effective low frame rate cameras (120 fps). The throughput achieved here is a notable progression in the field of diagnostics as it enables rapid quantitative testing and analysis. We demonstrate the applicability of the instrument to point-of-care diagnostics, by performing blood cell counting. We report a comparative analysis between the counts (in cells per µl) obtained from our instrument, with that of a commercially available hematology analyzer.

Acoustofluidic Fluorescence Activated Cell Sorter.

Selective isolation of cell subpopulations with defined biological characteristics is crucial for many biological studies and clinical applications. In this work, we present the development of an acoustofluidic fluorescence activated cell sorting (FACS) device that simultaneously performs on-demand, high-throughput, high-resolution cell detection and sorting, integrated onto a single chip. Our acoustofluidic FACS device uses the "microfluidic drifting" technique to precisely focus cells/particles three dimensionally and achieves a flow of single-file particles/cells as they pass through a laser interrogation region. We then utilize short bursts (150 µs) of standing surface acoustic waves (SSAW) triggered by an electronic feedback system to sort fluorescently labeled particles/cells with desired biological properties. We have demonstrated continuous isolation of fluorescently labeled HeLa cells from unlabeled cells at a throughput of ∼1200 events/s with a purity reaching 92.3 ± 3.39%. Furthermore, 99.18% postsort cell viability indicates that our acoustofluidic sorting technique maintains a high integrity of cells. Therefore, our integrated acoustofluidic FACS device is demonstrated to achieve two-way cell sorting with high purity, biocompatibility, and biosafety. We believe that our device has significant potential for use as a low-cost, high-performance, portable, and user-friendly FACS instrument.

Rapid cytometric antibiotic susceptibility testing utilizing adaptive multidimensional statistical metrics.

Flow cytometry holds promise to accelerate antibiotic susceptibility determinations; however, without robust multidimensional statistical analysis, general discrimination criteria have remained elusive. In this study, a new statistical method, probability binning signature quadratic form (PB-sQF), was developed and applied to analyze flow cytometric data of bacterial responses to antibiotic exposure. Both sensitive lab strains (Escherichia coli and Pseudomonas aeruginosa) and a multidrug resistant, clinically isolated strain (E. coli) were incubated with the bacteria-targeted dye, maltohexaose-conjugated IR786, and each of many bactericidal or bacteriostatic antibiotics to identify changes induced around corresponding minimum inhibition concentrations (MIC). The antibiotic-induced damages were monitored by flow cytometry after 1-h incubation through forward scatter, side scatter, and fluorescence channels. The 3-dimensional differences between the flow cytometric data of the no-antibiotic treated bacteria and the antibiotic-treated bacteria were characterized by PB-sQF into a 1-dimensional linear distance. A 99% confidence level was established by statistical bootstrapping for each antibiotic-bacteria pair. For the susceptible E. coli strain, statistically significant increments from this 99% confidence level were observed from 1/16x MIC to 1x MIC for all the antibiotics. The same increments were recorded for P. aeruginosa, which has been reported to cause difficulty in flow-based viability tests. For the multidrug resistant E. coli, significant distances from control samples were observed only when an effective antibiotic treatment was utilized. Our results suggest that a rapid and robust antimicrobial susceptibility test (AST) can be constructed by statistically characterizing the differences between sample and control flow cytometric populations, even in a label-free scheme with scattered light alone. These distances vs paired controls coupled with rigorous statistical confidence limits offer a new path toward investigating initial biological responses, screening for drugs, and shortening time to result in antimicrobial sensitivity testing.

A Cost-Effective and Labor-Saving Method for Detecting Human Leukocyte Antigen B27 Status via Sequence-Encoded Fluorescence Amplification Assay.

Identification of human leukocyte antigen B27 (HLA-B27) by flow cytometry (FCM) has been widely applied in clinical practice for auxiliary diagnosis of ankylosing spondylitis (AS). However, FCM requires freshly prepared samples and relies on expensive equipment, reagents, and an experienced operator. To provide a cheaper and more convenient method for HLA-B27 detection, we proposed a new method termed sequence-encoded fluorescence amplification assay (SEFA), which specially recognized sequences of HLA-B27 gene (HLA-B∗27) covering current common subtypes in a single closed tube. SEFA could detect as low as 10 pg (equal to 3 copies) genomic DNA per reaction and distinguish HLA-B∗27 from other HLA-B alleles with highly similar sequences. A total of 288 clinical samples were tested by SEFA, including 181 patients with AS and 107 healthy controls. Compared with the detection results from FCM, two controversial samples of patients with AS were obtained and further confirmed to be consistent with SEFA by Sanger sequencing, indicating that this method was more accurate than FCM. Moreover, SEFA could detect HLA-B27 status by using supernatant from crude extract of 10-µL blood without commercial reagents. Overall, SEFA has the potential to be an alternative for HLA-B27 identification with the advantage of convenience and low cost, especially suitable for early diagnosis of AS in areas with limited medical resources.

Clinical applicability and prognostic significance of molecular response assessed by fluorescent-PCR of immunoglobulin genes in multiple myeloma. Results from a GEM/PETHEMA study.

Minimal residual disease monitoring is becoming increasingly important in multiple myeloma (MM), but multiparameter flow cytometry (MFC) and allele-specific oligonucleotide polymerase chain reaction (ASO-PCR) techniques are not routinely available. This study investigated the prognostic influence of achieving molecular response assessed by fluorescent-PCR (F-PCR) in 130 newly diagnosed MM patients from Grupo Español Multidisciplinar de Melanoma (GEM)2000/GEM05 trials (NCT00560053, NCT00443235, NCT00464217) who achieved almost very good partial response after induction therapy. As a reference, we used the results observed with simultaneous MFC. F-PCR at diagnosis was performed on DNA using three different multiplex PCRs: IGH D-J, IGK V-J and KDE rearrangements. The applicability of F-PCR was 91·5%. After induction therapy, 64 patients achieved molecular response and 66 non-molecular response; median progression-free survival (PFS) was 61 versus 36 months, respectively (P = 0·001). Median overall survival (OS) was not reached (NR) in molecular response patients (5-year survival: 75%) versus 66 months in the non-molecular response group (P = 0·03). The corresponding PFS and OS values for patients with immunophenotypic versus non-immunophenotypic response were 67 versus 42 months (P = 0·005) and NR (5-year survival: 95%) versus 69 months (P = 0·004), respectively. F-PCR analysis is a rapid, affordable, and easily performable technique that, in some circumstances, may be a valid approach for minimal residual disease investigations in MM.

One-dimensional acoustic standing waves in rectangular channels for flow cytometry.

Flow cytometry has become a powerful analytical tool for applications ranging from blood diagnostics to high throughput screening of molecular assemblies on microsphere arrays. However, instrument size, expense, throughput, and consumable use limit its use in resource poor areas of the world, as a component in environmental monitoring, and for detection of very rare cell populations. For these reasons, new technologies to improve the size and cost-to-performance ratio of flow cytometry are required. One such technology is the use of acoustic standing waves that efficiently concentrate cells and particles to the center of flow channels for analysis. The simplest form of this method uses one-dimensional acoustic standing waves to focus particles in rectangular channels. We have developed one-dimensional acoustic focusing flow channels that can be fabricated in simple capillary devices or easily microfabricated using photolithography and deep reactive ion etching. Image and video analysis demonstrates that these channels precisely focus single flowing streams of particles and cells for traditional flow cytometry analysis. Additionally, use of standing waves with increasing harmonics and in parallel microfabricated channels is shown to effectively create many parallel focused streams. Furthermore, we present the fabrication of an inexpensive optical platform for flow cytometry in rectangular channels and use of the system to provide precise analysis. The simplicity and low-cost of the acoustic focusing devices developed here promise to be effective for flow cytometers that have reduced size, cost, and consumable use. Finally, the straightforward path to parallel flow streams using one-dimensional multinode acoustic focusing, indicates that simple acoustic focusing in rectangular channels may also have a prominent role in high-throughput flow cytometry.

A much convenient and economical method to harvest a great number of microglia.

Microglia, implicating in such neuro-pathologies as brain inflammation, neurodegeneration, glioma, and neurogenesis, play an important role in central nervous system. Advanced research on microglia is crucial in exploring the neuro-pathology and neuro-physiology of these diseases, so how to culture large numbers of microglia in vitro becomes the base of a research. The wildly used method, at present, obtaining microglia from murine cannot fulfill the requirement of research, costing too much time and needing too many rats. We intend to introduce an optimized method that can harvest large quantities of microglia with high purity. Neonatal 2-3 days old Wistar rats were sacrificed and the cerebral cortices were trypsinized. We primarily cultured mixed cortical cells for 8-10 days. The microglia were harvested from the liquid supernatant; the left cells in the mixed cortical glial culture were passaged at a 1:2 density. After another 8-10 days of culture, microglia were collected again. And then, we passaged the left cells again for acquiring microglia from the third collection. We did not add additional mitogens in the experiment. At last, on average, 7.0 × 10(6) microglia were collected from one neonatal rat. By this modified method, much more microglia can be effectively and easily harvested comparing with the usual protocol before. We compared the characteristics of microglia harvested from these three passages, such as morphology, phenotype, purity, and abilities on proliferation, secretion, and phagocytosis. The cells presented typical microglia morphology, having phenotype markers of CD11b/c and CD45. The microglia from these three passages retained similar phagocytosis and secretion functions. Expanded population of microglia for investigation can be provided by this easy method in a short time with little cost and few rats.

The utility of end-of-treatment bone marrow aspirates in pediatric acute lymphoblastic leukemia: a quality improvement project.

We reviewed the use, results and costs of end-of-treatment bone marrow aspirates (EOTBMAs) performed locally in patients diagnosed with ALL between 2000 and 2005. Of 193 patients, 188(97%) received EOTBMAs. Though 15/188(8.0%) patients experienced relapse at a median time of 1.1 years (range 0.1-4 years), no sign of relapse was detected on any EOTBMA. After communication of results to clinical staff, only 2/17 (12%) of patients with ALL finishing treatment in the subsequent 5 months received an EOTBMA (P < 0.0001). Our results confirm the futility of EOTBMAs in a large contemporary cohort. Disseminating local results may help ensure adherence to best practices.

Development of a five-plex flow cytometric immunoassay for the simultaneous detection of six coccidiostats in feed and eggs.

Coccidiostats are the only veterinary drugs still permitted to be used as feed additives to treat poultry for coccidiosis. To protect consumers, maximum levels for their presence in food and feed have been set by the European Union (EU). To monitor these coccidiostats, a rapid and inexpensive screening method would be a useful tool. The development of such a screening method, using a flow cytometry-based immunoassay, is described. The assay uses five sets of colour-coded paramagnetic microspheres for the detection of six selected priority coccidiostats. Different coccidiostats, with and without carrier proteins, were covalently coupled onto different bead sets and tested in combination with polyclonal antisera and with a fluorescent-labelled secondary antibody. The five optimal combinations were selected for this multiplex and a simple-to-use sample extraction method was applied for screening blank and spiked eggs and feed samples. A very good correlation (r ranging from 0.995 to 0.999) was obtained with the responses obtained in two different flow cytometers (Luminex 100 and FLEXMAP 3D). The sensitivities obtained were in accordance with the levels set by the EU as the measured limits of detection for narasin/salinomycin, lasalocid, diclazuril, nicarbazin (4,4'-dinitrocarbanilide) and monensin in eggs were 0.01, 0.1, 0.5, 53 and 0.1 µg/kg and in feed 0.1, 0.2, 0.3, 9 and 1.5 µg/kg, respectively.

Immune status monitoring of HIV/AIDS patients in resource-limited settings: a review with an emphasis on CD4+ T-lymphocyte determination.

CD4 T-lymphocytes play a vital role in maintaining the integrity of the human immune system. They are also the primary target cells for human immunodeficiency virus (HIV). The progressive depletion of these cells eventually results in weakening of the host's immune ability to fight against any pathogen, thus rendering the host susceptible to infections and leading ultimately to death of patients in the terminal stage of acquired immune deficiency syndrome (AIDS). Although several clinical and laboratory parameters have been used for monitoring disease progression and the effectiveness of HIV antiretroviral therapy (ART), it is the simple measurement of CD4+ T-lymphocytes that remains the single and most important parameter for management of HIV-infected patients in resource-limited settings. To date, flow cytometer is considered to be the most accepted technology for both percentage and absolute CD4+ T-lymphocyte determination because of its accuracy, precision and reproducibility. However, flow cytometer based CD4 testing is relatively expensive, complex and thus technically demanding. Simple innovative approaches applicable to the conventional flow cytometric system and new technologies have been successfully developed to increase cost saving especially for use in resource-challenged settings. Principles of the existing dual- and single-platform approaches as well as several affordable CD4 measurement technologies are discussed along with both internal and external quality control systems in the management of laboratories performing CD4 testing.

Rapid and cheap prototyping of a microfluidic cell sorter.

Development of a microfluidic device is generally based on fabrication-design-fabrication loop, as, unlike the microelectronics design, there is no rigorous simulation-based verification of the chip before fabrication. This usually results in extremely long, and hence expensive, product development cycle if micro/nano fabrication facilities are used from the beginning of the cycle. Here, we illustrate a novel approach of device prototyping that is fast, cheap, reliable, and most importantly, this technique can be adopted even if no state-of-the-art microfabrication facility is available. A water-jet machine is used to cut the desired microfluidic channels into a thin steel plate which is then used as a template to cut the channels into a thin sheet of a transparent and cheap polymer material named Surlyn® by using a Hot Knife™. The feature-inscribed Surlyn sheet is bonded in between two microscope glass slides by utilizing the techniques which has been being used in curing polymer film between dual layer automotive glasses for years. Optical fibers are inserted from the sides of chip and are bonded by UV epoxy. To study the applicability of this prototyping approach, we made a basic microfluidic sorter and tested its functionalities. Sample containing microparticles is injected into the chip. Light from a 532-nm diode laser is coupled into the optical fiber that delivers light to the interrogation region in the channel. The emitted light from the particle is collected by a photodiode (PD) placed over the detection window. The device sorts the particles into the sorted or waste outlets depending on the level of the PD signal. We used fluorescent latex beads to test the detection and sorting functionalities of the device. We found that the system could detect all the beads that passed through its geometric observation region and could sort almost all the beads it detected.

Reducing costs in flow-cytometric counting of residual white blood cells in blood products: utilization of a single-platform bead-free flow-rate calibration method.

BACKGROUND: Commercial flow-cytometric methods for counting residual white blood cells (rWBCs) in leukoreduced blood products use calibration beads for estimation of the measured sample volume. A bead-free flow-rate calibration method is developed and validated. STUDY DESIGN AND METHODS: The analyzed volume was calculated by acquisition time (ACQ). Twenty-nine spiking series of red blood cell (RBC) or platelet (PLT) products were prepared containing levels ranging from 0.08 × 10(6) up to 2048 × 10(6) WBCs/L. Nearly WBC-free triple-leukofiltered RBCs or PLT concentrates (PCs) served as background. Propidium iodide (PI) was used to identify rWBCs. Five RBC series were compared against a commercially available kit (LeukoSure, Beckman Coulter). Routine capabilities were tested on 41 RBC and 92 PC samples of two independent transfusion services. RESULTS: The lower detection limit in RBC was 0.08 × 10(6) rWBCs/L for ACQ and 0.16 for LeukoSure. Criteria for linearity, accuracy, and precision were fulfilled within the range of 0.5 × 10(6) to 512 × 10(6) WBCs/L. For PCs, all these criteria were fulfilled between 0.5 × 10(6) and 32 × 10(6) rWBCs/L (lower detection limit of 0.25) for PI. ACQ and LeukoSure agreed sufficiently (81%) when tested on routine RBCs or PCs. CONCLUSION: A residual WBC count of fewer than 0.5 × 10(6) WBCs/L can be accurately counted using the ACQ approach at a total reagent cost of less than 0.5€ per sample.

Simplified murine multipotent progenitor isolation scheme: Establishing a consensus approach for multipotent progenitor identification.

The mouse hematopoietic system has served as a paradigm for analysis of developmental fate decisions in tissue homeostasis and regeneration. However, multiple immunophenotypic definitions of, and sometimes divergent nomenclatures used to classify, murine multipotent progenitors (MPPs) have emerged in the field over time. This has created significant confusion and inconsistency in the hematology field. To facilitate easier comparison of murine MPP phenotypes between research laboratories, a working group of four International Society for Experimental Hematology (ISEH) members with extensive experience studying the functional activities associated with different MPP phenotypic definitions reviewed the current state of the field with the goal of developing a position statement toward a simplified and unified immunophenotypic definition of MPP populations. In November of 2020, this position statement was presented as a webinar to the ISEH community for discussion and feedback. Hence, the Simplified MPP Identification Scheme presented here is the result of curation of existing literature, consultation with leaders in the field, and crowdsourcing from the wider experimental hematology community. Adoption of a unified definition and nomenclature, while still leaving room for individual investigator customization, will benefit scientists at all levels trying to compare these populations between experimental settings.

Rapid, sensitive and cost-effective determination of immune checkpoint inhibitor activity using a magnetic bead-based binding assay.

Immune checkpoint Inhibitors (ICIs) are effective immunno-therapeutic agents for cancer. Rapid and sensitive determination of the blocking activity of ICIs is important for ICIs development and immunological research. Among various immune checkpoint (IC) binding assays, cell-based binding assays are widely regarded, and the functional ELISA is a convenient alternative. However, these methodologies are limited by time-consuming preparation of cell lines stably expressing IC molecules, or long turnaround time with high cost. In this study, two magnetic bead based binding assays were developed to evaluate activity of ICIs, which was determined by a soluble ligand/bead immobilized receptor based binding assay (sL/bR binding assay) that assessed efficacy to block binding of one soluble IC ligand on its cognate receptor immobilized beads, or by a soluble receptor/bead immobilized ligand based binding assay (sR/bL binding assay) that assessed efficacy to block binding of soluble IC receptor on its cognate ligand immobilized beads. Half maximal inhibitory concentration (IC50) values of ICIs were calculated to determine ICIs activity. The sL/bR binding assay accurately determined the activity of two TIGIT blocking antibodies, since the relative blocking activity of two TIGIT antibodies determined by the sL/bR binding assay established in this study and that by the cell based binding assay were almost identical. In contrast, the sR/bL binding assay showed significantly improved sensitivity to determine activity of two PD-1 blocking antibodies than the sL/bR binding assay that was tested in this study and previous reports. Moreover, both amount of the used recombinant protein of ICI receptor/ligand and turnaround time of the two binding assays were more than 10 times less than those of the functional ELISA. These data indicate that the two magnetic bead based binding assays are sensitive, rapid and cost-effective methods to determine blocking activity of ICIs.

Human mammalian cell sorting using a highly integrated micro-fabricated fluorescence-activated cell sorter (microFACS).

We demonstrate a high performance microfabricated FACS system with highly integrated microfluidics, optics, acoustics, and electronics. Single cell manipulation at a high speed is made possible by the fast response time (approximately 0.1 ms) of the integrated PZT actuator and the nozzle structure at the sorting junction. A Teflon AF-coated optofluidic waveguide along the microfluidic channel guides the illumination light, enabling multi-spot detection, while a novel space-time coding technology enhances the detection sensitivity of the microFACS system. The real-time control loop system is implemented using a field-programmable-gate-array (FPGA) for automated and accurate sorting. The microFACS achieves a high purification enrichment factor: up to approximately 230 fold for both polystyrene microbeads and suspended human mammalian cells (K562) at a high throughput (>1000 cells s(-1)). The sorting mechanism is independent of cell properties such as size, density, and shape, thus the presented system can be applied to sort out any pure sub-populations. This new lab-on-a-chip FACS system, therefore, holds promise to revolutionize microfluidic cytometers to meet cost, size, and performance goals.