Microfluid biosensor for detection of hpv in patient scraping samples: Determining E6 and E7 oncogenes.

E6 and E7 oncogenes are pivotal in the carcinogenic transformation in HPV infections and efficient diagnostic methods can ensure the detection and differentiation of HPV genotype. This study describes the development and validation of an electrochemical, label-free genosensor coupled with a microfluidic system for detecting the E6 and E7 oncogenes in cervical scraping samples. The nanostructuring employed was based on a cysteine and graphene quantum dots layer that provides functional groups, surface area, and interesting electrochemical properties. Biorecognition tests with cervical scraping samples showed differentiation in the voltammetric response. Low-risk HPV exhibited a lower biorecognition response, reflected in ΔI% values of 82.33 % ± 0.29 for HPV06 and 80.65 % ± 0.68 for HPV11 at a dilution of 1:100. Meanwhile, high-risk, HPV16 and HPV18, demonstrated ΔI% values of 96.65 % ± 1.27 and 93 % ± 0.026, respectively, at the same dilution. Therefore, the biorecognition intensity followed the order: HPV16 >HPV18 >HPV06 >HPV11. The limit of detection and the limit of quantification of E6E7 microfluidic LOC-Genosensor was 26 fM, and 79.6 fM. Consequently, the E6E7 biosensor is a valuable alternative for clinical HPV diagnosis, capable of detecting the potential for oncogenic progression even in the early stages of infection.

Streamlined integrated protein isoelectric focusing using microfluidic paper-based device.

An innovative integrated paper-based microdevice was developed for protein separation by isoelectric focusing (IEF), allowing for robust design thanks to a 3D-printed holder integrating separation channel, reservoirs, and electrodes. To reach robustness and precision, the optimization focused on the holder geometry, the paper nature, the reservoir design, the IEF medium, and various focusing parameters. A well-established and stable pH gradient was obtained on a glass-fiber paper substrate with simple sponge reservoirs, and the integration of the electrodes in the holder led to a straightforward system. The separation medium composed of water/glycerol (85/15, v/v) allowed for reducing medium evaporation while being an efficient medium for most hydrophobic and hydrophilic proteins, compatible with mass spectrometry detection for further proteomics developments. To our knowledge, this is the first report of the use of glycerol solutions as a separation medium in a paper-based microdevice. Analytical performances regarding pH gradient generation, pI determination, separation efficiency, and resolution were estimated while varying the IEF experimental parameters. The overall process led to an efficient separation within 25 min. Then, this methodology was applied to a sample composed of saliva doped with proteins. A minimal matrix effect was evidenced, underscoring the practical viability of our platform. This low-cost, versatile and robust paper-based IEF microdevice opens the way to various applications, ranging from sample pre-treatment to integration in an overall proteomic-on-a-chip device.

A 3D-printed tumor-on-chip: user-friendly platform for the culture of breast cancer spheroids and the evaluation of anti-cancer drugs.

Tumor-on-chips (ToCs) are useful platforms for studying the physiology of tumors and evaluating the efficacy and toxicity of anti-cancer drugs. However, the design and fabrication of a ToC system is not a trivial venture. We introduce a user-friendly, flexible, 3D-printed microfluidic device that can be used to culture cancer cells or cancer-derived spheroids embedded in hydrogels under well-controlled environments. The system consists of two lateral flow compartments (left and right sides), each with two inlets and two outlets to deliver cell culture media as continuous liquid streams. The central compartment was designed to host a hydrogel in which cells and microtissues can be confined and cultured. We performed tracer experiments with colored inks and 40 kDa fluorescein isothiocyanate dextran to characterize the transport/mixing performances of the system. We also cultured homotypic (MCF7) and heterotypic (MCF7-BJ) spheroids embedded in gelatin methacryloyl hydrogels to illustrate the use of this microfluidic device in sustaining long-term micro-tissue culture experiments. We further demonstrated the use of this platform in anticancer drug testing by continuous perfusion of doxorubicin, a commonly used anti-cancer drug for breast cancer. In these experiments, we evaluated drug transport, viability, glucose consumption, cell death (apoptosis), and cytotoxicity. In summary, we introduce a robust and friendly ToC system capable of recapitulating relevant aspects of the tumor microenvironment for the study of cancer physiology, anti-cancer drug transport, efficacy, and safety. We anticipate that this flexible 3D-printed microfluidic device may facilitate cancer research and the development and screening of strategies for personalized medicine.

Unveiling the intricacies of BPA and BPS: comprehensive insights into its toxic effects using a cutting-edge microphysiological system.

Concerns over Bisphenol A (BPA) and its substitute, Bisphenol S (BPS), have led to innovative exploration due to potential adverse health effects. BPS, replacing BPA in some regions to avoid toxic impacts, remains insufficiently studied. Besides this, the organ-on-a-chip technology emerges as a transformative solution in drug discovery and chemiclas toxicity testing, minimizing costs and aligning with ethical standards by reducing reliance on animal models, by integrating diverse tissues and dynamic cell environments enhances precision in predicting organ function. Here, we employ a 3-organ-on-a-chip microfluidic device with skin, intestine, and liver cultures to assess the effects of BPA and BPS via topical and oral administration. Our evaluation focused on gene markers associated with carcinogenicity, systemic toxicity, and endocrine disruption. BPA exhibited expected absorption profiles, causing liver injury and genetic modulation in related pathways. BPS, a safer alternative, induced adverse effects on gene expression, particularly in topical absorption, with distinct absorption patterns. Our findings underscore the urgency of addressing BPA and BPS toxicity concerns, highlighting the crucial role of organ-on-a-chip technology in understanding associated health risks. The study promotes the organ-on-a-chip methodology as a valuable tool for safe drug development and disease treatments, offering a novel liver toxicity screening alternative to traditional animal tests. This contributes to advancing comprehension of the biological effects of these compounds, fostering improved safety assessments in human health.

Noninvasive and Multiplex Self-Test of Kidney Disease Biomarkers with Graphene-Based Lab-on-a-Chip (G-LOC): Toward Digital Diagnostics in the Hands of Patients.

Chronic kidney disease is one of the major health issues worldwide. However, diagnosis is now highly centralized in large laboratories, resulting in low access to patient monitoring and poor personalized treatments. This work reports the development of a graphene-based lab-on-a-chip (G-LOC) for the digital testing of renal function biomarkers in serum and saliva samples. G-LOC integrates multiple bioelectronic sensors with a microfluidic system that enables multiplex self-testing of urea, potassium, sodium, and chloride. The linearity, limit of detection (LOD), accuracy, and coefficient of variability (CV) were studied. Accuracy values higher than 95.5% and CV lower than 9% were obtained for all of the biomarkers. The analytical performance was compared against three reference lab benchtop analyzers by measuring healthy- and renal-failure-level samples of serum. From receiver operating characteristic (ROC) plots, sensitivities (%) of 99.7, 97.6, 99.1, and 89.0 were obtained for urea, potassium, sodium, and chloride, respectively. Then, the test was evaluated in noninvasive saliva samples and compared against reference methods. Correlation and Bland-Altman plots showed good correlation and agreement of the G-LOC with the reference methods. It is noteworthy that the precision of G-LOC was similar to better than benchtop lab analyzers, with the advantage of being highly portable. Finally, a user testing study was conducted. The analytical performance obtained with untrained volunteers was similar to that obtained with trained chemists. Additionally, based on a user experience survey, G-LOC was found to have very simple usability and would be suitable for at-home diagnostics.

Portable platform for leukocyte extraction from blood using sheath-free microfluidic DLD.

Leukocyte count is routinely performed for diagnostic purposes and is rapidly emerging as a significant biomarker for a wide array of diseases. Additionally, leukocytes have demonstrated considerable promise in novel cell-based immunotherapies. However, the direct retrieval of leukocytes from whole blood is a significant challenge due to their low abundance compared to erythrocytes. Here, we introduce a microfluidic-based platform that isolates and recovers leukocytes from diluted whole blood in a single step. Our platform utilizes a novel, sheathless method to initially sediment and focus blood cells into a dense stream while flowing through a tubing before entering the microfluidic device. A hexagonal-shaped structure, patterned at the device's inlet, directs all the blood cells against the channel's outer walls. The focused cells are then separated based on their size using the deterministic lateral displacement (DLD) microfluidic technique. We evaluated various parameters that could influence leukocyte separation, including different focusing structures (assessed both computationally and experimentally), the orientation of the tubing-chip interface, the effects of blood sample hematocrit (dilution), and flow rate. Our device demonstrated the ability to isolate leukocytes from diluted blood with a separation efficiency of 100%, a recovery rate of 76%, and a purity of 80%, while maintaining a cell viability of 98%. The device operates for over 30 min at a flow rate of 2 µL min-1. Furthermore, we developed a handheld pressure controller to drive fluid flow, enhancing the operability of our platform outside of central laboratories and enabling near-patient testing. Our platform can be integrated with downstream cell-based assays and analytical methods that require high leukocyte purity (80%), ranging from cell counting to diagnostics and cell culture applications.

A review of the recent achievements and future trends on 3D printed microfluidic devices for bioanalytical applications.

3D printing has revolutionized the manufacturing process of microanalytical devices by enabling the automated production of customized objects. This technology promises to become a fundamental tool, accelerating investigations in critical areas of health, food, and environmental sciences. This microfabrication technology can be easily disseminated among users to produce further and provide analytical data to an interconnected network towards the Internet of Things, as 3D printers enable automated, reproducible, low-cost, and easy fabrication of microanalytical devices in a single step. New functional materials are being investigated for one-step fabrication of highly complex 3D printed parts using photocurable resins. However, they are not yet widely used to fabricate microfluidic devices. This is likely the critical step towards easy and automated fabrication of sophisticated, complex, and functional 3D-printed microchips. Accordingly, this review covers recent advances in the development of 3D-printed microfluidic devices for point-of-care (POC) or bioanalytical applications such as nucleic acid amplification assays, immunoassays, cell and biomarker analysis and organs-on-a-chip. Finally, we discuss the future implications of this technology and highlight the challenges in researching and developing appropriate materials and manufacturing techniques to enable the production of 3D-printed microfluidic analytical devices in a single step.

Renal proximal tubule-on-a-chip in PDMS: fabrication, functionalization, and RPTEC:HUVEC co-culture evaluation.

'On-a-chip' technology advances the development of physiologically relevant organ-mimicking architecture by integrating human cells into three-dimensional microfluidic devices. This method also establishes discrete functional units, faciliting focused research on specific organ components. In this study, we detail the development and assessment of a convoluted renal proximal tubule-on-a-chip (PT-on-a-chip). This platform involves co-culturing Renal Proximal Tubule Epithelial Cells (RPTEC) and Human Umbilical Vein Endothelial Cells (HUVEC) within a polydimethylsiloxane microfluidic device, crafted through a combination of 3D printing and molding techniques. Our PT-on-a-chip significantly reduced high glucose level, exhibited albumin uptake, and simulated tubulopathy induced by amphotericin B. Remarkably, the RPTEC:HUVEC co-culture exhibited efficient cell adhesion within 30 min on microchannels functionalized with plasma, 3-aminopropyltriethoxysilane, and type-I collagen. This approach significantly reduced the required incubation time for medium perfusion. In comparison, alternative methods such as plasma and plasma plus polyvinyl alcohol were only effective in promoting cell attachment to flat surfaces. The PT-on-a-chip holds great promise as a valuable tool for assessing the nephrotoxic potential of new drug candidates, enhancing our understanding of drug interactions with co-cultured renal cells, and reducing the need for animal experimentation, promoting the safe and ethical development of new pharmaceuticals.

Trends and challenges in microfluidic methods for protein manipulation-A review.

Proteins are important molecules involved in an immensely large number of biological processes. Being capable of manipulating proteins is critical for developing reliable and affordable techniques to analyze and/or detect them. Such techniques would enable the production of therapeutic agents for the treatment of diseases or other biotechnological applications (e.g., bioreactors or biocatalysis). Microfluidic technology represents a potential solution to protein manipulation challenges because of the diverse phenomena that can be exploited to achieve micro- and nanoparticle manipulation. In this review, we discuss recent contributions made in the field of protein manipulation in microfluidic systems using different physicochemical principles and techniques, some of which are miniaturized versions of already established macro-scale techniques.

Applications of Lab on a Chip in Antimicrobial Susceptibility of Staphylococcus aureus: A Systematic Review.

Background and Objectives: Staphylococcus aureus is a prevalent bacterium capable of inducing various infections, including skin and soft tissue infections, bloodstream infections, pneumonia, and surgical site infections. The emergence of antimicrobial resistance in S. aureus, particularly methicillin-resistant S. aureus, has raised substantial concerns within global healthcare settings. Prior to antibiotic prescription, the ideal approach is antimicrobial susceptibility testing (AST); however, this is frequently perceived as excessively complex and time-intensive. Lab-on-a-chip (LOC) technology holds promise in addressing these challenges and advancing fundamental microbiological research while also aiding in the development of therapeutic strategies. This systematic review aims to evaluate the potential utility of LOC for AST of S. aureus. Materials and Methods: This study adhered to the PRISMA guidelines. Various databases, including SCOPUS, PubMed/MEDLINE, SCIELO, and LILACS, in addition to gray literature sources, were employed in the review process. Results: Sixteen studies were included in this systematic review. All these studies detailed the effectiveness, rapidity, and predictability of LOC systems for assessing S. aureus susceptibility to various antibiotics. When comparing the LOC approach to traditional manual methods, it was evident that LOC requires a minimal quantity of reagents. Furthermore, most studies reported that the entire LOC procedure took 10 min to 7 h, with results being equally accurate as those obtained through traditional AST protocols. Conclusions: The potential application of LOC for AST of S. aureus is emphasized by its ability to provide rapid access to minimum inhibitory concentration data, which can substantially aid in selecting the most suitable antibiotics and dosages for treating challenging infections caused by this microorganism. Moreover, the rapid AST facilitated by LOC holds promise for enhancing the appropriateness and efficacy of therapy in clinical settings.

Simultaneous detection of dopamine and ascorbic acid by using a thread-based microfluidic device and multiple pulse amperometry.

This study presents a novel approach for the simultaneous detection of ascorbic acid (AA) and dopamine (DA) using an affordable and user-friendly microfluidic device. Microfluidic devices, when combined with electrochemical detectors like screen-printed electrodes (SPEs), offer numerous advantages such as portability, high sample throughput, and low reagent consumption. In this study, a 3D-printed microfluidic device called a µTED was developed, utilizing textile threads as microfluidic channels and an unmodified SPE as the amperometric detector. The method employed multiple pulse amperometry (MPA) with carefully selected potential values (+0.65 V and -0.10 V). The reduction current signals generated by dopamine o-quinone were used to calculate a correction factor for the oxidation signals of ascorbic acid, enabling simultaneous quantification. The developed microfluidic device ensured a stable flow rate of the carrier solution at 1.19 µL s-1, minimizing the consumption of samples and reagents (injection volume of 2.0 µL). Under the optimized experimental conditions, a linear range from 50 to 900 µmol L-1 was achieved for both DA and AA. The obtained sensitivities were 2.24 µA L mmol-1 for AA and 5.09 µA L mmol-1 for DA, with corresponding limits of detection (LOD) of 2.60 µmol L-1 and 1.54 µmol L-1, respectively. To confirm the effectiveness of the proposed method, it was successfully applied to analyze AA and DA in a commercial blood serum sample spiked at three different concentration levels, with a medium recovery rate of 70%. Furthermore, the MPA technique demonstrated its simplicity by enabling the simultaneous determination of AA and DA without the need for prior separation steps or the use of chemically modified electrodes.

An electrochemical microfluidic device for non-enzymatic cholesterol determination using a lab-made disposable electrode.

Cholesterol is an important steroid and hormone precursor, and its levels in the blood are associated with risk factors for cardiovascular diseases. In this work, a non-enzymatic methodology for cholesterol determination in serum samples is described. First, a working electrode was constructed using homemade ink and a plastic substrate by a simple dunking process. Next, the dunked electrode (DWE) was modified with nickel ions (Ni-DWE) and combined with a low-cost microfluidic platform, resulting in a thread-based electroanalytical device (µTED). The arrangement of µTED consists of two coupled electrodes (one reference in the inlet reservoir and an auxiliary electrode against the outlet reservoir) and a mobile support for facile working electrode exchange. After optimization of construction parameters, the system was applied for non-enzymatic determination of cholesterol under alkaline conditions using the redox pair Ni(II)/Ni(III) as a mediator. Under the best analytical conditions, a calibration curve was constructed with a linear dynamic range (LDR) from 0.25 to 25.0 µmol L-1, and the calculated limits of detection (LOD) and quantification (LOQ) were 0.074 and 0.24 µmol L-1, respectively. No effects of possible interferents on electrochemical response were found in the presence of ascorbic acid, uric acid, dopamine, cysteine, and glucose, suggesting that the proposed device can be used for the determination of cholesterol without significant matrix effects of human plasma. Finally, cholesterol analysis was carried out using spiked plasma samples, and good recovery values were achieved.

Paper-based electrochemical biosensors for the diagnosis of viral diseases.

Paper-based electrochemical analytical devices (ePADs) have gained significant interest as promising analytical units in recent years because they can be fabricated in simple ways, are low-cost, portable, and disposable platforms that can be applied in various fields. In this sense, paper-based electrochemical biosensors are attractive analytical devices since they can promote diagnose several diseases and potentially allow decentralized analysis. Electrochemical biosensors are versatile, as the measured signal can be improved by using mainly molecular technologies and nanomaterials to attach biomolecules, resulting in an increase in their sensitivity and selectivity. Additionally, they can be implemented in microfluidic devices that drive and control the flow without external pumping and store reagents, and improve the mass transport of analytes, increasing sensor sensitivity. In this review, we focus on the recent developments in electrochemical paper-based devices for viruses' detection, including COVID-19, Dengue, Zika, Hepatitis, Ebola, AIDS, and Influenza, among others, which have caused impacts on people's health, especially in places with scarce resources. Also, we discuss the advantages and disadvantages of the main electrode's fabrication methods, device designs, and biomolecule immobilization strategies. Finally, the perspectives and challenges that need to be overcome to further advance paper-based electrochemical biosensors' applications are critically presented.

Secretion Function Analysis of Ex Vivo Immune Cells in an Integrated Microfluidic Device.

Immune cells play a major role in the development of cancer, from being able to inhibit it by secreting pro-inflammatory mediators, to assist in its development by secreting growth factors, immunosuppressive mediators, and ECM-modifying enzymes. Therefore, the ex vivo analysis of the secretion function of immune cells can be employed as a reliable prognostic biomarker in cancer. However, one limiting factor in current approaches to probe the ex vivo secretion function of cells is their low throughput and the consumption of large quantities of sample. Microfluidics provides a unique advantage, by being able to integrate different components, such as cell culture and biosensors in a monolithic microdevice; it can increase the analytical throughput and leverage it with its intrinsic low sample requirement. Furthermore, the integration of fluid control elements also allows this analysis to be highly automatable, leading to increases in consistency in the results. Here, we describe an approach to analyze the ex vivo secretion function of immune cells using a highly integrated microfluidic device.

Optical Detection of Cancer Cells Using Lab-on-a-Chip.

The global need for accurate and efficient cancer cell detection in biomedicine and clinical diagnosis has driven extensive research and technological development in the field. Precision, high-throughput, non-invasive separation, detection, and classification of individual cells are critical requirements for successful technology. Lab-on-a-chip devices offer enormous potential for solving biological and medical problems and have become a priority research area for microanalysis and manipulating cells. This paper reviews recent developments in the detection of cancer cells using the microfluidics-based lab-on-a-chip method, focusing on describing and explaining techniques that use optical phenomena and a plethora of probes for sensing, amplification, and immobilization. The paper describes how optics are applied in each experimental method, highlighting their advantages and disadvantages. The discussion includes a summary of current challenges and prospects for cancer diagnosis.

Organ-on-a-Chip for Drug Screening: A Bright Future for Sustainability? A Critical Review.

Globalization has raised concerns about spreading diseases and emphasized the need for quick and efficient methods for drug screening. Established drug efficacy and toxicity approaches have proven obsolete, with a high failure rate in clinical trials. Organ-on-a-chip has emerged as an essential alternative to outdated techniques, precisely simulating important characteristics of organs and predicting drug pharmacokinetics more ethically and efficiently. Although promising, most organ-on-a-chip devices are still manufactured using principles and materials from the micromachining industry. The abusive use of plastic for traditional drug screening methods and device production should be considered when substituting technologies so that the compensation for the generation of plastic waste can be projected. This critical review outlines recent advances for organ-on-a-chip in the industry and estimates the possibility of scaling up its production. Moreover, it analyzes trends in organ-on-a-chip publications and provides suggestions for a more sustainable future for organ-on-a-chip research and production.

Selection of phage-displayed antibodies with high affinity and specificity by electrophoresis in microfluidic devices.

A method development aimed for high-throughput and automated antibody screening holds great potential for areas ranging from fundamental molecular interactions to the discovery of novel disease markers, therapeutic targets, and monoclonal antibody engineering. Surface display techniques enable efficient manipulation of large molecular libraries in small volumes. Specifically, phage display appeared as a powerful technology for selecting peptides and proteins with enhanced, target-specific binding affinities. Here, we present a phage-selection microfluidic device wherein electrophoresis was performed under two orthogonal electric fields through an agarose gel functionalized with the respective antigen. This microdevice was capable of screening and sorting in a single round high-affinity phage-displayed antibodies against virus glycoproteins, including human immunodeficiency virus-1 glycoprotein 120 or the Ebola virus glycoprotein (EBOV-GP). Phages were differentially and laterally swept depending on their antigen affinity; the high-affinity phages were recovered at channels proximal to the application site, whereas low-affinity phages migrated distal after electrophoresis. These experiments proved that the microfluidic device specifically designed for phage-selection is rapid, sensitive, and effective. Therefore, this is an efficient and cost-effective method that allowed highly controlled assay conditions for isolating and sorting high-affinity ligands displayed in phages.

"Do it yourself" protocol to fabricate dual-detection paper-based analytical device for salivary biomarker analysis.

This paper describes the design and construction of dual microfluidic paper-based analytical devices (dual-µPADs) as a lab-on-paper platform involving a "do-it-yourself" fabrication protocol. The device comprises a colorimetric and electrochemical module to obtain a dual-mode signal readout sensing strategy. A 3D pen polymeric resin was used to prepare graphite carbon-based electrodes and hydrophobic barriers on paper substrates. The proposed carbon-based ink was employed to manufacture electrodes on paper based on a stencil-printing approach, which were further characterized by electrochemical and morphological analyses. The analytical performance of the dual-µPADs was simultaneously evaluated for lactate, pH, nitrite, and salivary amylase (sAA) analysis. To demonstrate the proof-of-concept, saliva samples collected from both healthy individuals and those with periodontitis were successfully tested to demonstrate the feasibility of the proposed devices. Samples collected from individuals previously diagnosed with periodontitis showed high levels of nitrite and sAA (> 94 µmol L-1 and > 610 U mL-1) in comparison with healthy individuals (≤ 16 µmol L-1 and 545 U mL-1). Moreover, periodontitis saliva resulted in acid solution and almost null lactate levels. Notably, this protocol supplies a simple way to manufacture dual-µPADs, a versatile platform for sensitive detecting of biomarkers in saliva playing a crucial role towards the point-of-care diagnosis of periodontal disease.

Recent advances and challenges in temperature monitoring and control in microfluidic devices.

Temperature is a critical-yet sometimes overlooked-parameter in microfluidics. Microfluidic devices can experience heating inside their channels during operation due to underlying physicochemical phenomena occurring therein. Such heating, whether required or not, must be monitored to ensure adequate device operation. Therefore, different techniques have been developed to measure and control temperature in microfluidic devices. In this contribution, the operating principles and applications of these techniques are reviewed. Temperature-monitoring instruments revised herein include thermocouples, thermistors, and custom-built temperature sensors. Of these, thermocouples exhibit the widest operating range; thermistors feature the highest accuracy; and custom-built temperature sensors demonstrate the best transduction. On the other hand, temperature control methods can be classified as external- or integrated-methods. Within the external methods, microheaters are shown to be the most adequate when working with biological samples, whereas Peltier elements are most useful in applications that require the development of temperature gradients. In contrast, integrated methods are based on chemical and physical properties, structural arrangements, which are characterized by their low fabrication cost and a wide range of applications. The potential integration of these platforms with the Internet of Things technology is discussed as a potential new trend in the field.

Advances in Concentration Gradient Generation Approaches in a Microfluidic Device for Toxicity Analysis.

This systematic review aimed to analyze the development and functionality of microfluidic concentration gradient generators (CGGs) for toxicological evaluation of different biological organisms. We searched articles using the keywords: concentration gradient generator, toxicity, and microfluidic device. Only 33 of the 352 articles found were included and examined regarding the fabrication of the microdevices, the characteristics of the CGG, the biological model, and the desired results. The main fabrication method was soft lithography, using polydimethylsiloxane (PDMS) material (91%) and SU-8 as the mold (58.3%). New technologies were applied to minimize shear and bubble problems, reduce costs, and accelerate prototyping. The Christmas tree CGG design and its variations were the most reported in the studies, as well as the convective method of generation (61%). Biological models included bacteria and nematodes for antibiotic screening, microalgae for pollutant toxicity, tumor and normal cells for, primarily, chemotherapy screening, and Zebrafish embryos for drug and metal developmental toxicity. The toxic effects of each concentration generated were evaluated mostly with imaging and microscopy techniques. This study showed an advantage of CGGs over other techniques and their applicability for several biological models. Even with soft lithography, PDMS, and Christmas tree being more popular in their respective categories, current studies aim to apply new technologies and intricate architectures to improve testing effectiveness and reduce common microfluidics problems, allowing for high applicability of toxicity tests in different medical and environmental models.