Periodontitis and dental caries are two major bacterially induced, non-communicable diseases that cause the deterioration of oral health, with implications in patients’ general health. Early, precise diagnosis and personalized monitoring are essential for the efficient prevention and management of these diseases. Here, we present a disk-shaped microfluidic platform (OralDisk) compatible with chair-side use that enables analysis of non-invasively collected whole saliva samples and molecular-based detection of ten bacteria: seven periodontitis-associated (Aggregatibacter actinomycetemcomitans, Campylobacter rectus, Fusobacterium nucleatum, Prevotella intermedia, Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola) and three caries-associated (oral Lactobacilli, Streptococcus mutans, Streptococcus sobrinus). Each OralDisk test required 400 µL of homogenized whole saliva. The automated workflow included bacterial DNA extraction, purification and hydrolysis probe real-time PCR detection of the target pathogens. All reagents were pre-stored within the disk and sample-to-answer processing took < 3 h using a compact, customized processing device. A technical feasibility study (25 OralDisks) was conducted using samples from healthy, periodontitis and caries patients. The comparison of the OralDisk with a lab-based reference method revealed a ~90% agreement amongst targets detected as positive and negative. This shows the OralDisk’s potential and suitability for inclusion in larger prospective implementation studies in dental care settings.

Centrifugal microfluidics enables fully automated molecular diagnostics at the point-of-need. However, the integration of solid-phase nucleic acid extraction remains a challenge. Under this scope, we developed the magnetophoresis under continuous rotation for magnetic bead-based nucleic acid extraction. Four stationary permanent magnets are arranged above a cartridge, creating a magnetic field that enables the beads to be transported between the chambers of the extraction module under continuous rotation. The centrifugal force is maintained to avoid uncontrolled spreading of liquids. We concluded that below a frequency of 5 Hz, magnetic beads move radially inwards. In support of magnetophoresis, bead inertia and passive geometrical design features allow to control the azimuthal bead movement between chambers. We then demonstrated ferrimagnetic bead transfer in liquids with broad range of surface tension and density values. Furthermore, we extracted nucleic acids from lysed Anopheles gambiae mosquitoes reaching comparable results of eluate purity (LabDisk: A260/A280 = 1.6 ± 0.04; Reference: 1.8 ± 0.17), and RT-PCR of extracted RNA (LabDisk: Ct = 17.9 ± 1.6; Reference: Ct = 19.3 ± 1.7). Conclusively, magnetophoresis at continuous rotation enables easy cartridge integration and nucleic acid extraction at the point-of-need with high yield and purity.

Effective mosquito monitoring relies on the accurate identification and characterization of the target population. Since this process requires specialist knowledge and equipment that is not widely available, automated field-deployable systems are highly desirable. We present a centrifugal microfluidic cartridge, the VectorDisk, which integrates TaqMan PCR assays in two feasibility studies, aiming to assess multiplexing capability, specificity, and reproducibility in detecting disk-integrated vector-related assays. In the first study, pools of 10 mosquitoes were used as samples. We tested 18 disks with 27 DNA and RNA assays each, using a combination of multiple microfluidic chambers and detection wavelengths (geometric and color multiplexing) to identify mosquito and malaria parasite species as well as insecticide resistance mechanisms. In the second study, purified nucleic acids served as samples to test arboviral and malaria infective mosquito assays. Nine disks were tested with 14 assays each. No false positive results were detected on any of the disks. The coefficient of variation in reproducibility tests was <10%. The modular nature of the platform, the easy adaptation of the primer/probe panels, the cold chain independence, the rapid (2–3 h) analysis, and the assay multiplexing capacity are key features, rendering the VectorDisk a potential candidate for automated vector analysis.

Background: The growing demand for versatile laboratory automation is exemplified in the context of liquid biopsy, where multi-analyte approaches are increasingly recognised for their potential to enhance diagnostic sensitivity in oncology. However, current practice often necessitates the use of dedicated instruments and workflows for the extraction of each analyte, posing financial and logistical barriers for automated multi-analyte liquid biopsy. Methods: Here, we present Robotic Centrifugal Microfluidics (RoCM), an all-in-one platform that combines the versatility of centrifugal microfluidics and operational flexibility of robotic liquid handling. This combination enables the automation of complex micro- and macrofluidic protocols, realised through the use of (1). exchangeable microfluidic cartridges and (2). programmable robotic operations such as in-rotation liquid supply, magnetic bead manipulation, or microfluidic valving. In-rotation robotic liquid manipulation maintains fluid control under centrifugal forces and reduces the cartridge footprint associated with pre-loaded liquid reservoirs. Platform applicability was demonstrated using two exemplary liquid biopsy workflows: the extraction of cell-free DNA (cfDNA) from blood plasma using RoCM-cfDNA slices and the extraction of extracellular vesicles (EVs) from blood plasma using RoCM-EV slices. Results: In a pilot study with patient samples from different cancer entities, the RoCM-cfDNA slices yielded comparable variant allele frequencies to a commercial bead-based instrument, while the RoCM-EV slices achieved a recovery of a greater diversity of EV subpopulations than semi-automated size-exclusion chromatography. Conclusions: By simply exchanging cartridges, RoCM enables the extraction of diverse analytes within a single automated system. Its application can be extended to further analytes, such as circulating tumour cells (CTCs), or to applications beyond liquid biopsies, where versatile micro- and macrofluidic protocols benefit from implementation in a single automation instrument.

Tuberculosis (TB) is still one of the world’s deadliest infections. Fast detection of the pathogen M. tuberculosis (MTB) and its genetic resistance markers substantially improves treatment success and outcome. A key element for rapid genetic diagnostics is the efficient extraction of DNA from sputum for qPCR detection at the point of care. We present the fully automated sample preparation of MTB DNA from 3 mL of liquefied sputum and qPCR detection of MTB on a centrifugal microfluidic cartridge. Our method achieves a limit of detection (LoD) between 17 and 57 CFU/mL and provides a purified DNA solution for molecular downstream testing, such as targeted NGS.

Food allergies pose a serious and increasing public health challenge as many allergens can trigger potential life-threatening reactions in affected individuals. Reliable detection of allergens such as soy proteins is therefore a critical and essential tool to detect cross-contaminations, particularly in food processing and production environments. To enable monitoring directly at these sites, portable biosensing platforms are required to provide rapid point-of-need detection. However, the development of such systems requires the combination of biochemistry, material science, and precision engineering. In this article we present a centrifugal microfluidic platform integrating automated sample preparation, clean-up, and immunological detection in a single-use disposable cartridge. Targeting soybean protein as a model allergen, the system combines optimized immunoassay chemistry with advanced polymer microfabrication. Regioselective antibody coupling strategies were investigated for stable and oriented bead conjugation. N-glycan click chemistry was used to produce monodisperse, highly functional fluorescent microbeads. The microfluidic LabDisk was first manufactured by thermoforming and later transferred to injection molding with integrated optical elements to increase sensitivity. The fully integrated LabDisk detected soy protein at the lowest concentrations tested, namely 1 ppm in extraction buffer and 4 ppm in incurred complex matrices. This work shows the important steps from biochemical concept to scalable manufacturing, paving the way for mass-produced, portable allergen biosensors.