Advection-enhanced kinetics in microtiter plates for improved surface assay quantitation and multiplexing capabilities
Surface assays such as ELISA are pervasive in clinics and research and predominantly standardized in microtiter plates (MTP). MTPs provide many advantages but are often detrimental to surface assay efficiency due to inherent mass transport limitations. Microscale flows can overcome these and largely improve assay kinetics. However, the disruptive nature of microfluidics with existing labware and protocols has narrowed its transformative potential. We present WellProbe, a novel microfluidic concept compatible with MTPs. With it, we show that immunoassays become more sensitive at low concentrations (up to 9× signal improvement in 12x less time), richer in information with 3-4 different kinetic conditions, and can be used to estimate kinetic parameters, minimize washing steps and non-specific binding, and identify compromised results. We further multiplex single-well assays combining WellProbe’s kinetic regions with tailored microarrays. Finally, we demonstrate our system in a context of immunoglobulin subclass evaluation, increasingly regarded as clinically relevant.Read More
Automation for Life Science LaboratoriesThe automation of processes in all areas of the life sciences will continue to increase in the coming years due to an ever increasing number of samples to be processed Gentaur Labware, an increasing need to protect laboratory personnel from infectious material and increasing cost pressure. Depending on the requirements of the respective application, different concepts for automation systems are available, which have a different degree of automation with regard to data handling, transportation tasks, and the processing of the samples.
- Robots form a central component of these automation concepts. Classic stationary robots from the industrial sector will increasingly be replaced by new developments in the field of light-weight robots.
- In addition, mobile robots will also be of particular importance in the automation of life science laboratories in the future, especially for transportation tasks between different manual and (partially) automated stations.
- With an increasing number of different, highly diverse processes, the need for special devices and system components will also increase.
- This applies to both, the handling of the labware and the processing of the samples. In contrast to previous automation strategies with a highly parallel approach, future developments will increasingly be characterized by individual sample handling.
Fluorescence-based Single-cell Analysis of Whole-mount-stained and Cleared Microtissues and Organoids for High Throughput ScreeningThree-dimensional (3D) cell culture, especially in the form of organ-like microtissues (“organoids”), has emerged as a novel tool potentially mimicking human tissue biology more closely than standard two-dimensional culture. Typically, tissue sectioning is the standard method for immunohistochemical analysis. However, it removes cells from their native niche and can result in the loss of 3D context during analyses.
- Automated workflows require parallel processing and analysis of hundreds to thousands of samples, and sectioning is mechanically complex, time-intensive, and thus less suited for automated workflows.
- Here, we present a simple protocol for combined whole-mount immunostaining, tissue-clearing, and optical analysis of large-scale (approx. 1 mm) 3D tissues with single-cell level resolution.
- While the protocol can be performed manually, it was specifically designed to be compatible with high-throughput applications and automated liquid handling systems.
- This approach is freely scalable and allows parallel automated processing of large sample numbers in standard labware.
- We have successfully applied the protocol to human mid- and forebrain organoids, but, in principle, the workflow is suitable for a variety of 3D tissue samples to facilitate the phenotypic discovery of cellular behaviors in 3D cell culture-based high-throughput screens.
- Graphic abstract: Automatable organoid clearing and high-content analysis workflow and timeline.
Gold-Polyoxoborates Nanocomposite Prohibits Adsorption of Bacteriophages on Inner Surfaces of Polypropylene Labware and Protects Samples from Bacterial and Yeast InfectionsBacteriophages (phages) are a specific type of viruses that infect bacteria. Because of growing antibiotic resistance among bacterial strains, phage-based therapies are becoming more and more attractive. The critical problem is the storage of bacteriophages. Recently, it was found that bacteriophages might adsorb on the surfaces of plastic containers, effectively decreasing the titer of phage suspensions. Here, we showed that a BOA nanocomposite (gold nanoparticles embedded in polyoxoborate matrix) deposited onto the inner walls of the containers stabilizes phage suspensions against uncontrolled adsorption and titer decrease. Additionally, BOA provides antibacterial and antifungal protection. The application of BOA assures safe and sterile means for the storage of bacteriophages.
Adsorption of bacteriophages on polypropylene labware affects the reproducibility of phage researchHydrophobicity is one of the most critical factors governing the adsorption of molecules and objects, such as virions, on surfaces. Even moderate change of wetting angle of plastic surfaces causes a drastic decrease ranging from 2 to 5 logs of the viruses (e.g., T4 phage) in the suspension due to adsorption on polymer vials’ walls.
- The effect varies immensely in seemingly identical containers but purchased from different vendors. Comparison of glass, polyethylene, polypropylene, and polystyrene containers revealed a threshold in the wetting angle of around 95°: virions adsorb on the surface of more hydrophobic containers, while in more hydrophilic vials, phage suspensions are stable.
- The polypropylene surface of the Eppendorf-type and Falcon-type can accommodate from around 108 PFU/ml to around 1010 PFU/ml from the suspension.
- The adsorption onto the container’s wall might result in complete scavenging of virions from the bulk. We developed two methods to overcome this issue.
- The addition of surfactant Tween20 and/or plasma treatment provides a remedy by modulating surface wettability and inhibiting virions’ adsorption.
- Plastic containers are essential consumables in the daily use of many bio-laboratories.
- Thus, this is important not only for phage-related research (e.g., the use of phage therapies as an alternative for antibiotics) but also for data comparison and reproducibility in the field of biochemistry and virology.
Resonant acoustic rheometry for non-contact characterization of viscoelastic biomaterials read more