Dataset Information

From Microtiter Plates to Droplets-There and Back Again.

ABSTRACT: Droplet-based microfluidic screening techniques can benefit from interfacing established microtiter plate-based screening and sample management workflows. Interfacing tools are required both for loading preconfigured microtiter-plate (MTP)-based sample collections into droplets and for dispensing the used droplets samples back into MTPs for subsequent storage or further processing. Here, we present a collection of Digital Microfluidic Pipetting Tips (DMPTs) with integrated facilities for droplet generation and manipulation together with a robotic system for its operation. This combination serves as a bidirectional sampling interface for sample transfer from wells into droplets (w2d) and vice versa droplets into wells (d2w). The DMPT were designed to fit into 96-deep-well MTPs and prepared from glass by means of microsystems technology. The aspirated samples are converted into the channel-confined droplets' sequences separated by an immiscible carrier medium. To comply with the demands of dose-response assays, up to three additional assay compound solutions can be added to the sample droplets. To enable different procedural assay protocols, four different DMPT variants were made. In this way, droplet series with gradually changing composition can be generated for, e.g., 2D screening purposes. The developed DMPT and their common fluidic connector are described here. To handle the opposite transfer d2w, a robotic transfer system was set up and is described briefly.

SUBMITTER: Henkel T

PROVIDER: S-EPMC9316479 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:BackgroundConventional crop protection has major drawbacks, such as developing pest and pathogen insensitivity to pesticides and low environmental compatibility. Therefore, alternative crop protection strategies are needed. One promising approach treats crops with chemical compounds that induce the primed state of enhanced defense. However, identifying priming compounds is often tedious as it requires offline sampling and analysis. High throughput screening methods for the analysis of priming-active compounds have great potential to simplify the search for such compounds. One established method to identify priming makes use of parsley cell cultures. This method relies on measurement of fluorescence of furanocoumarins in the final sample. This study demonstrates for the first time the online measurement of furanocoumarins in microtiter plates. As not all plants produce fluorescence molecules as immune response, a signal, which is not restricted to a specific plant is required, to extend online screening methods to other plant cell cultures. It was shown that the breathing activity of primed parsley cell cultures increases, compared to unprimed parsley cell cultures. The breathing activity can by monitored online. Therefore, online identification of priming-inducing compounds by recording breathing activity represents a promising, straight-forward and highly informative approach. However, so far breathing has been recorded in shake flasks which suffer from low throughput. For industrial application we here report a high-throughput, online identification method for identifying priming-inducing chemistry.ResultsThis study describes the development of a high-throughput screening system that enables identifying and analyzing the impact of defense priming-inducing compounds in microtiter plates. This screening system relies on the breathing activity of parsley cell cultures. The validity of measuring the breathing activity in microtiter plates to drawing conclusions regarding priming-inducing activity was demonstrated. Furthermore, for the first time, the fluorescence of the priming-active reference compound salicylic acid and of furanocoumarins were simultaneously monitored online. Dose and time studies with salicylic acid-treated parsley cell suspensions revealed a wide range of possible addition times and concentrations that cause priming. The online fluorescence measuring method was further confirmed with three additional compounds with known priming-causing activity.ConclusionsDetermining the OTR, fluorescence of the priming-active chemical compound SA and of furanocoumarins in parsley suspension cultures in MTPs by online measurement is a powerful and high-throughput tool to study possible priming compounds. It allows an in-depth screening for priming compounds and a better understanding of the priming process induced by a given substance. Evaluation of priming phenomena via OTR should also be applicable to cell suspensions of other plant species and varieties and allow screening for priming-inducing chemical compounds in intact plants. These online fluorescence methods to measure the breathing activity, furanocoumarin and SA have the potential to accelerate the search for new priming compounds and promote priming as a promising, eco-friendly crop protection strategy.

Dataset Information

From Microtiter Plates to Droplets-There and Back Again.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets