A new flow cytometric transducer for fast sample throughput and time resolved kinetic studies of biological cells and other particles

Abstract

In state of the art flow cytometric transducers, the cells are supplied through tubes. Passage through the tube and washing between different samples is time consuming and limits the number of samples that can be processed in a given time. This is a drawback particularly with automatic routine instruments. For kinetic studies in the time range of seconds, it is necessary to perform the cell reactions directly in the transducer in order to have a short delay between the suspension vessel where the cell reaction is in progress and the point of measurement. A new one parameter electrical sizing transducer without a particle supplying tube is described and compared with a conventional Metricell transducer. The cells are directly supplied from an exchangeable vessel to the measuring point in the transducer. The vessel which is an inexpensive mass produced product, serves as the injection tip for passing the cell suspension into the focusing flow path. There is no interconnected tube to delay or intermix the cells in the stage between the reaction in progress in the vessel and flow analysis. Delay times of only 1 second are achieved with the new transducer, and by supplying each sample with its own vessel subsequent samples are analyzed without the necessity of cleaning a cell supplying tube. In this way a high throughput of samples per time unit is achieved and time kinetic experiments in the time range of seconds can be performed. The design of a tubeless multiparameter Fluvo Metricell transducer is explained.

Real-Time Spectroscopic Analysis of Ligand-Receptor Dynamics

“…A novel application of the system was in examination of ligand binding during continuous infusion of stimulus to understand the infl uence of rate of receptor occupancy as well as the number of receptors occupied in cell activation. Kachel et al (48) described a sample transducer with a delay of 1 sec. There is a clear need for a systematic adaptation of rapid sampling techniques to flow cytometry.…”

Application of the CYTOMIC 12 flow cytometric compact analyzer for automatic kinetic measurements

“…2. Example of a one parameter time sequence experiment with sheep erythrocytes (8) which were sized in a Metricell flow cytometer (2) equipped with a new tubeless transducer (5). A , isometric view onto the 64 one parameter erythrocyte volume distributions sequentially arranged in the form of a pseudo two parameter histogram.…”

“…For fast kinetic experiments in the time range of seconds, special transducers are required where the cells are directly introduced from the reaction vessel into the sensing zone of the flow cytometer ( 5 ) . At the same time, the requirements for the data acquisition and processing equipment differ from the requirements for conventional systems in which the cells are considered to have features that are stable with time.…”

Very early detection of changes associated with cellular activation using a modified flow cytometer

“…Key terms: Kinetics, calcium mobilization, Flow cytometric observation of early changes in cellular activation parameters has been hindered by the need to remove the sample in order to add reagents and by the time required to mix the sample and reestablish stable sample flow, typically 10-15 s. Several investigators have addressed the importance of on-line reagent addition andlor sample transit time in flow cytometry (1,(3)(4)(5)(6)(7). Increased dynamic resolution of cellular responses has application in the measurement of changes in intracellular Ca2+ concentration, membrane depolarization, changes in intracellular pH, oxidative bursts, and ligand-receptor binding.…”