Many in vivo tissue responses begin locally, yet most in vitro stimuli are delivered globally. Microfluidics has a unique ability to provide focal stimulation to tissue samples with precise control over fluid location, flow rate, and composition. However, previous devices utilizing fixed ports beneath the tissue required manual alignment of the tissue over the ports, increasing the risk of mechanical damage. Here we present a novel microfluidic device that allows the user to define the location of fluid delivery to a living tissue slice without manipulating the tissue itself. The device utilized a two-component SlipChip design to create a mobile port beneath the tissue slice. A culture chamber perforated by an array of ports housed a tissue slice and was separated by a layer of fluorocarbon oil from a single delivery port, fed by a microfluidic channel in the movable layer below. We derived and validated a physical model, based on interfacial tension and flow resistance, to predict the conditions under which fluid delivery occurred without leakage into the gap between layers. Aqueous solution was delivered reproducibly to samples of tissue and gel, and the width of the delivery region was controlled primarily by convection. Tissue slice viability was not affected by stimulation on the device. As a proof-of-principle, we showed that live slices of lymph node tissue could be sequentially targeted for precise stimulation. In the future this device may serve as a platform to study the effects of fluid flow in tissues and to perform local drug screening.
The lymph node is a highly organized
and dynamic structure that
is critical for facilitating the intercellular interactions that constitute
adaptive immunity. Most ex vivo studies of the lymph node begin by
reducing it to a cell suspension, thus losing the spatial organization,
or fixing it, thus losing the ability to make repeated measurements.
Live murine lymph node tissue slices offer the potential to retain
spatial complexity and dynamic accessibility, but their viability,
level of immune activation, and retention of antigen-specific functions
have not been validated. Here we systematically characterized live
murine lymph node slices as a platform to study immunity. Live lymph
node slices maintained the expected spatial organization and cell
populations while reflecting the 3D spatial complexity of the organ.
Slices collected under optimized conditions were comparable to cell
suspensions in terms of both 24-h viability and inflammation. Slices
responded to T cell receptor cross-linking with increased surface
marker expression and cytokine secretion, in some cases more strongly
than matched lymphocyte cultures. Furthermore, slices processed protein
antigens, and slices from vaccinated animals responded to ex vivo
challenge with antigen-specific cytokine secretion. In summary, lymph
node slices provide a versatile platform to investigate immune functions
in spatially organized tissue, enabling well-defined stimulation,
time-course analysis, and parallel read-outs.
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