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isolated lung systems
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Harvard Apparatus phone 508.893.8999 toll free U.S. 800.272.2775 fax 508.429.5732 www.harvardapparatus.com
PCLS Precision Cut Lung Slice Chamber
Assessment of lung functions under cell culture
conditions
From any species (murine, rat, human)
Study of airways of different sizes
Allows quantification of the responsiveness of
simple airway passages and simple vessels
Analyzing ciliary beating frequency
The incubation chamber was developed to allow incubation and observation of
slices by an inverted microscope. The chamber is made of Polycarbonate. It is
connected to a water bath to maintain constant temperature conditions. Two
incubation cells are positioned in the center of the chamber. The bottom of the
cells is sealed by glass, the cover is made of acrylic glass. The slices are fixed in
the incubation cells by positioning them under nylon strings fixed to a bent
platinum wire. The incubation cells can be filled with buffer, medium or drug
solutions through the filling pipe. Buffer solution can be removed from the cells
over a vacuum pipe. In addition, it is possible to gas the incubation cells in order
to use bicarbonate buffered media.
The incubation chamber is placed on the stage of an inverted microscope and
warmed to 37C. The slices are screened for airways and transferred to the
incubation chamber. Lung slices are selected for study using predefined criteria
(Martin et al. 1996). Airways and vessels are focused, and finally the images are
analyzed by image analysis software (e.g., Optimas or Metamorph).
Example of an Application
As an example Figure 1 shows exposure of a slice to increasing
concentrations of endothelin-1. Shown is a lung slice containing an airway (B),
a pulmonary artery (PA) and a pulmonary vein (PV). The pulmonary artery and
the airway contracted almost completely, while the pulmonary vein area
decreased to only 50% of its initial area. These responses are now easily
quantified by digital imaging technique.
It is a distinct advantage of this technique that in many ways precision-cut
slices can be treated like a cell culture. Thus, the slices can be incubated
under various conditions and gene as well as protein expression or mediator
release be determined. In contrast to cell culture models, in slices the
anatomical structure of the lung is largely maintained, so that the functional
consequences of gene expression and mediator release can be evaluated.
Figure 1. Exposure of a PCLS to increasing concentrations of endothelin-1.
Shown is one lung slice containing a small airway (B), a pulmonary artery (PA)
and a pulmonary vein (PV). The slice was imaged before (C) and after exposure to
increasing concentrations of endothelin-1, ranging from 10-10 M to 10-6 M.
Inlet From
Waterbath
Gas Inlet
Filling Pipes
Vacuum Pipes
Return to
waterbath
Platinum Wire with
Nylon Threads
Incubation Cells
Incubation Chamber Top View
Vacuum Pipes
Lung Functions Under the Microscope
Precision-cut lung slices (PCLS) offer a novel and unique way to assess lung
functions under cell culture conditions. They can be prepared from nearly any
species including mouse, rat and human lungs. The method allows the study of
the response of airways of different size (down to the terminal bronchioles) and
to relate these changes in lung functions to gene expression and mediator
release. Slices are viable for at least three days. They can be placed under an
inverted microscope, where digital image techniques allow quantification not
only of the responsiveness of single airways, but also of single vessels. In
addition, it is possible to analyze the ciliary beating frequency. More than 20
slices can be obtained from one lung, thus this method is very economical in
terms of experimental costs and animal use. Tissue cores are prepared from the
lungs filled with agarose solution, after cooling to 4C. From the cores, slices
(220 20m) are cut using a tissue slicer
Order # Product
73-2370 Precision Cut Lung Slice Chamber
Accessories
73-0125 Thermostatic Circulator E 103, 230 VAC,
73-2802 Thermostatic Circulator E 103, 115 VAC
73-0113 Roller Pump Reglo Analogue ISM 827, |
4 Channels, 0.002 to 30 ml/min
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