Zeiss Lightsheet Z.1, Operating Manual

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CHAPTER 2 - SAMPLE PREPARATION
Lightsheet Z.1 Sample Mounting for LSFM Carl Zeiss
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Another possibility is to use a polymer to make the chamber. The polymer, similar to the gelling agent,
must be transparent or at least have an optical index as close as possible to water or the buffer used
during the experiment. The polymer is usually used as a sheet that can be formed as required. The other
possibility is to approach commercial manufacturers to make polymer chambers at the specific sizes and
lengths required.
Fusing polymer sheets can be done using a welding iron with controlled temperature or a welding device
use for melting together plastic bags. As described in Fig. 12, the polymer foil is folded to an appropriate
size. This can be made easier by using a guide or template, in this case a micropipette. The polymer is
fused together. The tube generated is finally fused together on the other side to make a complete
container. The polymer chamber can be easily mounted on the LSFM by using a clip, a slotted metal
capillary or glued to a micropipette. However, the last two options have the disadvantage of partially
obscuring the field of view.
This technique has been successfully used to image living cells (Engelbrecht et al., 2007) and cell clusters
(Pampaloni et al., 2007).
2.2.4 FEP Tubing
More recently, the availability of Fluorinated Ethylene Propylene (FEP) tube of different diameters has
been successfully used for long term imaging of Zebrafish embryos (Kaufman et al., 2012). Here, we refer
only to the paper by Anna Kaufmann, Michaela Mickoleit, Michael Weber and Jan Huisken in
Development 139, 3242-3247 (2012) ("Multilayer mounting enables long-term imaging of zebrafish
development in a light sheet microscope") and emphasize the fact that the mounting method described
in this article is fully compatible with the Lightsheet Z.1.
Fig. 12 Making and mounting an incubation polymer foil chamber.
A piece of polymer foil (A) is folded and either heat- or glue-sealed to generate a tube of a pre-
defined size (B). Excess foil can be removed or used to glue the tube on to a specific holder (capillary,
thread, metal rod…) (D). One side of the tube can be then glued or heat-sealed to close the chamber
(C). The polymer used must be suitable for microscopy and easy to seal. The chamber can be glued to
a support, held by forceps, or inserted into a slit rod.