Zeiss Lightsheet Z.1 Operating Manual

Contents

CHAPTER 2 - SAMPLE PREPARATIONCarl Zeiss Sample Mounting for LSFM Lightsheet Z.122 000000-1790-528 02/2013A few points must be taken into account when choosing a particular mount:− Compatibility. This is a crucial issue. The mount must be compatible with the object you want toimage (chemistry, temperature etc.), but it must also be compatible with the stage holder.− Stability (mechanical, optical, chemical).− Tightness. In the case of embedded samples, once the gel has solidified, the cylinder of gellingagent is pushed through the capillary out of the distal end by a plunger fitting into the capillary.The system must be air tight to avoid air entry leading to a displacement of the gel rod. Theplunger can be sealed with a drop of wax, acrylamide or nail polish, i.e. anything that prevents theplunger and hence the agarose containing the sample from moving.− Cost.2.3.3 Sample HolderFor technical description of the sample chamber, hearting devices and sample holder, you can refer toCHAPTER 1 HARDWARE.Once the specimen is prepared and properly labeled, it is ready to be imaged. While in conventionalimaging there is a suitable platform on which to place the glass slide or the chamber, in LSFM the objectmust be held from above via the sample holder. Depending on the size of the sample there are twodifferent types of sample holders available: sample holder for capillaries and syringes (Fig. 14/C-F and I).Always use the minimal cylinder diameter necessary for your specimen size to avoid excessive amounts ofagarose.The largest sample holder has been designed to accommodate a 1ml syringe that can be inserted fromthe top with a plunger that can be operated once the sample holder is mounted on the stage. Onceinserted, the syringe is perfectly fitted to the sample holder as the two flaps used for injection fit theupper part of the holder. In this way the object support is well maintained, an essential issue for imagingand multiview imaging as the object is moved through the light sheet by the stage.Capillaries have been extensively used to image small embedded objects, as hooks for very large objects,and as support for enclosed objects, so the capillary has become commonly used for LSFM sampleembedding.Capillaries are made of glass. They can break. They are also sliding when wet. Please handlethem with care and dispose them properly.