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ZEISS Lightsheet Z.1Sample Preparation


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ZEISS Lightsheet Z.1

Sample Preparation

Author: Flood P.M., Kelly R., Gutirrez-Heredia L.and E.G. Reynaud

School of Biology and Environmental Science,University College Dublin, Ireland

Date: Sept 2013

This paper describes theoretical and practical aspects of sample preparation for Light Sheet Fluorescence

Microscopy (LSFM). We present general rules for sample handling and mounting, as well as guidelines with

respect to the best preparative technique to use, taking into account sample type, structure and properties.

Step by step protocols and recommended materials for ZEISS Lightsheet Z.1 samples are included.

These protocols cover sample preparation ranging from micrometer-sized fluorescent beads to millimeter-sized

insects, providing detailed information relating to preparation and observation techniques. Finally, this paper

identifies the main artifacts and problems that can result from the preparation techniques.

CONTENTS (click text to go to page):

1 INTRODUCTION

2 SAMPLE MOUNTING FOR LSFM

2.1 The perfect Sample for LSFM

2.2 Holding the Sample

2.2.1 Embedded Samples

2.2.2 Hanging Samples

2.2.3 Enclosed Samples

2.2.4 FEP Tubing

2.3 Materials and Equipment

2.3.1 Sample Chambers

2.3.2 Molding and Mounting Supports

2.3.3 Sample Holder

2.3.4 Gels and Polymers

2.3.5 Hydrogel Preparation

2.4 Fixation and Fixatives

2.5 Stains and Staining

2.5.1 Choosing a Fluorescent Label

2.6 Antifading Agents

2.7 Cleaning, Labelling and Storing Samples

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3 SPECIFIC EXAMPLES OF SAMPLE

PREPARATION3.1 Preparation of Fluorescent Beads

3.2 Preparation of a Medaka Fish Embryo

(Oryza latypes)

3.3 Preparation of a Fly Pupa

(Drosophila melanogaster)

3.4 Preparation of a Plant Root

(Arabidopsis thaliana)

3.5 Imaging Cell Cysts in an Extracellular Matrix Gel

3.6 Immunostaining and Preparation of

MDCK Cell Cysts

3.7 Preparation of a Whole Mount of a Mosquito

(Anopheles gambiae)

4 TIPS, TROUBLESHOOTING AND

ADDITIONAL INFORMATION

4.1 Tips

4.2 Troubleshooting

4.3 Suggested Additional Sources of Information

4.4 References and Further Reading

5 Index


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EN_41_011_058|CZ10-2013|Design,scopeofdeliveryandtechnicalprogresssubjecttochangewithoutnotice.|CarlZeissMicroscopyGmbH
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lightsheetz1 sample-preparation zeiss (1)

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