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Summary

Abstract

Introduction

Protocol

Representative Results

Discussion

Acknowledgements

Materials

References

Neuroscience

The Cutting and Floating Method for Paraffin-embedded Tissue for Sectioning

Published: September 5th, 2018

DOI:

10.3791/58288

1Institute of Life Science, Nanchang University, 2Queen Mary School, Medical Department, Nanchang University, 3School of Life Science, Nanchang University
* These authors contributed equally

Here, we present a protocol to improve paraffin sectioning. This method combines cutting and floating using a simple thermostatic chamber to avoid the transfer process required by the conventional method. As a result, the efficiency and the number of intact paraffin sections were greatly improved.

Sectioning of the paraffin-embedded tissue is widely used in histology and pathology. However, it is tedious. To improve this method, several commercial companies have devised complex section transfer systems using fluid water. To simplify this technology, we created a simple method using homemade equipment that combines cutting and floating within a simple thermostatic chamber; therefore, the sections automatically enter the water bath on the water surface. The hippocampus from adult mouse brains, adult mouse kidneys, embryonic mouse brains, and adult zebrafish eyes were cut using both conventional paraffin sectioning and the presented method for comparison. Statistical analysis shows that our improved method saved time and produced higher quality sections. In addition, paraffin sectioning of a whole specimen in a short time is easy for junior operators.

Morphological study is important in the biological research. Although new technology has allowed researchers to observe their targets directly from the whole tissue or organisms1,2,3, cutting the specimen into thin sections, followed by staining, remains the primary method fornot only tissue morphology but also protein targeting directly in the tissue. Light microscopy uses three section types: paraffin, frozen, and semithin. Although cryosectioning is common for protecting tissue antigenicity, and the specimen preparation is simple, the retained tissue morphology is poor and....

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All methods described here have been approved by the Animal Care and Use Committee of Nanchang University.

1. Assemble the Equipment and Connect the Microtome

  1. Design the parameter per the requirements (Supplementary Figure 1).
  2. Submit the parameter to a local factory to manufacture the acrylic boards.
  3. Assemble all parts in sequence: Use chloroform to combine 7 commercial acrylic boards into a tank with a section chan.......

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The improved method increased the number of intact paraffin sections. We tested this new method on adult mouse hippocampal tissue, adult mouse kidneys, embryonic mouse brains, and zebrafish eyes. Water was added to the tank, and the water temperature was maintained between 38.0 °C to 40.0 °C. After a serially preparing the tissue samples, they were sectioned and compared to conventional sectioning. The new method avoided section loss and increased the proportion of intact sectio.......

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To improve paraffin section morphology and solve the problem of wasted time during conventional paraffin sectioning, we created an improved paraffin sectioning method that combines cutting and unfolding. This improved method relies on simple equipment that comprises a section channel, a water bath and a heater with a temperature detection switch. The section ribbon enters the water bath through the section channel and unfolds automatically while cutting. Therefore, this method improves paraffin sectioning quality and eff.......

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This work was supported by the National Natural Science Foundation of China (Grant No. 31400936, 31460260) and the Natural Science Foundation of Jiangxi Province of China (20171BAB215020). We also thank the joint program between Nanchang University and Queen Mary University of London for supporting this work.

....

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Name Company Catalog Number Comments
Incubator Boekel Scientific 133000-2
Ethanol  Sinopharm Chemical Reagent Co.,Lid 64-17-5
Xylene  Sinopharm Chemical Reagent Co.,Lid 1330-20-7
Paraplast Leica 39601006
Heated Paraffin Embedding Module Leica
Commercial acrylic board
Trichloromethane Sinopharm Chemical Reagent Co.,Lid 67-66-3
Tubular electric heating element(12V 200W)
Temperature controller(12v 120w) Mingsuo XH-W3002
Rotary microtome  Leica
Neutral silicone sealant Link the water channel with the microtome knife holder
Voltage transformer Dearll S-250-12
Disposable blade Accu-Edge 4689
Hematoxylin Baso Diagnostics Inc. BA-4025
Eosin  Baso Diagnostics Inc. BA-4025
Microslide Sail Brand 7105
Neutral balsam Sinopharm Chemical Reagent Co.,Lid 10004160
Coverslip  Citoglas 10212424C
Microscope Carl Zeiss
Hydrochloric acid Xilong Chemical 7647-01-0
Water bath for paraffin sections Leica
HistoCore Arcadia C - Cold Plate Leica
paraffin repellent spray  Thermo Scientific 9990420

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