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Biology

Bir Deniz Tubeworm Canlı Optik ve Elektron Mikroskobu Teknikleri Kullanılarak kalsifikasyon Olaylar Karakterizasyonu

Published: February 28, 2017 doi: 10.3791/55164
Automatic Translation
1, 2, 3, 3, 4, 1
1Department of Biological Sciences, Clemson University, 2Department of Marine Biodiversity Research (BioDive), Japan Agency for Marine-Earth Science and Technology (JAMSTEC), 3Advanced Material Research Laboratory (AMRL), Clemson University, 4Swire Institute of Marine Sciences and School of Biological Sciences, The University of Hong Kong

Introduction

Biyomineralizasyon zarif sipariş mineral 1 üretimi ile sonuçlanan hücre faaliyetlerinin bir paketi köprü etkinlikleri kompleks dizisidir. meydan dinamik hücresel süreci ve optik ve elektron mikroskopisi yöntemleri bir arada kullanarak sofistike mineral yapıları hem karakterize etmektir. Hücre içi pH yükselmesi dolayısıyla, artan bir pH değerine sahip döneminde ortaya tanımlayan kalsifikasyon 2, 3 meydana olması muhtemeldir zaman ortaya Caco 3 kristallerin oluşumunu desteklemektedir.

Etti Serpulidae gelen tüp kurtları okyanus 4 ortak calcifiers bulunmaktadır. Özellikle biyolojik kirliliğe 5, 6, aynı zamanda deniz araştırmaları için popüler bir omurgasız modelidir. Bu çalışmada, hidrotermal bölümlerinde kalsifikasyon süreci During Biyomineralizasyon görülmektedir. Metamorfoz hızlı işlem kalsiyum karbonat yapıları 7, 8 çıkmasını içerir.

Biz tubeworm yapılabilir nasıl iç pH ölçümleri göstermek ve yaşam evreleri ve kalsifikasyon hakkında dokular nasıl taranabilir. ilgi hayat aşaması tanımlandıktan sonra, kireçlenme sorumlu doku elektron mikroskopisi yöntemleri kullanılarak daha yüksek bir çözünürlükte karakterize edilebilir. Floresan mikroskobu kullanarak, metamorfik indüksiyondan sonra görünmesi kalsiyum karbonat için gereken zamanı belirler. hayatın benzer bir sahne daha sonra element bileşimi dağıtımı için SEM-EDS ile görüntülendi ve biriken mineral, iki farklı elektron mikroskopi yöntemleri, özellikle SEM-EBSD ve FIB-TEM kullanılarak analiz edildi.

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Materials

Name Company Catalog Number Comments
Hexamethyldisilazane  Electron Microscopy Sciences 16700(EM)
Osmium Tetroxide 2% Aqueous Solution Electron Microscopy Sciences 19192
IBMX 3-Isobutyl-1-methylxanthine ThermoFisher Scientific PHZ1124
Nigericin, Free Acid ThermoFisher Scientific N7143-5MG
35 mm diameter dish, hole size 27 mm, Glass No.0, Non-coat ThermoFisher Scientific D110400
5-(and-6)-Carboxy SNARF-1, Acetoxymethyl Ester, Acetate ThermoFisher Scientific C-1271
BDH Potassium Chloride, ACS Grade VWR BDH0258-500G
Paraformaldehyde
reagent grade, crystalline		 Sigma P6148
1 M Hydrochloric Acid for Volumetric Analysis Wako Pure Chemical Industries, Ltd 083-01095
0.05 M Sodium Hydroxide Solution for Volumetric Analysis Wako Pure Chemical Industries, Ltd 199-02185
Calcein Sigma C0875
FASW Iwaki Co. Ltd. Rei-sea Marine
Mixed Cellulose Ester Membranes; 47 mm dia, 0.45 µm ADVANTEC A045A047A
ethanol Wako Pure Chemical Industries, Ltd 051-00476
Artificial seawater for buffers by SOP06 of DOE (1994), cdiac.ornl.gov/ftp/cdiac74/sop06.pdf
Sodium Chloride Wako Pure Chemical Industries, Ltd 191-01665
Potassium Chloride Wako Pure Chemical Industries, Ltd 163-03545
Magnesium Chloride Hexahydrate Wako Pure Chemical Industries, Ltd 135-00165
Calcium Chloride Wako Pure Chemical Industries, Ltd 039-00475
Sodium Sulfate Wako Pure Chemical Industries, Ltd 197-03345
Hydrochloric Acid Wako Pure Chemical Industries, Ltd 089-08415
2-amino-2-hydroxymethyl-1,3-propanediol (tris) Wako Pure Chemical Industries, Ltd 207-06275
2-aminopyridine Wako Pure Chemical Industries, Ltd 011-02775
Orion 5-star Plus pH meter Thermo Scientific
PrpHecT ROSS Micro Combination pH Electrode 8220BNWP Thermo Scientific
Axiovision, Version 4.6, Axio Observer Z1 Zeiss
ImageJ NIH, Bethesda, MD, USA
HRTEM H500 Hitachi
SU6600 VPSEM Hitachi
NB5000 Focused Ion and Electron Beam (FIB-SEM) system Hitachi 

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References

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Bir Deniz Tubeworm Canlı Optik ve Elektron Mikroskobu Teknikleri Kullanılarak kalsifikasyon Olaylar Karakterizasyonu
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Cite this Article

Chan, V. B. S., Toyofuku, T.,… More

Chan, V. B. S., Toyofuku, T., Wetzel, G., Saraf, L., Thiyagarajan, V., Mount, A. S. Characterization of Calcification Events Using Live Optical and Electron Microscopy Techniques in a Marine Tubeworm. J. Vis. Exp. (120), e55164, doi:10.3791/55164 (2017).

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