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Peer-reviewed

Preparation of Candida albicans Biofilms for Transmission Electron Microscopy

制备白色念珠菌生物膜用于透射电镜分析

HT Heather T. Taff
David R. Andes David R. Andes

发布: 2013年07月20日第3卷第14期 DOI: 10.21769/BioProtoc.822 浏览次数: 10684

评审: Fanglian He

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Cover of PLOS Pathogens, featuring study using the protocol.

Aug 2012

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Abstract

Transmission Electron Microscopy is a form of microscopy that allows for imaging of distinct portions of an individual cell. For Candida albicans biofilms, it is often used to visualize the cell walls of fixed samples of yeast and hyphae. This protocol describes how to grow, harvest, and fix Candida albicans biofilms in preparation for Transmission Electron Microscopy.

Materials and Reagents

  1. Plate containing grown Candida albicans colonies
  2. 500 ml 0.22 μm filter (Corning, catalog number: 431118 )
  3. Aluminum foil
  4. Bacto-peptone (BD Biosciences, catalog number: 211677 )
  5. Bacto-yeast extract (BD Biosciences, catalog number: 212750 )
  6. Cell scraper (BD Biosciences, Falcon®, catalog number: 353085 )
  7. Difco Dextrose (BD Biosciences, catalog number: 215530 )
  8. Glutaraldehyde (Sigma-Aldrich, catalog number: G5882 )
  9. Lead nitrate (PbNO3) (Electron Microscopy Sciences, catalog number: 17900 )
  10. MOPS (Thermo Fisher Scientific, catalog number: BP308 )
  11. 4% Osmium tetroxide (Electron Microscopy Sciences, catalog number: 19140 )
  12. Parafilm
  13. Paraformaldehyde (Electron Microscopy Sciences, catalog number: 15710 )
  14. Propylene oxide (Electron Microscopy Sciences, catalog number: 20410 )
  15. RPMI 1640 (Sigma-Aldrich, catalog number: R6504 )
  16. Sodium cacodylate trihydrate (Sigma-Aldrich, catalog number: C0250 )
  17. Sodium chloride (Thermo Fisher Scientific, catalog number: S671-3 )
  18. Sodium citrate (Electron Microscopy Sciences, catalog number: 21140 , 500 gm)
  19. Sodium hydroxide (Sigma-Aldrich, catalog number: 59223C )
  20. Sodium phosphate dibasic anhydrous (Thermo Fisher Scientific, catalog number: BP332-500 )
  21. Sodium phosphate monobasic monohydrate (Thermo Fisher Scientific, catalog number: S369-500 )
  22. Spurr’s Resin Kit (Polyscience, catalog number: 01916-1 )
  23. Uridine (Sigma-Aldrich, catalog number: U3750 )
  24. Uranyl acetate (Electron Microscopy Sciences, catalog number: 22400 )
  25. YPD + uridine (see Recipes)
  26. RPMI + MOPS (see Recipes)
  27. 1x PBS (see Recipes)
  28. Fixative (see Recipes)
  29. Reynold’s Lead Citrate Strain (see Recipes)

Equipment

  1. 15 ml conical tube (Corning, catalog number: CLS430055 )
  2. 6 well polystyrene plate (Grenier Bio-one, catalog number: 657160 )
  3. Hemocytometer
  4. Shaking incubator with adjustable temperatures and speeds
  5. Transmission Electron Microscope facility with trained microscopist

Procedure

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文章信息

版权信息

© 2013 The Authors; exclusive licensee Bio-protocol LLC.

如何引用

Taff, H. T. and Andes, D. R. (2013). Preparation of Candida albicans Biofilms for Transmission Electron Microscopy. Bio-protocol 3(14): e822. DOI: 10.21769/BioProtoc.822.

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分类

微生物学 > 微生物生物膜 > 生物膜培养

微生物学 > 微生物细胞生物学 > 细胞成像

细胞生物学 > 细胞成像 > 电子显微镜

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