Easy flat embedding of oriented samples in hydrophilic resin (LR White) under controlled atmosphere: application allowing both nucleic acid hybridizations (CARD-FISH) and ultrastructural observations

Acta histochemica
Olivier Gros, Leslie Candice Maurin

Abstract

Hydrophilic resins present the advantage of making possible both hybridization experiments involving either antibodies or oligonucleotide probes and ultrastructural observations. Whereas various embedding protocols are available, only very few concern flat-embedded preparations. In this study we describe an easy protocol for flat embedding of small-oriented biological samples in hydrophilic resins (LR White). The most important constraints are (i) to polymerize the samples under argon-saturated atmosphere (avoiding oxygen which is an inhibitor of LR White polymerization) and (ii) to use transparent flat embedding molds. Two kinds of samples were analyzed: small pieces of large tissue that need to be accurately oriented for a valuable analysis and very small organisms such as free-living nematodes, which are very hard to investigate with conventional paraffin wax embedding techniques. Semi-thin sections strongly reinforce the quality of the observations from oligonucleotidic in situ hybridization experiments by reducing the background usually encountered in oligonucleotide probe hybridization experiments from sections. Such protocols could also permit a cheap alternative to the use of laser scanning confocal microscopes for olig...Continue Reading

References

Oct 7, 2004·Applied and Environmental Microbiology·Raju SekarJakob Pernthaler
Nov 3, 2004·Journal of the American Society for Mass Spectrometry·Alan M KleinfeldClaude Lechene
May 20, 2006·Nature·Andrea D NussbaumerMonika Bright
Feb 22, 2007·Proceedings. Biological Sciences·Mathieu PerniceNicole Dubilier

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