Comparison of treatment regimens to sensitize in situ hybridization for low-abundance calmodulin transcripts in the white matter of the rat spinal cord

Csaba Szigeti, Beatrix Kovacs, Elod Kortvely, K. Gulya

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

We compared two in situ hybridization protocols for the detection of low-abundance calmodulin (CaM) I mRNA populations in the lipid-rich white matter of the rat lumbar spinal cord. Digoxigenin-labeled, CaM I gene-specific antisense and sense cRNA probes were used in these experiments. Although microwave heating followed by chloroform and 0.1% triton X-100 treatment resulted in the specific labeling of several cells in the gray matter of the spinal cord with a low nonspecific signal, it did not detect any CaM I expressing cells in the white matter. The protocol involving a hybridization solution adjusted to a slightly alkaline pH (pH 8.0), however, resulted in the detection of a large number of CaM-expressing cells not only in the gray matter, but also in the white matter of the spinal cord, with a nonspecific hybridization signal that was essentially identical to that of the background, and it also retained a much better overall tissue quality as compared with the protocol involving microwave heating and triton X-100 treatment. Numerous medium-sized astrocyte-like cells and smaller cells resembling oligodendrocytes were detected that expressed CaM I mRNAs throughout the white matter of the lumbar spinal cord. Thus, in situ hybridization carried out at a slightly alkaline pH is a far superior method for the detection of low-abundance mRNA populations in lipid-rich regions of the central nervous system, such as the white matter areas, as compared with microwave heating combined with triton X-100 treatment.

Original languageEnglish
Pages (from-to)1-6
Number of pages6
JournalActa Biologica Szegediensis
Volume47
Issue number1-4
Publication statusPublished - 2003

Fingerprint

calmodulin
Calmodulin
spinal cord
in situ hybridization
In Situ Hybridization
Rats
Spinal Cord
Microwave heating
microwave treatment
Octoxynol
rats
Microwaves
Heating
Messenger RNA
cells
hybridization
Lipids
Digoxigenin
Complementary RNA
digoxigenin

Keywords

  • Alkaline pH
  • Calmodulin
  • In situ hybridization
  • Microwave
  • Rat
  • Spinal cord
  • Triton X-100
  • White matter

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Cell Biology
  • Neuroscience(all)
  • Applied Microbiology and Biotechnology

Cite this

Comparison of treatment regimens to sensitize in situ hybridization for low-abundance calmodulin transcripts in the white matter of the rat spinal cord. / Szigeti, Csaba; Kovacs, Beatrix; Kortvely, Elod; Gulya, K.

In: Acta Biologica Szegediensis, Vol. 47, No. 1-4, 2003, p. 1-6.

Research output: Contribution to journalArticle

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