Utilization of a rate enhancement hybridization buffer system for rapid in situ hybridization for the detection of porcine circovirus in cell culture and in tissues of pigs with postweaning multisystemic wasting syndrome

Theerapol Sirinarumitr, Igor Morozov, Porntippa Nawagitgul, Steven D. Sorden, Perry A. Harms, Prem S Paul

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

A rapid in situ hybridization (ISH) technique for the detection of porcine circovirus (PCV) nucleic acid in cell culture and formalin-fixed paraffin-embedded tissues was developed. A fluorescein-labeled RNA probe was transcribed from a plasmid containing 530 bp of the ORF1 of a PCV isolated from a pig with postweaning multisystemic wasting syndrome (PMWS). Hybridization using standard hybridization buffer was performed at 42 C for 16 hours and was compared to hybridization using rate enhancement hybridization (REH) buffer at 67 C for 2 hours. Hybridization was detected with an alkaline phosphatase-conjugated antifluorescein antibody. In both cultured cells and tissues from pigs with PMWS, the signal intensity and number of labeled cells in sections hybridized with REH buffer were equal to those of sections hybridized with standard hybridization buffer. The total time required for ISH using the REH buffer is 7-8 hours, thus making this protocol suitable for application in routine PCV diagnosis.

Original languageEnglish (US)
Pages (from-to)562-565
Number of pages4
JournalJournal of Veterinary Diagnostic Investigation
Volume12
Issue number6
DOIs
StatePublished - Jan 1 2000

Fingerprint

Circovirus
Porcine circovirus
Wasting Syndrome
postweaning multisystemic wasting syndrome
in situ hybridization
In Situ Hybridization
Buffers
cell culture
Swine
hybridization
buffers
Cell Culture Techniques
swine
RNA Probes
Fluorescein
Paraffin
Nucleic Acids
Formaldehyde
Alkaline Phosphatase
RNA probes

ASJC Scopus subject areas

  • veterinary(all)

Cite this

Utilization of a rate enhancement hybridization buffer system for rapid in situ hybridization for the detection of porcine circovirus in cell culture and in tissues of pigs with postweaning multisystemic wasting syndrome. / Sirinarumitr, Theerapol; Morozov, Igor; Nawagitgul, Porntippa; Sorden, Steven D.; Harms, Perry A.; Paul, Prem S.

In: Journal of Veterinary Diagnostic Investigation, Vol. 12, No. 6, 01.01.2000, p. 562-565.

Research output: Contribution to journalArticle

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