Optimal probe size and fixation time for the detection of Porcine circovirus-2 DNA by in situ hybridization in formalin-fixed, paraffin-embedded tissue.
Probe size and fixation time for detecting Porcine circovirus-2 (PCV-2) by in situ hybridization in formalin-fixed, paraffin-embedded lymph nodes from experimentally infected pigs were optimized. In situ hybridization using a 169-base pair (bp) probe detected significantly fewer PCV-2-positive cells than when using 8 other larger probes (P < 0.05). The difference in hybridization intensity between smaller probes (169 and 225 bp) and larger probes (416, 473, 571, 631, 693, and 753 bp) was statistically significant (P < 0.05). The PCV-2-positive cells were consistently detected in lymph nodes fixed up to 3 days; thereafter, the number of positive cells declined. The PCV-2-positive cells were detected in lymph nodes fixed for up to 730 days. The difference in hybridization intensity between samples fixed for a short term (1 or 3 days) and a longer term (4-730 days) was statistically significant (P < 0.05). The data demonstrates that the optimal probe size and fixation time for detecting PCV-2 in formalin-fixed, paraffin-embedded lymph nodes is 473 bp and 1-3 days, respectively.
Ha
Yooncheol
Y
Department of Veterinary Pathology, College of Veterinary Medicine, Seoul National University, San 56-1, Shillim-dong, Gwanak-gu, Seoul 151-742, Republic of Korea.
Chae
Chanhee
C
D000818:Animals; D018173:Circoviridae Infections / Q000235:genetics / Q000473:pathology / Q000662:veterinary; D017925:Circovirus / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D017931:DNA Primers; D004279:DNA, Viral / Q000032:analysis / Q000235:genetics*; D008198:Lymph Nodes / Q000821:virology; D009693:Nucleic Acid Hybridization / Q000379:methods; D016612:Paraffin Embedding; D012680:Sensitivity and Specificity; D013552:Swine; D013553:Swine Diseases / Q000175:diagnosis / Q000235:genetics / Q000473:pathology
D017931:DNA Primers; D004279:DNA, Viral