Comparison of polymerase chain reaction and immunohistochemistry for the detection of Mycoplasma pulmonis in paraffin-embedded tissue.
The polymerase chain reaction (PCR) is a reliable method of detecting specific DNA sequences. The purpose of this study was to develop a PCR method of detecting Mycoplasma pulmonis in paraffin-embedded tissue sections and to compare the sensitivity of this method with that of the avidin-biotin complex (ABC) immunohistochemistry technique. We infected mice intranasally with 10(7) M. pulmonis and sacrificed them 28 days later. Using a published oligonucleotide primer sequence for M. pulmonis, we examined 8-microns paraffin sections by PCR for a 710 base-pair segment of the genome. A sequential 8-microns paraffin section was stained for M. pulmonis, using standard ABC immunohistochemistry methods. By PCR and dot blot hybridization, 60 of 62 paraffin-embedded lungs were positive for M. pulmonis, whereas only 17 of 62 lungs were positive, using the ABC method. In this study, we demonstrated that PCR of paraffin-embedded tissues followed by dot blot hybridization of the PCR products was much more sensitive than the ABC method in identifying mouse lungs infected with M. pulmonis.
Brunnert
S R
SR
Institute of Comparative Medicine, College of Physicians & Surgeons, Columbia University, New York, NY 10032.
Dai
Y
Y
Kohn
D F
DF
D000818:Animals; D001360:Avidin; D001483:Base Sequence; D001710:Biotin; D004269:DNA, Bacterial / Q000032:analysis; D007150:Immunohistochemistry; D008168:Lung / Q000382:microbiology* / Q000473:pathology; D051379:Mice; D008969:Molecular Sequence Data; D009174:Mycoplasma / Q000235:genetics / Q000302:isolation & purification*; D009693:Nucleic Acid Hybridization; D010232:Paraffin; D016133:Polymerase Chain Reaction; D013047:Specific Pathogen-Free Organisms; D016610:Tissue Embedding
D004269:DNA, Bacterial; D001360:Avidin; D001710:Biotin; D010232:Paraffin