Chromogenic in-situ hybridization technique for the detection of porcine circovirus 3 in formalin-fixed paraffin-embedded lung and lymphoid tissues
Background and Aim: Porcine circovirus 3 (PCV3) was recently reported in Malaysian commercial pig population in 2020 by conventional polymerase chain reaction (PCR), revealing a molecular prevalence of 17.02 in the sampled domestic pig population. This study aims to describe a chromogenic in situ hy...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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Veterinary World
2023
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| Online Access: | http://psasir.upm.edu.my/id/eprint/107025/ http://psasir.upm.edu.my/id/eprint/107025/1/Chromogenic%20in-situ%20hybridization%20technique%20for%20the%20detection%20of%20porcine%20circovirus%203%20in%20formalin-fixed%20paraffin-embedded%20lung%20and%20lymphoid%20tissues.pdf |
| _version_ | 1848864855120936960 |
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| author | Tan, Chew Yee Lee, Kah Chun Chiou, Ming-Tang Lin, Chao-Nan Ooi, Peck Toung |
| author_facet | Tan, Chew Yee Lee, Kah Chun Chiou, Ming-Tang Lin, Chao-Nan Ooi, Peck Toung |
| author_sort | Tan, Chew Yee |
| building | UPM Institutional Repository |
| collection | Online Access |
| description | Background and Aim: Porcine circovirus 3 (PCV3) was recently reported in Malaysian commercial pig population in 2020 by conventional polymerase chain reaction (PCR), revealing a molecular prevalence of 17.02 in the sampled domestic pig population. This study aims to describe a chromogenic in situ hybridization (ISH) technique using digoxigenin (DIG)- labeled cloned PCV3 open reading frame 1 (ORF1) fragment DNA to detect and localize the PCV3 antigen in formalinfixed, paraffin-embedded lung, and lymphoid tissue specimens. Materials and Methods: Since PCV3 was mainly detected in lung and lymphoid tissues, we obtained tissue specimens from these organs from the previous Malaysian PCV3 study. Digoxigenin-labeled ISH probes were designed to target a 69 bp region of PCV3 ORF1 spanning from the nucleotide positions (282“350). Results: Light microscopy analysis revealed that chromogenic staining of PCV3 antigens was visualized within the cytoplasm of pneumocytes and lymphocytes, indicating positive ISH results. The results of molecular detection of PCV3 using PCR and ISH showed a high agreement of 90.91, including for the negative PCV3 status for all samples. Conclusion: This study reports a chromogenic ISH technique using DIG-labeled probes targeting PCV3 ORF1 to detect PCV3 antigens in lung and lymphoid tissues. Despite the limited availability of PCV3 antibodies, ISH remains relevant for investigating PCV3 replication and pathogenesis and can be used complementarily with PCR for evaluating the localization of antigens in infected tissues. |
| first_indexed | 2025-11-15T13:55:26Z |
| format | Article |
| id | upm-107025 |
| institution | Universiti Putra Malaysia |
| institution_category | Local University |
| language | English |
| last_indexed | 2025-11-15T13:55:26Z |
| publishDate | 2023 |
| publisher | Veterinary World |
| recordtype | eprints |
| repository_type | Digital Repository |
| spelling | upm-1070252024-10-17T06:48:34Z http://psasir.upm.edu.my/id/eprint/107025/ Chromogenic in-situ hybridization technique for the detection of porcine circovirus 3 in formalin-fixed paraffin-embedded lung and lymphoid tissues Tan, Chew Yee Lee, Kah Chun Chiou, Ming-Tang Lin, Chao-Nan Ooi, Peck Toung Background and Aim: Porcine circovirus 3 (PCV3) was recently reported in Malaysian commercial pig population in 2020 by conventional polymerase chain reaction (PCR), revealing a molecular prevalence of 17.02 in the sampled domestic pig population. This study aims to describe a chromogenic in situ hybridization (ISH) technique using digoxigenin (DIG)- labeled cloned PCV3 open reading frame 1 (ORF1) fragment DNA to detect and localize the PCV3 antigen in formalinfixed, paraffin-embedded lung, and lymphoid tissue specimens. Materials and Methods: Since PCV3 was mainly detected in lung and lymphoid tissues, we obtained tissue specimens from these organs from the previous Malaysian PCV3 study. Digoxigenin-labeled ISH probes were designed to target a 69 bp region of PCV3 ORF1 spanning from the nucleotide positions (282“350). Results: Light microscopy analysis revealed that chromogenic staining of PCV3 antigens was visualized within the cytoplasm of pneumocytes and lymphocytes, indicating positive ISH results. The results of molecular detection of PCV3 using PCR and ISH showed a high agreement of 90.91, including for the negative PCV3 status for all samples. Conclusion: This study reports a chromogenic ISH technique using DIG-labeled probes targeting PCV3 ORF1 to detect PCV3 antigens in lung and lymphoid tissues. Despite the limited availability of PCV3 antibodies, ISH remains relevant for investigating PCV3 replication and pathogenesis and can be used complementarily with PCR for evaluating the localization of antigens in infected tissues. Veterinary World 2023-07-09 Article PeerReviewed text en http://psasir.upm.edu.my/id/eprint/107025/1/Chromogenic%20in-situ%20hybridization%20technique%20for%20the%20detection%20of%20porcine%20circovirus%203%20in%20formalin-fixed%20paraffin-embedded%20lung%20and%20lymphoid%20tissues.pdf Tan, Chew Yee and Lee, Kah Chun and Chiou, Ming-Tang and Lin, Chao-Nan and Ooi, Peck Toung (2023) Chromogenic in-situ hybridization technique for the detection of porcine circovirus 3 in formalin-fixed paraffin-embedded lung and lymphoid tissues. Veterinary World, 16 (7). 1444 - 1450. ISSN 0972-8988; ESSN: 2231-0916 https://www.veterinaryworld.org/Vol.16/July-2023/10.html 10.14202/vetworld.2023.1444-1450 |
| spellingShingle | Tan, Chew Yee Lee, Kah Chun Chiou, Ming-Tang Lin, Chao-Nan Ooi, Peck Toung Chromogenic in-situ hybridization technique for the detection of porcine circovirus 3 in formalin-fixed paraffin-embedded lung and lymphoid tissues |
| title | Chromogenic in-situ hybridization technique for the detection of porcine circovirus 3 in formalin-fixed paraffin-embedded lung and lymphoid tissues |
| title_full | Chromogenic in-situ hybridization technique for the detection of porcine circovirus 3 in formalin-fixed paraffin-embedded lung and lymphoid tissues |
| title_fullStr | Chromogenic in-situ hybridization technique for the detection of porcine circovirus 3 in formalin-fixed paraffin-embedded lung and lymphoid tissues |
| title_full_unstemmed | Chromogenic in-situ hybridization technique for the detection of porcine circovirus 3 in formalin-fixed paraffin-embedded lung and lymphoid tissues |
| title_short | Chromogenic in-situ hybridization technique for the detection of porcine circovirus 3 in formalin-fixed paraffin-embedded lung and lymphoid tissues |
| title_sort | chromogenic in-situ hybridization technique for the detection of porcine circovirus 3 in formalin-fixed paraffin-embedded lung and lymphoid tissues |
| url | http://psasir.upm.edu.my/id/eprint/107025/ http://psasir.upm.edu.my/id/eprint/107025/ http://psasir.upm.edu.my/id/eprint/107025/ http://psasir.upm.edu.my/id/eprint/107025/1/Chromogenic%20in-situ%20hybridization%20technique%20for%20the%20detection%20of%20porcine%20circovirus%203%20in%20formalin-fixed%20paraffin-embedded%20lung%20and%20lymphoid%20tissues.pdf |