Immunohistochemical detection of enteroviruses in pancreatic tissues of patients with type 1 diabetes using a polyclonal antibody against 2A protease of Coxsackievirus
ABSTRACT Aims/Introduction The need for antiserum for immunohistochemical (IHC) detection of enterovirus (EV) in formaldehyde‐fixed and paraffin‐embedded samples is increasing. The gold standard monoclonal antibody (clone 5D8/1) against EV‐envelope protein (VP1) was proven to cross‐react with other...
| Main Authors: | , , , , , , |
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| Format: | Article |
| Language: | English |
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Wiley
2022-03-01
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| Series: | Journal of Diabetes Investigation |
| Subjects: | |
| Online Access: | https://doi.org/10.1111/jdi.13700 |
| _version_ | 1819121612173082624 |
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| author | Erika Jimbo Tetsuro Kobayashi Akira Takeshita Keiichiro Mine Seiho Nagafuchi Tomoyasu Fukui Soroku Yagihashi |
| author_facet | Erika Jimbo Tetsuro Kobayashi Akira Takeshita Keiichiro Mine Seiho Nagafuchi Tomoyasu Fukui Soroku Yagihashi |
| author_sort | Erika Jimbo |
| collection | DOAJ |
| description | ABSTRACT Aims/Introduction The need for antiserum for immunohistochemical (IHC) detection of enterovirus (EV) in formaldehyde‐fixed and paraffin‐embedded samples is increasing. The gold standard monoclonal antibody (clone 5D8/1) against EV‐envelope protein (VP1) was proven to cross‐react with other proteins. Another candidate marker of EV proteins is 2A protease (2Apro), which is encoded by the EV gene and translated by the host cells during EV replication, and participates processing proproteins to viral capsid proteins. Materials and Methods We raised polyclonal antiserum by immunizing a rabbit with an 18‐mer peptide of Coxsackievirus B1 (CVB1)‐2Apro, and examined the specificity and sensitivity for EV on formaldehyde‐fixed and paraffin‐embedded tissue samples. Results Enzyme‐linked immunosorbent assay study showed a high titer of antibody for 18‐mer peptide of CVB1‐2Apro, cross‐reacting with CVB3‐2Apro peptide. IHC showed that antiserum against 2Apro reacted with CVB1‐infected and VP1‐positive Vero cells. Confocal laser scanning microscopy showed that antigen stained by the 2Apro antibody located in the same cell with VP1 stained by 5D8/1. IHC using 2Apro antiserum showed dense staining in the islets of EV‐associated fulminant type 1 diabetes pancreas and that located in the same cell stained positive for VP1 (5D8/1). Specificity of 2Apro antiserum by IHC staining was confirmed by negative 2Apro in 14 VP1‐negative non‐diabetes control pancreases. Conclusions Our study provides a new polyclonal antiserum against CVB1‐2Apro, which might be useful for IHC of EV‐infected human tissues stored as archive of formaldehyde‐fixed and paraffin‐embedded tissue samples. |
| first_indexed | 2024-12-22T06:39:19Z |
| format | Article |
| id | doaj.art-516a8a33dc2f44948c0555f7543a908f |
| institution | Directory Open Access Journal |
| issn | 2040-1116 2040-1124 |
| language | English |
| last_indexed | 2024-12-22T06:39:19Z |
| publishDate | 2022-03-01 |
| publisher | Wiley |
| record_format | Article |
| series | Journal of Diabetes Investigation |
| spelling | doaj.art-516a8a33dc2f44948c0555f7543a908f2022-12-21T18:35:28ZengWileyJournal of Diabetes Investigation2040-11162040-11242022-03-0113343544210.1111/jdi.13700Immunohistochemical detection of enteroviruses in pancreatic tissues of patients with type 1 diabetes using a polyclonal antibody against 2A protease of CoxsackievirusErika Jimbo0Tetsuro Kobayashi1Akira Takeshita2Keiichiro Mine3Seiho Nagafuchi4Tomoyasu Fukui5Soroku Yagihashi6Division of Immunology and Molecular Medicine Okinaka Memorial Institute for Medical Research Tokyo JapanDivision of Immunology and Molecular Medicine Okinaka Memorial Institute for Medical Research Tokyo JapanDepartment of Endocrinology and Metabolism Toranomon Hospital Tokyo JapanDivision of Metabolism and Endocrinology Department of Internal Medicine Faculty of Medicine Saga University Saga JapanDivision of Metabolism and Endocrinology Department of Internal Medicine Faculty of Medicine Saga University Saga JapanDivision of Diabetes, Metabolism and Endocrinology Department of Medicine Showa University School of Medicine Tokyo JapanDepartment of Exploratory Medicine on Nature, Life, and Man Toho University School of Medicine Chiba JapanABSTRACT Aims/Introduction The need for antiserum for immunohistochemical (IHC) detection of enterovirus (EV) in formaldehyde‐fixed and paraffin‐embedded samples is increasing. The gold standard monoclonal antibody (clone 5D8/1) against EV‐envelope protein (VP1) was proven to cross‐react with other proteins. Another candidate marker of EV proteins is 2A protease (2Apro), which is encoded by the EV gene and translated by the host cells during EV replication, and participates processing proproteins to viral capsid proteins. Materials and Methods We raised polyclonal antiserum by immunizing a rabbit with an 18‐mer peptide of Coxsackievirus B1 (CVB1)‐2Apro, and examined the specificity and sensitivity for EV on formaldehyde‐fixed and paraffin‐embedded tissue samples. Results Enzyme‐linked immunosorbent assay study showed a high titer of antibody for 18‐mer peptide of CVB1‐2Apro, cross‐reacting with CVB3‐2Apro peptide. IHC showed that antiserum against 2Apro reacted with CVB1‐infected and VP1‐positive Vero cells. Confocal laser scanning microscopy showed that antigen stained by the 2Apro antibody located in the same cell with VP1 stained by 5D8/1. IHC using 2Apro antiserum showed dense staining in the islets of EV‐associated fulminant type 1 diabetes pancreas and that located in the same cell stained positive for VP1 (5D8/1). Specificity of 2Apro antiserum by IHC staining was confirmed by negative 2Apro in 14 VP1‐negative non‐diabetes control pancreases. Conclusions Our study provides a new polyclonal antiserum against CVB1‐2Apro, which might be useful for IHC of EV‐infected human tissues stored as archive of formaldehyde‐fixed and paraffin‐embedded tissue samples.https://doi.org/10.1111/jdi.137002A proteaseEnterovirusPolyclonal antiserum |
| spellingShingle | Erika Jimbo Tetsuro Kobayashi Akira Takeshita Keiichiro Mine Seiho Nagafuchi Tomoyasu Fukui Soroku Yagihashi Immunohistochemical detection of enteroviruses in pancreatic tissues of patients with type 1 diabetes using a polyclonal antibody against 2A protease of Coxsackievirus Journal of Diabetes Investigation 2A protease Enterovirus Polyclonal antiserum |
| title | Immunohistochemical detection of enteroviruses in pancreatic tissues of patients with type 1 diabetes using a polyclonal antibody against 2A protease of Coxsackievirus |
| title_full | Immunohistochemical detection of enteroviruses in pancreatic tissues of patients with type 1 diabetes using a polyclonal antibody against 2A protease of Coxsackievirus |
| title_fullStr | Immunohistochemical detection of enteroviruses in pancreatic tissues of patients with type 1 diabetes using a polyclonal antibody against 2A protease of Coxsackievirus |
| title_full_unstemmed | Immunohistochemical detection of enteroviruses in pancreatic tissues of patients with type 1 diabetes using a polyclonal antibody against 2A protease of Coxsackievirus |
| title_short | Immunohistochemical detection of enteroviruses in pancreatic tissues of patients with type 1 diabetes using a polyclonal antibody against 2A protease of Coxsackievirus |
| title_sort | immunohistochemical detection of enteroviruses in pancreatic tissues of patients with type 1 diabetes using a polyclonal antibody against 2a protease of coxsackievirus |
| topic | 2A protease Enterovirus Polyclonal antiserum |
| url | https://doi.org/10.1111/jdi.13700 |
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