JP5306943B2 - 弱毒ウイルスの作出方法 - Google Patents
弱毒ウイルスの作出方法 Download PDFInfo
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- JP5306943B2 JP5306943B2 JP2009192750A JP2009192750A JP5306943B2 JP 5306943 B2 JP5306943 B2 JP 5306943B2 JP 2009192750 A JP2009192750 A JP 2009192750A JP 2009192750 A JP2009192750 A JP 2009192750A JP 5306943 B2 JP5306943 B2 JP 5306943B2
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- 241000700605 Viruses Species 0.000 title claims description 112
- 230000002238 attenuated effect Effects 0.000 title claims description 25
- 210000004027 cell Anatomy 0.000 claims description 89
- 238000000034 method Methods 0.000 claims description 33
- 210000003501 vero cell Anatomy 0.000 claims description 19
- 238000012258 culturing Methods 0.000 claims description 4
- 238000004519 manufacturing process Methods 0.000 claims description 4
- 241001135989 Porcine reproductive and respiratory syndrome virus Species 0.000 claims description 2
- 230000000120 cytopathologic effect Effects 0.000 description 22
- 238000011081 inoculation Methods 0.000 description 18
- 238000012360 testing method Methods 0.000 description 12
- 210000001132 alveolar macrophage Anatomy 0.000 description 9
- 241001465754 Metazoa Species 0.000 description 8
- 241000282887 Suidae Species 0.000 description 8
- 238000011084 recovery Methods 0.000 description 8
- 229960005486 vaccine Drugs 0.000 description 7
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- 208000024891 symptom Diseases 0.000 description 4
- 241000282898 Sus scrofa Species 0.000 description 3
- 210000004748 cultured cell Anatomy 0.000 description 3
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 3
- 208000015181 infectious disease Diseases 0.000 description 3
- 238000004264 monolayer culture Methods 0.000 description 3
- 238000012770 revaccination Methods 0.000 description 3
- 208000005342 Porcine Reproductive and Respiratory Syndrome Diseases 0.000 description 2
- 206010037660 Pyrexia Diseases 0.000 description 2
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- 206010000234 Abortion spontaneous Diseases 0.000 description 1
- 241000282693 Cercopithecidae Species 0.000 description 1
- 241000282552 Chlorocebus aethiops Species 0.000 description 1
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- 241000144290 Sigmodon hispidus Species 0.000 description 1
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- 238000007918 intramuscular administration Methods 0.000 description 1
- 210000003292 kidney cell Anatomy 0.000 description 1
- 231100001231 less toxic Toxicity 0.000 description 1
- 229940124590 live attenuated vaccine Drugs 0.000 description 1
- 229940023012 live-attenuated vaccine Drugs 0.000 description 1
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- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
Description
野外飼育豚から、以下に示す通りに独自に豚繁殖・呼吸障害症候群(PRRS)ウイルスを分離した。ウイルスゲノム配列の一部を解析した結果から、当該分離株は北米型PRRSウイルスであることが判明した。
MARC145細胞で培養したウイルス液(回収培養液)をベロ細胞(理研セルバンクより入手)に接種し、37℃で5〜7日間培養した。ベロ細胞はMEM培地中で培養し、単層培養に調整して接種に用いた。接種後、CPEは確認されなかったが、抗血清を用いた間接蛍光抗体法により細胞内のウイルスの存在は確認できた。
(1) 各種細胞への感受性試験
予備試験として、上記で得たウイルス株のうち33代目(PRRSVwt-7/Vero-33T)について、豚肺胞マクロファージへの感受性を確認した。コントロールのウイルス株として、MARC145細胞で7代及び27代継代(ウイルス回収・再接種)した株を用いた。回収培養液(ウイルス液)を豚肺胞マクロファージ(Mφ)、MARC145細胞、ベロ細胞に接種し、各ウイルスの力価を算出して比較した。試験に用いたウイルス株と試験の結果をそれぞれ表1及び表2に示す。
下記表3に示す通り、4週齢のPRRS陰性SPF豚5頭に1頭当たり106.8TCID50のウイルス液を筋肉内接種し、4週間観察した。比較のため、PRRSVwt-7/MARC-7T(比較例)及び市販の弱毒生ワクチン製剤(参考例)を接種した。市販のワクチンは販売者が推奨する用法・用量通りに用いた。
Claims (2)
- ウイルスが感染した株化細胞を培養し、次いで、ウイルス非感染細胞へのウイルスの再接種をすることなく、感染細胞を継代培養する工程を含む、弱毒ウイルスの作出方法であって、前記ウイルスが豚繁殖・呼吸障害症候群ウイルスであり、前記細胞がベロ細胞である、方法。
- 前記工程が15〜35回行われる請求項1記載の方法。
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JP2009192750A JP5306943B2 (ja) | 2009-08-24 | 2009-08-24 | 弱毒ウイルスの作出方法 |
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JP2009192750A JP5306943B2 (ja) | 2009-08-24 | 2009-08-24 | 弱毒ウイルスの作出方法 |
Publications (2)
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JP2011041534A JP2011041534A (ja) | 2011-03-03 |
JP5306943B2 true JP5306943B2 (ja) | 2013-10-02 |
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JP2009192750A Active JP5306943B2 (ja) | 2009-08-24 | 2009-08-24 | 弱毒ウイルスの作出方法 |
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Families Citing this family (2)
Publication number | Priority date | Publication date | Assignee | Title |
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US20210277361A1 (en) * | 2018-08-27 | 2021-09-09 | Mican Technologies Inc. | Method for evaluating anti-infective drugs, vaccines, etc. using immortalized monocytic cells and induced cells |
JP7079901B1 (ja) * | 2021-08-24 | 2022-06-02 | Vaxosome合同会社 | 細胞外粒子の産生方法 |
Family Cites Families (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CA2076744C (en) * | 1991-08-26 | 2000-06-27 | Danny W. Chladek | Viral agent associated with mystery swine disease |
US6042830A (en) * | 1992-08-05 | 2000-03-28 | Boehringer Ingelheim Vetmedica, Inc. | Viral agent associated with mystery swine disease |
DK0835930T3 (da) * | 1996-10-09 | 2001-06-18 | Akzo Nobel Nv | Europæiske vaccinestammer af porcint reproduktions- og respirationssyndrom-virus. (PRRSV) |
CA2370372C (en) * | 1999-04-22 | 2009-02-24 | United States Department Of Agriculture | Porcine reproductive and respiratory syndrome vaccine, based on isolate ja-142 |
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