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METHOD FOR OBTAINING INFORMATION OF ANTIGEN-SPECIFIC ANTIBODY

Foreign code F190009687
File No. S2017-0535-C0
Posted date Jan 23, 2019
Country WIPO
International application number 2018JP015671
International publication number WO 2018194015
Date of international filing Apr 16, 2018
Date of international publication Oct 25, 2018
Priority data
  • P2017-084431 (Apr 21, 2017) JP
Title METHOD FOR OBTAINING INFORMATION OF ANTIGEN-SPECIFIC ANTIBODY
Abstract Provided is a method whereby a high-affinity antibody can be efficiently produced with the use of a natural type protein as an antigen. Also provided is a method for obtaining information of an antigen-specific antibody derived exclusively from a blistered fish without being affected by microorganisms existing in the rearing environment. Information of a high-affinity antibody, i.e., an antigen-specific antibody is obtained with the use of a natural type protein as an antigen through the following steps, etc.: (A) a step for administering an antigen to a blistered fish and allowing the fish to produce an antibody; (B) a step for collecting the blister fluid containing the antibody from the fish treated in (A); and (C) a step for constructing an antigen-specific polyclonal antibody gene library from the blister fluid collected in (B). Thus, information of the antigen-specific antibody is obtained using primers, said primers being capable of amplifying only the gene of the antigen-specific antibody derived exclusively from the blistered fish but not amplifying genes of microorganisms existing in the rearing environment such as Propionibacterium acnes .
Outline of related art and contending technology BACKGROUND ART
In the prior art, a variety of mammalian immune by an animal such as a mouse or a variety of antibodies such as phage display method is the method has been developed. However, these methods having a high affinity for the target protein of a naturally occurring antibody can be acquired, and a high purity target protein, and in the three-dimensional structure while maintaining the required conditions. In addition, the antibody by the immune system DNA are known in this method, this method is a dendritic cell can be expressed as in the native part of the protein when the target is effective. However, proteins cannot be expressed in a dendritic cell, a small amount of target protein as the expression of high affinity antibody was extremely difficult to acquire. Therefore, these known in the art depending on the method for antibody production, post-translational protein, sugar chain, fatty acid or the like is modified with the native protein as an antigen, such as drug discovery can be used to high-affinity antibodies have to be made efficiently.
Therefore, the present invention an administrator of the native protein as an antigen, and to efficiently manufacture high-affinity antibodies can be object of the present invention is to provide methods and. Then, in order to provide a method, an antibody specific for an antigen by fish having bubbles of information have been acquired. Having a raised by the bubbles because the fish in water, such as acne bacteria were present in the environment without being affected by the microorganisms, having only the fish and bubbles from the antigen-specific antibody may acquire the information of the object of the present invention. In addition, the inventors of the present invention, in Patent Document 1, and fish having bubbles by the object of the challenge, while making the state of the fish, the fish antibodies produced can be repeatedly obtained to develop a method of manufacturing an antibody.
Scope of claims (In Japanese)請求の範囲 [請求項1]
次の(A)~(D)の工程を含む抗原特異的な抗体の情報を取得する方法。
(A)水泡を有する魚類に抗原を投与し、抗体を産生させる工程
(B)上記(A)の魚類から抗体を含む水泡液を採取する工程
(C)上記(B)にて採取した水泡液より抗原特異的なポリクローナル抗体遺伝子ライブラリーを構築する工程
(D)上記(C)にて構築された抗原特異的なポリクローナル抗体遺伝子ライブラリーをファージディスプレイ化する工程

[請求項2]
抗原特異的な抗体の情報が抗原特異的なVH領域および/またはVL領域の塩基配列情報および/またはアミノ酸配列情報である請求項1に記載の方法。

[請求項3]
抗原特異的なポリクローナル抗体遺伝子ライブラリーの構築にあたり、配列表配列番号1に記載の塩基配列を含むプライマーおよび/または配列表配列番号2に記載の塩基配列を含むプライマーを用いる請求項1または2に記載の方法。

[請求項4]
水泡を有する魚類が水泡眼またはらんちゅうである請求項1~3のいずれかに記載の方法。

[請求項5]
請求項1~4のいずれかに記載の方法により取得される抗原特異的な抗体の塩基配列情報。

[請求項6]
請求項1~4のいずれかに記載の方法により取得される抗原特異的な抗体のアミノ酸配列情報。

[請求項7]
次の(A)~(E)の工程を経て得られる、抗原特異的なポリクローナル抗体の全部または一部が示されたファージ提示組換えタンパク質。
(A)水泡を有する魚類に抗原を投与し、抗体を産生させる工程
(B)上記(A)の魚類から抗体を含む水泡液を採取する工程
(C)上記(B)にて採取した水泡液より抗原特異的なポリクローナル抗体遺伝子ライブラリーを構築する工程
(D)上記(C)にて構築された抗原特異的なポリクローナル抗体遺伝子ライブラリーをファージディスプレイ化する工程
(E)上記(D)の工程により構築されたファージライブラリから抗原に結合する抗原特異的なポリクローナル抗体の全部または一部が示されたファージ提示組換えタンパク質をスクリーニングする工程

[請求項8]
配列表配列番号31~34のいずれかに記載の塩基配列によってコードされる、または配列表配列番号35~38のいずれかに記載のアミノ酸配列によって示される請求項7に記載のファージ提示組換えタンパク質。

[請求項9]
配列表配列番号1に記載の塩基配列を含むプライマーおよび/または配列表配列番号2に記載の塩基配列を含むプライマー。

[請求項10]
抗原特異的なポリクローナル抗体遺伝子ライブラリーの構築のための請求項9に記載のプライマー。

  • Applicant
  • ※All designated countries except for US in the data before July 2012
  • MIE UNIVERSITY
  • Inventor
  • TAMARU Yutaka
IPC(International Patent Classification)
Specified countries National States: AE AG AL AM AO AT AU AZ BA BB BG BH BN BR BW BY BZ CA CH CL CN CO CR CU CZ DE DJ DK DM DO DZ EC EE EG ES FI GB GD GE GH GM GT HN HR HU ID IL IN IR IS JO JP KE KG KH KN KP KR KW KZ LA LC LK LR LS LU LY MA MD ME MG MK MN MW MX MY MZ NA NG NI NO NZ OM PA PE PG PH PL PT QA RO RS RU RW SA SC SD SE SG SK SL SM ST SV SY TH TJ TM TN TR TT TZ UA UG US UZ VC VN ZA ZM ZW
ARIPO: BW GH GM KE LR LS MW MZ NA RW SD SL SZ TZ UG ZM ZW
EAPO: AM AZ BY KG KZ RU TJ TM
EPO: AL AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HR HU IE IS IT LT LU LV MC MK MT NL NO PL PT RO RS SE SI SK SM TR
OAPI: BF BJ CF CG CI CM GA GN GQ GW KM ML MR NE SN ST TD TG
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