JPWO2007032241A1 - 微粒子−タンパク質複合体およびその作製方法、半導体装置、蛍光標識方法 - Google Patents
微粒子−タンパク質複合体およびその作製方法、半導体装置、蛍光標識方法 Download PDFInfo
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- JPWO2007032241A1 JPWO2007032241A1 JP2007535431A JP2007535431A JPWO2007032241A1 JP WO2007032241 A1 JPWO2007032241 A1 JP WO2007032241A1 JP 2007535431 A JP2007535431 A JP 2007535431A JP 2007535431 A JP2007535431 A JP 2007535431A JP WO2007032241 A1 JPWO2007032241 A1 JP WO2007032241A1
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Classifications
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- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
- C07K14/47—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B82—NANOTECHNOLOGY
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- C09K11/00—Luminescent, e.g. electroluminescent, chemiluminescent materials
- C09K11/02—Use of particular materials as binders, particle coatings or suspension media therefor
- C09K11/025—Use of particular materials as binders, particle coatings or suspension media therefor non-luminescent particle coatings or suspension media
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- C09K11/00—Luminescent, e.g. electroluminescent, chemiluminescent materials
- C09K11/08—Luminescent, e.g. electroluminescent, chemiluminescent materials containing inorganic luminescent materials
- C09K11/56—Luminescent, e.g. electroluminescent, chemiluminescent materials containing inorganic luminescent materials containing sulfur
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
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- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
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- G01N33/531—Production of immunochemical test materials
- G01N33/532—Production of labelled immunochemicals
- G01N33/533—Production of labelled immunochemicals with fluorescent label
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- H—ELECTRICITY
- H01—ELECTRIC ELEMENTS
- H01L—SEMICONDUCTOR DEVICES NOT COVERED BY CLASS H10
- H01L29/00—Semiconductor devices specially adapted for rectifying, amplifying, oscillating or switching and having potential barriers; Capacitors or resistors having potential barriers, e.g. a PN-junction depletion layer or carrier concentration layer; Details of semiconductor bodies or of electrodes thereof ; Multistep manufacturing processes therefor
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- H01L29/423—Electrodes ; Multistep manufacturing processes therefor characterised by their shape, relative sizes or dispositions not carrying the current to be rectified, amplified or switched
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- H01L29/42316—Gate electrodes for field effect devices for field-effect transistors
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- H01L29/66—Types of semiconductor device ; Multistep manufacturing processes therefor
- H01L29/68—Types of semiconductor device ; Multistep manufacturing processes therefor controllable by only the electric current supplied, or only the electric potential applied, to an electrode which does not carry the current to be rectified, amplified or switched
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- Inorganic Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Crystallography & Structural Chemistry (AREA)
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Abstract
Description
2、7、12 外殻
3、9、13 モノマーサブユニット
101 半導体基板
102 素子分離用絶縁膜
103 トンネル絶縁膜
103a、105a SiO2膜
104 微粒子
105 絶縁膜
106 コントロールゲート
107a ソース領域
107b ドレイン領域
108 層間絶縁膜
109 コンタクトホール
110 タングステンプラグ
111a、111b Al配線
120 半導体装置
130 チャネル
132 アポフェリチン
135 硫化物イオン(金属イオン)
137 微粒子−アポフェリチン複合体
図2(a)は、本発明の第1の実施形態に係る化合物半導体−アポフェリチン複合体の作製方法を示すフローチャートであり、(b)は、各試薬の最終濃度を示す図である。本実施形態では、硫化カドミウム(CdS)からなる微粒子をアポフェリチン内に形成させる方法を説明する。
図9(a)は、本発明の第2の実施形態に係る化合物半導体−アポフェリチン複合体の作製方法を示すフローチャートであり、(b)は、各試薬の最終濃度を示す図である。本実施形態では、硫化亜鉛(ZnS)からなる微粒子をアポフェリチン内に形成させる方法を説明する。
図12(a)は、本発明の第3の実施形態に係る化合物半導体−リステリアアポフェリチン複合体の作製方法を示すフローチャートであり、(b)は、各試薬の最終濃度を示す図である。本実施形態では、CdSからなる微粒子をリステリアアポフェリチン内に形成させる方法を説明する。
図12(c)は、第3の実施形態の変形例に係る化合物半導体−リステリアアポフェリチン複合体の作製条件を示す図である。
図14(a)は、本発明の第4の実施形態に係る化合物半導体−リステリアアポフェリチン複合体の作製方法を示すフローチャートであり、(b)は、各試薬の最終濃度を示す図である。本実施形態では、硫化亜鉛(ZnS)からなる微粒子をリステリアアポフェリチン内に形成させる方法を説明する。
図14(c)は、第3の実施形態の変形例に係る化合物半導体−リステリアアポフェリチン複合体の作製条件を示す図である。
本発明の第5の実施形態として、遺伝子組み換え技術を用いて作製されたリコンビナントアポフェリチンを用いて化合物半導体からなる微粒子−タンパク質複合体を形成させる方法を説明する。
これ以降の実施形態では、化合物半導体−アポフェリチン(またはリステリアフェリチン)複合体の応用例について説明する。
図18は、本発明の第6の実施形態に係る半導体装置を示す断面図である。同図に示す半導体装置は、化合物半導体からなる量子ドットを利用した不揮発性メモリセルである。
図19(a)〜(f)は、本実施形態の半導体装置の製造方法を説明するための断面図である。
本発明の第7の実施形態として、本発明の化合物半導体−タンパク質複合体を蛍光標識として用いる方法について説明する。
図20(a)〜(g)は、本実施形態の第1の蛍光標識方法を説明するための図である。ここでは、化合物半導体−タンパク質複合体を用いて標的物質を蛍光標識する方法を説明する。
図21(a)、(b)は、本実施形態の第2の蛍光標識方法を説明するための図である。
本発明の第8の実施形態として、基板に対して蛍光標識を行う方法について説明する。
図23は、本発明の第9の実施形態に係る化合物半導体−アポフェリチン複合体の作成方法を示すフローチャートである。本実施形態では、CdSからなる微粒子をアポフェリチン内に形成させる方法を説明する。
Claims (18)
- 複数のモノマーサブユニットから構成され、内部に空洞が形成された外殻と、
前記外殻内に形成され、化合物半導体からなり、励起された場合に蛍光を発する微粒子とを備え、
前記外殻は、N末端の一部を欠失させた単一のモノマーサブユニットからなるリコンビナントアポフェリチンまたは、N末端の一部を欠失させた単一のモノマーサブユニットからなるリコンビナントリステリアアポフェリチンである、微粒子−タンパク質複合体。 - 前記微粒子は、CdSまたはZnSを含むことを特徴とする請求項1に記載の微粒子−タンパク質複合体。
- 水溶液中で金属のアンモニウム錯体を形成させる工程(a)と、
内部に空洞が形成されたタンパク質の溶液とチオ酢酸とを前記水溶液中で混合して、前記タンパク質の前記空洞内に前記金属の硫化物からなる微粒子を形成させる工程(b)とを備えていることを特徴とする微粒子−タンパク質複合体の作製方法。 - 前記タンパク質はアポフェリチン、リステリアアポフェリチンまたはこれらの組み換え体であることを特徴とする請求項3に記載の微粒子−タンパク質複合体の作製方法。
- 前記金属は、CdまたはZnであることを特徴とする請求項3に記載の微粒子−タンパク質複合体の作製方法。
- 前記工程(a)では、酢酸イオンとアンモニウムイオンとを含む溶液と、前記金属の酢酸塩溶液と、アンモニア水とを混合することを特徴とする請求項5に記載の微粒子−タンパク質複合体の作製方法。
- 前記金属はCdまたはZnであり、
前記工程(b)では、前記水溶液中のアンモニア濃度によって直径の異なる微粒子が形成されることを特徴とする請求項6に記載の微粒子−タンパク質複合体の作製方法。 - 前記タンパク質はリステリアアポフェリチンまたはリステリアアポフェリチンの組み換え体であり、
前記工程(a)および前記工程(b)は、共に0℃以上15℃以下で行うことを特徴とする請求項3に記載の微粒子−タンパク質複合体の作製方法。 - 前記工程(b)で形成された前記微粒子と前記タンパク質との複合体を精製する工程(c)と、
前記工程(c)で精製された前記微粒子と前記タンパク質との複合体を含む溶液と前記金属を含む溶液とを混合する工程(d)と、
前記工程(d)で作製された混合液にチオ酢酸を加え、前記タンパク質内に形成された前記微粒子を成長させる工程(e)と、
前記工程(e)の後、前記混合液に前記金属を含む溶液と前記チオ酢酸とを交互に加える工程を繰り返して前記微粒子を成長させる工程(f)とをさらに備えていることを特徴とする請求項3に記載の微粒子−タンパク質複合体の作製方法。 - 内部に空洞が形成されたタンパク質と、前記空洞内に形成された金属の硫化物からなる微粒子との複合体を含む溶液を作製する工程(a)と、
前記微粒子と前記タンパク質との複合体を含む溶液に前記金属を含む溶液を加える工程(b)と、
前記工程(b)の後に、前記微粒子と前記タンパク質との複合体を含む溶液にチオ酢酸を加えて前記微粒子を成長させる工程(c)と、
前記工程(b)と前記工程(c)とを繰り返して前記微粒子を成長させる工程(d)とを備えている微粒子−タンパク質複合体の作製方法。 - 水溶液中でCdまたはZnのアンモニウム錯体を形成させる工程(a)と、
内部に空洞が形成されたタンパク質の溶液とチオ酢酸とを前記水溶液中で混合して、前記空洞内にCdSまたはZnSを含み、励起された場合に蛍光を発する微粒子を形成させ、微粒子−タンパク質複合体を作製する工程(b)と、
前記微粒子−タンパク質複合体を標的物質に結合させて前記標的物質を標識する工程(c)とを備えていることを特徴とする蛍光標識方法。 - 内部に空洞を有する前記タンパク質はアポフェリチン、リステリアアポフェリチンまたはこれらの組み換え体であることを特徴とする請求項11に記載の蛍光標識方法。
- 前記工程(c)の前に、N末端にシステインを付加した第1の変異アポフェリチンまたは第1の変異リステリアアポフェリチンを作製する工程(d)と、
前記工程(c)の前に、前記標的物質をSH基を有する化合物で修飾する工程(e)とをさらに備え、
前記工程(c)では、前記第1の変異アポフェリチンまたは前記第1の変異リステリアアポフェリチンと前記標的物質とをジスルフィド結合によって結合させることを特徴とする請求項12に記載の蛍光標識方法。 - 前記標的物質は外表面にアミノ基を有しており、
前記工程(c)では、カップリング剤を用いて前記標的物質の外表面に位置するアミノ基と前記アポフェリチンまたは前記リステリアアポフェリチンの外表面に位置するカルボキシル基とを縮合反応させてアミド結合を形成させることを特徴とする請求項12に記載の蛍光標識方法。 - 前記タンパク質は、N末端にTi認識ペプチドが付加された第2の変異アポフェリチンであり、
前記標的物質は表面の一部にTi膜を有する基板であることを特徴とする請求項12に記載の蛍光標識方法。 - 前記工程(a)の前に、遺伝子組み換え技術を用いて前記第2の変異アポフェリチンを作製する工程をさらに備えていることを特徴とする請求項15に記載の蛍光標識方法。
- 水溶液中でCdまたはZnのアンモニウム錯体を形成させる工程(a)と、
内部に空洞が形成されたタンパク質の溶液とチオ酢酸とを前記水溶液中で混合して、前記空洞内にCdSまたはZnSを含み、励起された場合に蛍光を発する微粒子を形成させ、微粒子−タンパク質複合体を作製する工程(b)と、
内部に空洞を有するタンパク質を標的物質に結合させる工程(c)と、
前記タンパク質内にCdSまたはZnSを含み、励起された場合に蛍光を発する微粒子を形成させ、微粒子−タンパク質複合体を作製する工程(d)とを備えていることを特徴とする蛍光標識方法。 - 内部に空洞を有する前記タンパク質はアポフェリチン、リステリアアポフェリチンまたはこれらの組み換え体であることを特徴とする請求項17に記載の蛍光標識方法。
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