In vitro Biotinylation of Avitag with Enzyme
| Avidity 主要開發及販售與分子結合相關的產品。 其專利的 AviTag™ 是可以被 BirA 酵素進行生物素鍵結反應 (Biotinylation) 的胜肽鏈 (peptide)，結合 Biotin 可與 avidin 或 streptavidin 專一性鍵結的特性，提供了理想的分子交互作用分析工具。其產品包含了in vivo, in vitro biotinylation 產品，含 AviTaq 的殖體 (vector)、Biotinylated protein、純化辨識 Avitag 的抗體…等。 |
| https://www.avidity.com/ |
| 主力產品: BirA biotin-protein ligase bulk reaction kit |
| The BirA biotin-protein ligase adds d-biotin covalently to biotin-acceptor peptides/proteins via an ATP intermediate (biotinyl 5′-adenylate) in a highly efficient and targeted manner. This kit includes 300 µg of BirA enzyme for those who require larger scale reactions or more frequent use of the BirA biotinylation reaction in their work. Please Note: We strongly recommend the use of the AviTag amino acid sequence over other Biotinylation of Peptides sequences. The BirA enzyme biotinylates the AviTag sequence at a reaction rate 2x the natural substrate (BCCP) and as much as an order of magnitude or more over the other peptide sequences in use. The AviTag™ requires smaller amounts of enzyme and shorter incubation times than other Biotinylation of Peptides sequences thus minimizing ancillary problems associated with proteases and protein instability. |
| Cell lines- in vivo biotinylationAVB101AVB101- glycerol stock CVB101- chemically competent cellsEVB101- electro-competent cellsAVB100 AVB100- glycerol stockEVB100- electro-competent cellsAVB99AVB99-glycerol stockBIP-300- positive control plasmid for induction Enzyme- in vitro biotinylationBiotin ligaseBirA500- 40ug biotin ligaseBulk BirA- 300 ug biotin ligaseLarge quantities of biotin ligaseBIS-300 positive and negative control protein kitBIO-200 biotin solutionBIO-500 biotin solution Vectors containing AviTagC-terminus AvitagpAC4AC5pAC6All threeSequencing primersN-terminus AvitagpAN4pAN5pAN6All threeSequencing primers Purified Biotinylated Proteins and support chemicalsGaussia princeps LuciferaseGluc-10Gluc-100Renilla reniformis Green Fluorescent ProteinStreptavidin-conjugated proteinsStreptavidin-Gaussia princeps LuciferaseStreptavidin-Renilla reniformis GFPCoelenterazine Purification and Identification of Avitag’d proteinsAbC- antibody against C-terminus AvitagAbCA -Abc antibody conjugated to agaroseBIS-300 positive and negative control protein kit |
產品
BirA Enzyme and buffers for in vitro biotinylation
BirA500: BirA biotin-protein ligase standard reaction kit
BirA500-RT: BirA biotin-protein ligase lyophilized reaction kit
Bulk BirA: BirA biotin-protein ligase bulk reaction kit
Bulk BirA-RT: BirA biotin-protein ligase bulk lyophilized reaction kit
BIS-300 positive and negative control protein kit
Cell lines for in vivo biotinylation
AVB101 – E.coli B strain for efficient in vivo biotinylation
CVB101- Chemically competent cells
EVB101- Electro Competent E. coli B strain for in vivo biotinylation
BIP-300 – positive control plasmid for induction
Avidity ATUM AviTag vectors and other plasmids
pBirAcm – plasmid for creation of BirA-overproducing E.coli cell line
| Vector product name(Order) | Promoter | N- or C- terminal AviTag | Short, Flexible linker(SF) or Rigid, Extended linker (RE) |
| p3A-T5-N-Avtg-SF | T5 | N- | SF (AviTag-flexible linker-your protein) |
| p3A-T5-N-Avtg-RE | T5 | N- | RE (AviTag-stiff linker-your protein) |
| p3A-T5-C-Avtg-SF | T5 | -C | SF (your protein-flexible linker-AviTag) |
| p3A-T5-C-Avtg-RE | T5 | -C | RE (your protein-stiff linker-AviTag) |
| p3A-Rha-N-Avtg-SF | Rha | N- | SF (AviTag-flexible linker-your protein) |
| p3A-Rha-N-Avtg-RE | Rha | N- | RE (Avitag-stiff linker-your protein) |
| p3A-Rha-C-Avtg-SF | Rha | -C | SF (your protein-flexible linker-AviTag) |
| p3A-Rha-C-Avtg-RE | Rha | -C | RE (your protein-stiff linker-AviTag) |
Purification and Identification of Avitag’d proteins
AbC- mab antibody against C-terminus AviTag
Purified Proteins and support chemicals
Biotinylated Twinkle� CFP – 100 ug
Biotinylated Holly� CFP – 100 ug
Biotinylated Dasher� CFP – 100 ug
Fluorescent Streptavidin Twinkle� Fusion Protein – 100 ug
一、核心技術平台：AviTag® / BirA® 系統
Avidity 的所有產品幾乎都圍繞「專一性生物素化技術」發展。
1. AviTag®
- 一段 15 個胺基酸的短序列，能被 BirA 專一酵素 唯一 生物素化。
- 可融合於 N 端／C 端，不影響蛋白質摺疊。
- 使用場景：蛋白純化、蛋白定量、表面固定、生物感測（Sensor）、免疫偵測等。
2. BirA® 生物素化酵素
- 專一性給 AviTag 上的 lysine 加上生物素（1:1、生理條件）。
- 兩種主要形式：
- Recombinant BirA（體外反應）
- Cellular BirA（細胞內表現）
二、產品分類
(A) AviTag® & BirA® Reagents（標記核心）
1. AviTag Peptide / Vectors
- AviTag peptide（純胜肽）
- AviTag-fusion vectors
- 各式 promoter、tag variant、linker 選項
- 可用於：細胞表達、細胞外蛋白、重組蛋白工程
(B) In Vitro Biotinylation Kits（體外生物素化）
1. BirA Biotin-Protein Ligase Kit
- 完整配方（ATP / Mg²⁺ / D-biotin / buffers）
- 適用範圍廣，支援小量或大量蛋白生產
- 高效率單位點生物素化（避免 over-biotinylation）
2. TurboBiotinylation / Enhanced biotin kits（某些版本為加速型）
- 針對高通量、快速生物素化需求
(C) Protein Purification Reagents（蛋白純化）
1. NeutrAvidin（去糖基化 Avidin）
- 低背景、少非特異性 binding
- 種類：
- NeutrAvidin resin
- NeutrAvidin agarose
- NeutrAvidin magnetic beads
2. Streptavidin 系列
- Streptavidin resin
- Streptavidin HRP
- Streptavidin–Biotin binding 系列偵測試劑
3. Biotin-Binding Capture Tools
- Columns
- Spin columns
- Magnetic beads
- High-capacity capture resin
(D) Detection Reagents（蛋白偵測）
1. Anti-AviTag 抗體
- Monoclonal antibodies
- Conjugated versions：HRP、AP、fluorophores
2. Streptavidin / NeutrAvidin 標定探針
- HRP-conjugated
- AP-conjugated
- Fluorescence (Alexa / FITC) 標定版
- 用於 Western、ELISA、流式、pull-down
3. Biotinylated Controls / Standards
- Biotinylated BSA
- Oligos
- Recombinant standards
(E) Custom Services（客製化服務）
1. Custom Biotinylation Services
- 客戶帶蛋白 → Avidity 執行單一位點生物素化（QC 保證）
- 適合需要高生物活性/特異性的診斷與製藥公司
2. Custom AviTag Fusion Protein Production
- 基因構築
- 表現
- 小量純化
- Biotinylation + QC（HPLC / MS）
三、Avidity 產品的典型應用場景
1. 生物感測（Bio-layer interferometry / SPR）
- ForteBio Octet、GE Biacore
- AviTag 蛋白 → BirA 生物素化 → 固定於 SA biosensor
👉 高再現性、高純度、低批次差異。
2. 抗體親和力測定（K_D、kon、koff）
- Avidity system 全世界都在用來跑 Octet kinetic
3. CAR-T、抗體研發中的抗原呈現技術
- 維持抗原 native conformation
- 單一位點 biotin，避免 multivalent effect干擾親和性
4. ELISA / Lateral Flow 免疫檢測
- 精準控制配體方向性（oriented immobilization）
5. 蛋白工程、重組蛋白純化
6. 高靈敏度 pull-down、Co-IP
四、產品線總表（簡明表格）
| 類別 | 品項 | 用途 | 特點 |
|---|---|---|---|
| Tag 系列 | AviTag peptide、vectors | 蛋白標記 | 單一位點標記 |
| 酵素 | BirA ligase（in vitro / in vivo） | 生物素化 | 高特異性、1:1 修飾 |
| 試劑套組 | Biotinylation kits | 蛋白生物素化 | 標準化流程 |
| 純化材料 | NeutrAvidin / SA beads、resin | 抓取 biotin-proteins | 低背景、高親和 |
| 偵測材料 | Anti-AviTag Ab、SA-HRP、SA-FITC | Western / ELISA / Flow | 多平台通用 |
| 標準品 | Biotinylated proteins / oligos | assay control | 高穩定性 |
| 客製服務 | Custom biotinylation / fusion | 蛋白工程 | GMP 前端支援 |