RJH Biosciences Inc.


RJH Biosciences社ホームページ
RJH Biosciences社は、カナダのアルバータ州エドモントンを拠点とするバイオテクノロジー企業です。RJH Biosciences社は、様々な種類の核酸をヒト細胞や培養細胞株に運搬する今までにないトランスフェクション試薬や輸送システムを開発しています。RJH Biosciences社は「生物医学の研究開発企業に向けたトランスフェクション試薬」と、「前臨床・臨床用の核酸輸送用媒体」という商業上の2つのセグメントにおいて、付加価値のある商品を開発いたします。血液癌や免疫細胞の修飾をテーマとするRJH Biosciences社独自の研究開発プロジェクトでも、同様のトランスフェクション試薬が用いられています。

 

製品:トランスフェクション試薬

試薬の概要








All-Fect

siRNA pDNAトランスフェクション試薬として設計されており、siRNAによるノックダウンやプラスミドDNAのトランスフェクション、siRNAとpDNAの同時輸送に使用できます。All-Fectは、特に臍帯血・骨髄由来の間葉系幹細胞や、高度に分化した平滑筋細胞、内皮細胞での使用に適しています。

 

資料をダウンロード:All-Fect Brochure

詳細情報:メーカーページ

試薬の特徴:

  • 高効率のトランスフェクション

    血清の存在下で従来製品の2-3倍の効力を発揮します。

  • シンプルなプロトコル

    トランスフェクション中に組織の培養液を交換する必要がありません。

  • 毒性

    市販のトランスフェクション試薬より毒性が低く、正常な細胞の生理活性の維持に優れています。

リファレンス:

  • Hsu and Uludağ. Biomaterials (2012) 33: 7834-7848.
  • Remant Bahadur et al., J. Materials Chemistry B (2015) 3: 3972-3982.
  • Wang et al., J. Surgical Research (2013) 183: 8-17.
All-fect transfection of cells GFP

写真:臍帯血由来の間葉系幹細胞にGFPプラスミドをトランスフェクションした際に見られた、All-Fectの典型的な性能。この実験ではAll-Fectと他社のリポフェクション試薬をそれぞれの最適条件下で使用し、性能を比較しています。導入遺伝子の発現の度合いは、任意単位でのEGFPの発現度合いに基づいてフローサイトメトリーにより測定されました。上の写真は、トランスフェクション後のGFPの発現を示す典型的な蛍光顕微鏡像を示しています。

製品の特徴

RJH Biosciences社は、プラスミドDNAやsiRNA、mRNA、その他の核酸を用いて哺乳類細胞を改変するために、幅広く活用できるトランスフェクション試薬を開発いたしました。RJH Biosciences社のトランスフェクション試薬の基盤となっているのは、正電荷と疎水性(脂質)グループのバランスが最適な陽イオン性の脂質のポリマーです。ポリマーの骨格と脂質グループの性質を体系的に変えることで、トランスフェクション試薬のライブラリが作成されました。ある種のトランスフェクション試薬は異なる細胞種や核酸で幅広く機能し、他の種類の試薬は特定の細胞種へ特定の核酸運搬に非常に効果的に機能します。

RJH Biosciences社の輸送体の利点:

  • 核酸との多価の相互作用によってカーゴの強固な結合が形成され、細胞膜を通過する際の破壊的作用に耐える
  • 陽イオン性と脂質性の結合による相乗効果によりカーゴが覆われ、ヌクレアーゼから保護される
  • 脂質成分により、細胞膜との相互作用や細胞内への輸送が促される
  • pH緩衝能によりエンドソームからのカーゴの脱出が促される
  • 細胞質内に移行したフリーの核酸が目的に合わせて処理される

トランスフェクション試薬は以下の細胞モデルや用途に最適化されています。

初代細胞

接着細胞

  • VSMC (Vascular Smooth Muscle Cells)
  • HUVEC (Human Umbilical Vein Endothelial Cells)
  • Human Foreskin Fibroblast Cells
  • BMSC (Bone Marrow Stromal Cells)
  • Human Myoblasts
  • Rat Primary Sympathetic Neurons

浮遊細胞

  • UCB-MSC (Umbilical Cord Blood Derived Mesenchymal Stem Cells)
  • BM-MSC (Bone Marrow Derived Mesenchymal Stem Cells)
  • Mononuclear Cells from Leukemia Patients
  • Mononuclear Cells from Normal Human Blood
細胞株

接着細胞

  • 293-T (Kidney Fibroblast Cells)
  • MDA-231 (Breast Cancer Cells)
  • MDA-436 (Breast Cancer/ Melanoma Cells)
  • MDA-468, Sum-149PT, MCF-7 (Breast Cancer Cells)
  • A549 (Human Lung Cancer Cells)
  • MDCK (Kidney Epithelial Cells)
  • HCT-116 (Human Colon Cancer Cells)
  • C2C12 Myoblast Cells
  • MC3T3-1 (Preosteoblast Cells)
  • Green Monkey Vero Cells

浮遊細胞

  • U-937 (Human Lymphoma Cells)
  • K562 (Chronic Myeloid Leukemia Cells)
  • KG1, KG1A and THP-1 (Acute Myeloid Leukemia Cells)
  • Jurkat T-Cells
動物モデル
  • Systemic and local injection of RNAi mediator siRNA
  • Local injection of pDNA and mRNA expression vectors

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Background of Service

Our R&D Focus

One area of focus for RJH Biosciences is to implement RNA interference (RNAi) via delivery of short interfering RNA (siRNA). Our initial therapeutic application is blood cancers, while recognizing that the RNAi activity can be implemented in the treatment of a large range of human cancers and other diseases. Another focus is direct administration of plasmid DNA (pDNA) to express therapeutic proteins in situ, with applications in immunotherapy.

Why study blood cancers and immunotherapies with nucleic acid therapeutics?

There are three types of blood cancers: Leukemia, lymphoma, and myeloma. Leukemia is characterized by highly proliferating, abnormal white blood cells [1]. Lymphoma and myeloma are respectively cancers of the lymphatic system and plasma cells which greatly effect the immune system [2,3]. These three cancers are difficult to treat and the current treatments are limited in efficacy, especially at the end stage of the disease.

The use of nucleic acid-based therapeutics can eradicate these cancers in two primary ways, with RNAi technology and cell-based immunotherapy. The use of RNAi is being increasingly explored in the treatment of the blood cancers. Polynucleotides such as siRNA has aided the downregulation of oncogenes and can be designed to support specific abnormalities in individual patients, making it a ‘personal’ strategy with a universal technological design [4]. Due to siRNA’s potential in blood cancer therapies, we are currently focusing on siRNA therapeutics in our R&D projects, targeting disease-driving oncogenes and inducing apoptosis in the malignant cells.

Another strategy that is being explored for treating blood cancers is the use of immunotherapy. Immunotherapeutic strategies include the use of antibodies, stem cell transplants, cytokines, small molecules among others [5]. However, a more recent approach is genetic therapy by using engineered cells, also known as Cell Transfer Therapy. This method works by taking patients’ own immune cells such as but not limited to T-cells, B-cells, and NK cells. The immune cells genome is engineered to support various therapeutic strategies that may involve neoantigen expression and presentation on immune cell surface, and then are reintroduced into the host [5]. The modified cells are ultimately designed to target and remove the malignant cells. This strategy is highly advantageous as T-cells can ‘seek’ and destroy the malignant cells in the blood system. While this approach has been promising in blood cancers, it can be also used in other solid cancers. As the foundation of immunotherapy relies on nucleic acid introduction into patient cells, efficient delivery of nucleic acids is imperative for success. Our transfection reagents offer the best in class vehicles to undertake such a delivery.

Nanomedicine based on nucleic acid therapeutics is a large component to personalized cancer therapies and immunotherapies. The RJH Biosciences strives to provide quality transfection reagents, whether it involves the delivery of our own nucleic acid candidates or our customers’.

Jean, C. and Dick, J. (2005) Cancer stem cells: lessons from leukemia. Trends in cell biology. 15, 494-501.
Woods, N. et al. (2006) Therapueti gene causing lymphoma. Nature. 440, 1123.
Mahindra, A. et al. (2012) Latest advances and current challenges in the treatment of multiple myeloma. Nature Reviews Clinical Oncology. 9, 135-143.
Uludağ, H. et al. (2016) Current attempts to implement siRNA-based RNAi in leukemia models. Drug Discovery Today. 21, 1412-1420.
Zou, W. (2006) Regulatory T cells, tumour immunity and immunotherapy. Nature Reviews Immunology. 6, 295-307.

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製品一覧

製品一覧

製品名 説明 容量
All-Fect 幅広い種類の細胞に対応している、pDNAやsiRNAの輸送または同時輸送用の試薬 0.75 ml/1.5 ml
Prime-Fect 導入が難しい、初代細胞や幹細胞へのトランスフェクションに特化した試薬 0.75 ml/1.5 ml
Leu-Fect A & B 白血病細胞や浮遊細胞へのトランスフェクションに特化した試薬 0.75 ml/1.5 ml
Trans-Booster 接着細胞や浮遊細胞への、DNAやmRNAのトランスフェクション効率を向上するよう設計された試薬 0.75 ml/1.5 ml
In Vivo DNA-Fect 動物モデルや動物細胞へのpDNA輸送に効果的な試薬 0.75 ml/1.5 ml
In Vivo RNA-Fect 動物モデルへの全身性のsiRNA輸送に効果的な試薬 0.75 ml/1.5 ml
mRNA-Fect mRNAの輸送に最適な、高効率のトランスフェクション試薬 0.75 ml/1.5 ml
CRISP-Fect 接着細胞や浮遊細胞へのリボ核タンパク質(RNP)の輸送に最適な、高効率のトランスフェクション試薬 0.75 ml/1.5 ml

トランスフェクション試薬の選択ガイド

以下の表は、細胞タイプ・核酸ごとに適したトランスフェクション試薬をまとめたものです。トランスフェクション試薬の効果は、プラスミドDNA (pDNA)、低分子干渉RNA (siRNA)、メッセンジャーRNA (mRNA)を用いて評価されています。
トランスフェクション試薬がpDNAとsiRNAのどちらの輸送にも適している場合、pDNA/siRNAと表記されています。

Cell Type All-Fect Trans-Booster Leu-Fect-A Leu-Fect-B Prime-Fect mRNA-Fect CRISP-Fect
Primary Cells
Umbilical Cord Blood Derived Mesenchymal Stem Cells (UCB-MSC) pDNA pDNA
mRNA
pDNA
Bone Marrow Derived Mesenchymal Stem Cells (BM-MSC) pDNA pDNA
mRNA
pDNA
Vascular smooth muscle Cells (VSMCs) pDNA
mRNA
pDNA
Human Umbilical Vein Endothelial Cells (HUVECs) pDNA
mRNA
Mononuclear Cells from CML patients (MNC) pDNA
mRNA
siRNA siRNA
Human Foreskin Fibroblast Cells pDNA
Rat Primary Sympathetic Neurons pDNA mRNA
Cell Lines
Kidney Fibroblast Cells (293-T) pDNA pDNA
mRNA
pDNA
Breast Cancer Cells (MDA-MB-231) pDNA
siRNA
co-delivery
pDNA
mRNA
pDNA siRNA mRNA
Kidney Epithelial Cells (MDCK) siRNA
Breast Cancer/Melanoma Cells (MDA-MB-436) pDNA
mRNA
siRNA mRNA RNP
Breast Cancer Cells (MDA-MB-468) siRNA
Breast Cancer Cells (Sum-149PT) pDNA
mRNA
siRNA
Breast Cancer Cells (MCF-7) pDNA pDNA
mRNA
pDNA
siRNA
mRNA
Human Lymphoma Cells (U-937) pDNA pDNA
Chronic Myeloid Leukaemia Cells (K562) siRNA pDNA
mRNA
siRNA mRNA
Acute Myeloid Leukemia Cells (KG1 and KG1A) siRNA siRNA
Acute Myeloid Leukemia Cells (THP1) siRNA siRNA mRNA
Human Lung Cancer Cells (A549) siRNA
Human Colon Cancer (HCT-116) siRNA siRNA
Human Myoblasts ASO
Jurkat Cells pDNA mRNA RNP

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使用例

Feedback from independent researchers

Implementing CRISPR-Cas9 Technology using Transfection Reagents from RJH Biosciences
Killing Lung Cancer A549 Cells with RJH Reagents and Cytotoxic siRNAs
mRNA Transfection of Endothelial Cells with RJH Reagents
Transfecting Human Astrocytes with RJH Reagents: GFP Expression and siRNA Uptake
Transfection of A549 Lung Cancer Cells with RJH Reagents to Silence Tumor Suppressor p53 Expression
Transfecting Colon Cancer HCT-116 Cells with RJH Reagents to silence Polynucleotide Kinase 3′-Phosphatase (PNKP) Expression
Use of RJH Transfection Reagents in Antisense Oligonucleotide Delivery
Reagents for pDNA and siRNA delivery for TNBC cells
 

Application Notes

Reagents for Antisense Oligonucleotide (ASO) Delivery
Comparing Lipofection to RJH Reagents
mRNA Modification of PBMCs
mRNA-Fect Transfection Reagent to Deliver mRNA in Breast Cancer Cells
Transfecting Triple-Negative Breast Cancer MDA-MB-231 Cells with Plasmid DNA and siRNA by using ALL-Fect and Prime-Fect
Use of RJH Transfection Reagents in Co-Delivery of Plasmid DNA and short interfering RNA
RJH Transfection Reagents in siRNA Library Screens
microRNA Delivery to Leukemic Cells
 
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Q&A

As the first step, we suggest consulting the ‘Transfection Reagent Selection Guide’ to find out if the cells of interest have been previously tested. If your cell type is not on the list, we suggest using the reagents optimized for similar cells (i.e., attachment-dependent/suspension or established cell line/primary cells), or contact us by e-mail for an informed suggestion.

We recommend small scale preliminary experiments to optimize the performance of our transfection reagents. As with all transfection reagents, the complex formation, cell seeding density, and culture and incubation conditions will affect the final performance. Please consult our technical sheet on optimizing transfection for this purpose (link).

Our transfection reagents are compatible with a wide range of serum-free media, including DMEM, RPMI, MEM and others. Complexes are expected to be functional in serum-containing medium.

The recommended nucleic acid to transfection reagent ratio ranges from (w/w) 1:1 for relatively toxic reagents to 5-20:1 for biocompatible reagents. This ratio should be optimized for each application. For suggested ranges for each reagent, please consult the specific reagent manual.

No. It not necessary to remove the complexes from treated cells. Complexes can be left in culture with cells until end-point analysis.

Depending on the application, incubation times may vary from 2 hours to 24 hours. It may be possible to centrifuge the treated cells to accelerate the transfection process and minimize the complex incubation times.

Transfection efficiency can be determined by different approaches. Reporter genes, such as GFP or RFP, are convenient ways to assess transfection by using microscopy or flow cytometry techniques. Direct assessment of the induced gene product (e.g., by ELISA, western blot), or silenced gene expression (e.g., protein levels or mRNA levels by PCR) are important. One can also use functional outcomes as an indirect measure of transfection, although care must be paid to complicating factors in this case. In all studies, we recommend employing a control (i.e., non-active) agent similar in nature to the nucleic acid being investigated.

A methodical analysis of the factors contributing to transfection, as outlined in the ‘Technical Tips to Improve Transfection’, is a good place to start. Other resources to improve transfection efficiencies can be found in our ‘suggested reading’ tab. We are here to help as well, so do send us a message/e-mail to see how we can assist you.

All transfection reagents display a certain extent of cytotoxicity on cells, depending on the amount used. The key is to achieve transfection without disrupting the physiology of the cells significantly. One can minimize exposure time to transfection complexes, speed up the transfection process by centrifugation and optimize reagent/nucleic acid concentrations to eliminate unnecessary exposure. The purity of the nucleic acid is also important to eliminate unforeseen toxicities.

These are important criteria that affect the transfection efficiency. In general, transfection efficiency decreases with increased cell density and passage number, as cells settle in senescence. For other factors affecting transfection efficiency, please consult ‘Technical Tips to Improve Transfection’.

The recommend storage temperature is at 4 degC (short term) or -20 degC (long term). The reagents are designed to be stable for 1 year under these conditions.

We recommend to use 1:5 ratio of nucleic acid to transfection reagent, so that 1 mL vial (1 mg) is suitable for 200 transfections of 1 mg nucleic acid. However, optimal ratio may change depending on the application and cell type.

Yes. Our in vivo transfection reagents display broad activities so that they could be effective under in vitro conditions.

It is likely for our transfection reagents to work with different nucleic acids. This is not universally applicable, but most reagents seem to handle different nucleic acids. ALL-Fect transfection reagent can handle both DNA and RNA.

Our reagents are based on an optimal balance of cationic charge and hydrophobicity. They are polymeric in nature that interact with nucleic acids via multivalent interactions. These reagents provide effective condensation of anionic charge of nucleic acids, while displaying little toxicity on mammalian cells.

To indicate the source of the transfection reagent, you can state the reagent name and that it was obtained from RJH Biosciences Inc. (Edmonton, AB, Canada).

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【商品取扱元】株式会社 東京未来スタイル
info@tokyofuturestyle.com
TEL:029-851-9222 FAX:029-851-9220

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