Resources

RJH is committed to providing our customers with the tools, transfection protocols and advice necessary for success with our products.

This page has some useful references and resources to help you succeed. Including optimization of transfection protocols, transfection reagent selection guidance for your cells, and frequently asked questions. If you require any further assistance please do not hesitate to contact us and our team will be happy to help!

Transfection Reagent Selection Guide

The table below summarizes optimal transfection reagents for nucleic acids in different cell types.
The efficiency of the transfection reagents was assessed by using plasmid DNA (pDNA) or short interfering RNA (siRNA).
Where the transfection reagent was suitable for both pDNA or siRNA delivery, it was indicated with pDNA/siRNA.

Cell Type All-Fect 293-Fect Leu-Fect-A Leu-Fect-B Prime-Fect
Primary Cells
Umbilical Cord Blood Derived Mesenchymal Stem Cells (UCB-MSC) pDNA pDNA pDNA
Bone Marrow Derived Mesenchymal Stem Cells (BM-MSC) pDNA pDNA pDNA
Vascular smooth muscle Cells (VSMCs) pDNA pDNA
Human Umbilical Vein Endothelial Cells (HUVECs) pDNA
Mononuclear Cells from CML patients (MNC) siRNA siRNA
Human Foreskin Fibroblast Cells pDNA
Cell Lines
Kidney Fibroblast Cells (293-T) pDNA pDNA pDNA
Breast Cancer Cells (MDA-MB-231) pDNA
siRNA
co-delivery
pDNA pDNA siRNA
Kidney Epithelial Cells (MDCK) siRNA
Breast Cancer/Melanoma Cells (MDA-MB-435) siRNA
Breast Cancer Cells (MDA-MB-468) siRNA
Breast Cancer Cells (Sum-149PT) siRNA
Breast Cancer Cells (MCF-7) pDNA pDNA pDNA
siRNA
Human Lymphoma Cells (U-937) pDNA pDNA
Chronic Myeloid Leukaemia Cells (K562) siRNA siRNA
Acute Myeloid Leukemia Cells (KG1 and KG1A) siRNA siRNA
Acute Myeloid Leukemia Cells (THP1) siRNA siRNA
Human Lung Cancer Cells (A549) siRNA
Human Colon Cancer (HCT-116) siRNA siRNA
Human Myoblasts ASO

Frequently Asked Questions

1How should I select the best reagent for my cell type?
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.
2Do I need to optimize reagent after purchasing and how can I go about that?
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).
4Which serum-free medium should I use for preparing nucleic acid complexes with transfection reagents?
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.
5How much transfection reagent and nucleic acid (DNA and/or RNA) should I use for complex formation?
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.
6Do I need to remove complexes from cells after transfection?
No. It not necessary to remove the complexes from treated cells. Complexes can be left in culture with cells until end-point analysis.
7How long should I leave the transfection complexes with cells to find the intended effect?
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.
8How can I determine transfection efficiency for my cell type?
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.
9How can I improve transfection efficiency further?
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.
10How can I reduce cytotoxicity in my experiments?
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.
11Can cell density (or confluence) and passage number affect transfection?
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'.
12How should the transfection reagents be stored and what are their shelf-life?
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.
13How many transfections can be performed with 1 ml vial?
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.
14Can I use in vivo reagent for in vitro experiments?
Yes. Our in vivo transfection reagents display broad activities so that they could be effective under in vitro conditions.
15Can I use the reagents for a different type of nucleic acid?
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.
16What is the technology behind RJH Reagents
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.
17How shall I cite your product for a paper I am writing?
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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