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www. .com Applications: • mRNA translation and degradation studies • Protein biochemistry studies – synthesis, folding, processing, stability, localization, degradation • mRNA transfer into primary cells without generating genetically modified organisms (GMO) • Genome engineering by RNA-only CRISPR/Cas technology Highly efficient, especially in primary cells (e.g., neurons or HUVECs) and stem cells Immediate mRNA translation – protein synthesis detectable after 15-30 minutes Immediate mRNA delivery – no endocytosis or lysosomal degradation Extremely low cytotoxicity in cooperation with Fuse-It-mRNA The Shortcut for Protein Expression Primary cortical neurons + eGFP-mRNA Primary myocytes + LifeAct ® - TagGFP2-mRNA

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Page 1: Fuse-It-mRNA · 60500 Fuse-It-mRNA, infrared fluorescent: 2 x 150 µl solution 60501 Fuse-It-mRNA, infrared fluorescent: 2 x 300 µl solution For additional Fuse-It products, please

www. .com

Applications:

• mRNA translation and degradation studies

• Protein biochemistry studies – synthesis, folding, processing, stability, localization, degradation

• mRNAtransferintoprimarycellswithoutgeneratinggeneticallymodifiedorganisms(GMO)

• GenomeengineeringbyRNA-onlyCRISPR/Castechnology

Highlyefficient,especiallyinprimarycells(e.g.,neuronsorHUVECs)andstemcells

ImmediatemRNAtranslation–protein synthesis detectable after15-30minutes

ImmediatemRNAdelivery–no endocytosis or lysosomal degradation

Extremely low cytotoxicity

in cooperation with

Fuse-It-mRNATheShortcutforProtein Expression

Primary cortical neurons + eGFP-mRNA

Primary myocytes + LifeAct®-TagGFP2-mRNA

Page 2: Fuse-It-mRNA · 60500 Fuse-It-mRNA, infrared fluorescent: 2 x 150 µl solution 60501 Fuse-It-mRNA, infrared fluorescent: 2 x 300 µl solution For additional Fuse-It products, please

ImmediatemRNADelivery

The Fuse-It liposomal carrier, which includes the mRNA, simply fuses with the cell membrane and then releases the mRNA directly into the cytoplasm. mRNA translation starts immediately, without the interfering processes of endocytosis, lysosomal degradation, or mitosis. Unlike classical lipoplex-based delivey methods, cells do not internalize the mRNA by endocytosis.

MembraneFusion–ACompletelyNewTechnology

Cytotoxicity of Fuse-It-mRNA versus classical lipoplex-based method:In contrast to classical lipoplex-based methods, Fuse-It-mRNA fusion does not harm cells. Sixteen hours after eGFP-mRNA transfer by membrane fusion of primary human epithelial cells (nHEK) with Fuse-It-mRNA, the vitality of the cells is high, whereas significant cell death occurred in the cells treated with classical lipoplex-based methods.

HighEfficiency,EveninPrimaryCells

Based on the charge of natural cell membranes, Fuse-It-mRNA liposomes are able to effectively fuse with most cell types. Cell lines, non-proliferating cells (e.g., neurons), and a broad spectrum of difficult-to-transfect primary cells can directly translate the mRNA in the cytosol. Membrane fusion with Fuse-It-mRNA results in fast and highly efficient protein expression with no risk of genomic integration.

mRNA Transfer versus DNA Transfection: Fuse-It-mRNA results in extremely fast mRNA expression. Six hours after eGFP-mRNA transfer by membrane fusion of CHO-K1 cells with Fuse-It-mRNA, nearly 100% of the cells express eGFP, whereas the number of eGFP-expressing cells after plasmid-transfection is below 10%.

ExtremelylowCytotoxicity

In contrast to classical lipoplex-based methods, Fuse-It-mRNA requires only brief incubation times - between 5 and 20 minutes for maximal nucleotide transfer rates. Furthermore, low amounts of liposomal lipids result in maximal mRNA transfer, without the need for chemical compounds for endosomal release. This feature protects sensitive cells from potential toxic effects of carrier reagents.

Easy Handling

The Fuse-It-mRNA protocol is easy to use because the ratio between fusion reagent and mRNA remains constant in each experiment. The reagent can be used without restrictions because no genetically modified organisms are generated.

Membrane Fusion DNA Transfection

6 h 6 h

2 h 2 h

Fuse-It-mRNAClassical lipoplex-based method

Page 3: Fuse-It-mRNA · 60500 Fuse-It-mRNA, infrared fluorescent: 2 x 150 µl solution 60501 Fuse-It-mRNA, infrared fluorescent: 2 x 300 µl solution For additional Fuse-It products, please

Fuse-It-mRNATheShortcutforProteinExpression

Fuse-It-mRNA Membrane Fusion

Cytoplasm

mRNA

mRNA

MEMBRANEFUSION

Fuse-It mRNA

LiposomalCarrier

TRANSCRIPTION

PlasmidTransfectionreagent

DNA lipid complex

DNA TRANSFECTION

RELEASE OF DNA

Endosome

ENDO-CYTOSIS

ENDO-CYTOSIS

mRNA lipid complex

mRNA TRANSFECTION

mRNA

RELEASE OF mRNA

Endosome

Transfectionreagent

TRANSLATION

Protein

MembraneFusion–TheDirectWaytoProteinExpression

Page 4: Fuse-It-mRNA · 60500 Fuse-It-mRNA, infrared fluorescent: 2 x 150 µl solution 60501 Fuse-It-mRNA, infrared fluorescent: 2 x 300 µl solution For additional Fuse-It products, please

ProteinSynthesisaftertheTransferofeGFP-mRNAinCHO-K1CellsUsingFuse-It-mRNA:

Tel.: +4989/5204617-0Fax: +4989/5204617-59 E-Mail: [email protected] © ibidi GmbH, V 1.0, 2016 / 04

ibidiGmbHAmKlopferspitz1982152Planegg/MartinsriedGermany

EfficiencyofmRNAExpressioninVariousCellTypesafterTreatmentwithFuse-It-mRNA:

OrderingInformation:

Cat.No. Description

60500 Fuse-It-mRNA, infrared fluorescent: 2 x 150 µl solution

60501 Fuse-It-mRNA, infrared fluorescent: 2 x 300 µl solution

ForadditionalFuse-Itproducts,please refer to:

www.ibidi.com

Forfurthercelltypes,pleasereferto:www.ibidi.com

Within 2 hours of starting the experiment, the eGFP is already detectable in the cells. The eGFP amount increases and remains stable for more than 3 days (90 hours).

NOTE: Using functionally capped and polyadenylated mRNAs will achieve the best results.

CellType Organism IncubationTimeEfficiencyofmRNA

Expression

PrimaryCells

Cortical Neurons (embryonal) rat 5-15 min 70-90%Hepatocytes human 15-25 min 60-80%Keratinocytes (foreskin) human 10-15 min 70-80%Monocytes / Macrophages human 10-15 min 30-50%Myocytes (embryonal) rat 15-25 min 40-60%Myofibroblasts (embryonal) rat 15-20 min 30-50%

StemCells

Cor.4U Myocytes (iPSC derived cardiomyocytes)

human 10‐20 min 80‐100%

Dopa.4U (iPSC derived dopaminergic neurons)

human 5‐10 min 70‐90%

iPSC human 10‐20 min 70‐90%Peri.4U (iPSC derived peripheral neurons)

human 5‐10 min 70‐90%

CellLines

CHO-K1 hamster 15-20 min 80-100%HeLa human 15-20 min 70-90%MCF10A human 10-15 min 80-100%RAW 264.7 rat 15-20 min 50-70%

2 h 20h 40h 90h

Formoreinformationonourproducts,pleasevisitusat:

www.ibidi.com

Fuse-It-mRNATheShortcutforProteinExpression


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