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Cell fusion reagent
GenomONETM-CF

GenomONE-CF is a cell fusion kit composed of HVJ Envelope (HVJ-E) and special buffers. It can be used with both adhering cells and floating cells. Fusion of cells of the same or different types is possible with this kit in only 30 minutes.

*This product is sold for research purpose only. It may not be used for treatment or other clinical purposes or for intra- and extracorporeal diagnosis in humans or animals.

Applications

  • Hybridoma preparation
    (Fusion of antigen-sensitized B cells and myeloma cells)
  • Cancer vaccinations and cancer immunology
    (Fusion of cancer cells and dendritic cells)
  • Regenerative medicine and cytotherapy
    (Fusion of somatic cells and stem cells)
  • Embryonic development/differentiation
    (Nuclear transfer, nuclear replacement)
  • An alternative method to PEG and electrofusion for cell fusion

Specification・price

Cat. # Freeze-dried HVJ-E
(Equivalent to 0.26mL/vial)
HVJ-E Suspending Buffer
(0.5mL/vial)
Cell Fusion Buffer
(10mL/vial)
Price
Excluding Tax
Price
Including Tax
CF001 1 1 1 ¥18,000 ¥19,800
CF004 4 3 4 ¥55,000 ¥60,500

What is HVJ Envelope (HVJ-E)?

Hemagglutinating virus of Japan (HVJ) Envelope (HVJ-E) is a nonproliferative and noninfectious vesicle about 300 nm in diameter on average purified after complete  inactivation of Sendai virus genomic RNA. Since the F protein distributed on the HVJ-Envelope has high membrane-fusing potential comparable to that of live virus,  it is possible to use HVJ-E itself as a cell-fusing agent or to introduce genes, proteins, anti-cancer agents, etc. in HVJ-E-incorporated form into cells for analysis  of their functions.
HVJ (Hemagglutinating virus of Japan) is also called Sendai virus (SeV) or Mouse Parainfluenza virus type 1

*HVJ:Hemagglutinating Virus of Japan

Principle for cell fusion triggered by HVJ-E

If HVJ-E is added in amounts of more than several hundred HVJ-E per cell at low temperatures (0-8°C), HVJ-E is immediately adsorbed on the cell surface mediated by the receptor (acetyl type sialic acid recognized by HN protein) (Step 1), and cells undergo agglutination cross-linked by HVJ-E particles  (Step 2). At this stage, the hydrophobic domain at the N-terminal of cleaved F protein (F1) penetrates into the double lipid layer of the cell membrane, causing distortion of the membrane severe enough to allow an inflow of ions.

If this cell / HVJ-E complex is heated at 37°C, the distortion of the cell membrane is further expanded, accompanied by temporary alteration of the cell membrane lining structure. This change is transient  and the membrane soon returns to its normal structure. However, if a strong hydrophobic connective  force is applied at this stage, fusion between cell membranes takes place (Step 3).

1. Fusion of suspension cells

Cells labeled with red fluorescence*1 were combined with cells labeled with green fluorescence*2 (each 1-2 x105 cells) in a fusion buffer (50µL). HVJ-E was added to the mixture, followed by incubation on ice for 5 minutes and further incubation at 37°C for 15 minutes. Fused cells (yellow) were obtained.

2. Fusion of adherent cells and suspension cells

Cells labeled with red fluorescence*1 were combined with cells labeled with green fluorescence*2 (each 1-2 x105 cells) in a fusion buffer (50µL). HVJ-E was added to the mixture, followed by incubation on ice for 5 minutes and further incubation at 37°C for 15 minutes. Fused cells (yellow) were obtained.

3. Comparison with PEG method

Rat MSC cells (rat bone marrow-derived mesenchymal stem cells) labeled with red fluorescence*1 were combined with rat primary cardiac myocytes labeled with green fluorescence*2 (each 2 x 105 cells) in 50 µL of a buffer for cell fusion. The mixture was incubated on ice for 5 minutes and then at 37°C for 15 minutes. As a result, fused cells (yellow) were formed (GenomONE-CF suspension method). Fused cells adhering to the plate were also observed after 1-2 days of culturing. In the PEG-treated group, high cytotoxicity appeared immediately after cell fusion, reducing the number of fused cells obtained.

(*1)Labeled with PKH26 Red Fluorescent Cell Linker Kit (Sigma)

(*2)Labeled with PKH67 Green Fluorescent Cell Linker Kit (Sigma)

4. Comparison with PEG method in hybridoma preparation

Hybridoma supplement・・・BM Condimed H1 (Roche) is prepared from the supernatant of a mouse lymphoma cell line stimulated with PMA. It contains a complex mixture of growth factors and cytokines that have a marked stimulatory effect on the growth of hybridoma cells after fusion and during cloning.

Use of GenomONE-CF resulted in more efficient formation of antibody-producing hybridoma than that of PEG. The efficiency of cell fusion mediated by GenomONE-CF was increased by the addition of hybridoma supplement to the medium used for incubation after cell fusion.

Suspension Method

The suspension method is used to fuse suspension cells with suspension cells of the same or different type.

Plating Method

The plating method is used to fuse adherent cells with suspension cells.

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