Lamin-Chromobody plasmid

Description
DNA plasmid encoding for anti-Lamin VHH (anti-Lamin Nanobody) fused to fluorescent protein.

Specificity
Lamin A/C, tested in human cell lines (HeLa, U2OS) and insect S2 cells

Applications

Trace the nuclear lamina in live cells in real-time
Monitor the nuclear integrity and morphology
Use the Lamin-Chromobody® as a biosensor for real-time assays of mitosis or apoptosis

Lamin-Chromobody plasmid
Lamin-Chromobody plasmid (TagGFP2)

Product Size Code Price Buy
Product Lamin-Chromobody plasmid (TagGFP2) Size 20 µg Code lcg Price $ 999
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Lamin-Chromobody format

 

Lamin-Chromobody
plasmid (tagGFP)

Vector type

mammalian
expression vector

Promoter

constitutive CMV IE

Reporter

TagGFP2

Codon usage

mammalian

Selection

Kan/Neo

Nuclear localization signal (NLS)

No

Please contact us for more details.

Specificity
Lamin A/C, tested in human cell lines (HeLa, U2OS) and insect S2 cells

Encoded Nanobody/ VHH
Monoclonal anti-Lamin single domain antibody (sdAb) fragment

Sequence
With the Lamin-Chromobody plasmid you receive the sequence information of the Alpaca antibody to Lamin fused to TagGFP2, as well as the full vector sequence.

Microscopy techniques
Wide-field epifluorescence microscopy; confocal microscopy

Transfection
Transfection of Chromobody plasmids into mammalian cells can be done with standard DNA-transfection methods, e.g. lipofection (Lipofectamine 2000® from Thermo Scientific), according to the manufacturer’s protocol for the transfection reagent. Please choose the transfection method that works the best for your cell type.

Storage instructions
Shipped at ambient temperature. Store at -20 °C

Rothbauer U, Zolghadr K, Tillib S, Nowak D, Schermelleh L, Gahl A, Backmann N, Conrath K, Muyldermans S, Cardoso MC, Leonhardt H. Targeting and tracing antigens in live cells with fluorescent nanobodies. Nat Methods. 2006 Nov;3(11):887-9. PMID: 17060912

Schmidthals K, Helma J, Zolghadr K, Rothbauer U, Leonhardt H. Novel antibody derivatives for proteome and high-content analysis. Anal Bioanal Chem. 2010 Aug;397(8):3203-8. doi: 10.1007/s00216-010-3657-0. Epub 2010 Apr 7. PMID: 20372881

Zolghadr K, Gregor J, Leonhardt H, Rothbauer U. Case study on live cell apoptosis-assay using lamin-chromobody cell-lines for high-content analysis. Methods Mol Biol. 2012;911:569-75. doi: 10.1007/978-1-61779-968-6_36. PMID: 22886277

 

Are Chromobodies constitutively expressed?

Yes, Chromobody expression is regulated by immediate early promotor CMV. This promotor allows constitutive Chromobody expression.

Do Chromobodies only work in live cells?

Yes, the Chromobody plasmid is only expressed in live cells. Cells should be transfected with the Chromobody plasmid at least overnight to observe the Chromobody location signal. Alternatively, cells can be fixed prior to imaging.
Note: We don't recommend fixation of cells for the Histone-Chromobody.

When should I image my cells after transfection with the Chromobody plasmid?

The Chromobody signal is maintained up to 3 days in the cell. However, this also depends strongly on the cell type.
We recommend to image the cells 16-24 hours after transfection.

Do the Chromobodies diffuse through the cell membrane into growth medium?

No, Chromobodies are small proteins being expressed in the cytosol. They are not secreted into the medium and remain in the cell as long as the cell maintains its plasma membrane integrity.

Are Chromobodies fluorogenic or do they only emit fluorescence when bound to a target?

Chromobodies are chimeric proteins consisting of a VHH fused to a fluorescent protein. They maintain their fluorescence regardless of whether they are bound to a target or not.

Can I amplify the Chromobody plasmid in bacteria?

Yes, the Chromobody plasmids can be propagated in E.coli by standard techniques.

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Only for research applications, not for diagnostic or therapeutic use!