RFP-Booster (Atto 488)

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Antigen
Reactivity
Discosoma
Host
Camelid (Camelidae)
Antibody Type
Recombinant Antibody
Conjugate
Atto 488
Application
Fluorescence Microscopy (FM), Immunofluorescence (IF)
Options
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Purpose With our Booster you reactivate, boost, stabilizate the signals of your fusion proteins.
Brand RFP-Trap®
Specificity RFP-Booster efficiently highlights, enhances and stabilizes monomeric red fluorescent proteins such as mRFP1, mCherry or mPlum but also mRuby.
Characteristics
  • Enhance, stabilize and reactivate your fl uorescent proteins
  • RFP-Booster high specifi city for various monomeric red fl uorescent proteins derived from DsRed
  • Coupled to bright and photostable chemical dyes from ATTO-TEC
Components RFP-Trap® coupled to fluorescent dye ATTO 488
Alternative Name RFP
Background Red fluorescent proteins (RFP) and variants thereof are widely used to study protein localization and dynamics in living cells. However, photo stability and quantum efficiency of RFP are not sufficient for Super-Resolution Microscopy (e.g. 3D-SIM or STED) of fixed samples. In addition, many cell biological methods such as BrdU-staining, EdU-Click-iT™ treatment or Fluorescent In Situ Hybridization result in disruption of the RFP signal.The RFP-Booster_Atto594, a specific RFP-binding protein coupled to the fluorescent dye ATTO 594, reactivates, boosts and stabilizes your RFP signal.
Research Area Tags/Labels
Application Notes For the immunofluorescence staining of RFP-fusion proteins in fixed cells

ATTO 488:
Excitation range 480 - 510 nm (λabs= 501 nm)
Emission range 520 - 560 nm (λfl= 523 nm)
Comment

Nano-Booster are specific GFP- or RFP-binding protein coupled to the superior fluorescent dyes from ATTO-TEC. With our Booster you reactivate, boost and stabilize the signals of your fluorescent proteins.

Assay Procedure
  • 1. Fixation: 4 % paraformaldehyde (PFA) or 1:10 formalin (37 % formaldehyde, 10-15 % MetOH) in PBS, 10 min., RT.
  • 2. Wash 3x with PBS containing 0.1 % Tween 20 (PBST). Critical: do not let coverslips ""dry"".
  • 3. Permeabilisation: PBS containing 0.5 % Triton X-100, 5 min., RT. Alternatively, permeabilise by incubating in 100 % methanol for 5 min at -20 °C.
  • 4. Wash 2x with PBST.
  • 5. Blocking: 4 % BSA in PBST, 10 min, RT.
  • 6. RFP-Booster incubation: dilute RFP-Booster 1:200 - 1:400 in blocking buffer and incubate 1 h, RT.
    Note: For multiplexing protocols you can combine RFP-Booster with any other antibody.
  • 7. Wash 3x 5-10 min in PBST.
  • 8. If required, counterstain with DNA fluorescent dyes, e.g. DAPI.
  • 9. Before mounting, coverslips can be very briefly rinsed in water to prevent salt crystals to form.
  • 10. Mount in VectaShield (Vector Labs) or other mounting media with anti-fading agents and seal mounted coverslips with clear nail polish.
Restrictions For Research Use only
Format Liquid
Buffer PBS, 0.01 % Sodium azide
Preservative Sodium azide
Precaution of Use This product contains sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.
Handling Advice Do not freeze. Protect from light.
Storage 4 °C
Expiry Date 6 months
Product cited in: Vietri, Schink, Campsteijn, Wegner, Schultz, Christ, Thoresen, Brech, Raiborg, Stenmark: "Spastin and ESCRT-III coordinate mitotic spindle disassembly and nuclear envelope sealing." in: Nature, Vol. 522, Issue 7555, pp. 231-5, 2015 (PubMed).

Osterfield, Schüpbach, Wieschaus, Shvartsman: "Diversity of epithelial morphogenesis during eggshell formation in drosophilids." in: Development (Cambridge, England), Vol. 142, Issue 11, pp. 1971-7, 2015 (PubMed).

Platonova, Winterflood, Junemann, Albrecht, Faix, Ewers: "Single-molecule microscopy of molecules tagged with GFP or RFP derivatives in mammalian cells using nanobody binders." in: Methods (San Diego, Calif.), 2015 (PubMed).

Bleck, Itano, Johnson, Thomas, North, Bieniasz, Simon: "Temporal and spatial organization of ESCRT protein recruitment during HIV-1 budding." in: Proceedings of the National Academy of Sciences of the United States of America, Vol. 111, Issue 33, pp. 12211-6, 2014 (PubMed).

Winterflood, Ewers: "Single-Molecule Localization Microscopy using mCherry." in: Chemphyschem : a European journal of chemical physics and physical chemistry, Vol. 15, Issue 16, pp. 3447-51, 2014 (PubMed).

Biermann, Sokoll, Klueva, Missler, Wiegert, Sibarita, Heine: "Imaging of molecular surface dynamics in brain slices using single-particle tracking." in: Nature communications, Vol. 5, pp. 3024, 2014 (PubMed).

Hasegawa, Ryu, Kaláb: "Chromosomal gain promotes formation of a steep RanGTP gradient that drives mitosis in aneuploid cells." in: The Journal of cell biology, Vol. 200, Issue 2, pp. 151-61, 2013 (PubMed).

Franz, Roque, Saurya, Dobbelaere, Raff: "CP110 exhibits novel regulatory activities during centriole assembly in Drosophila." in: The Journal of cell biology, Vol. 203, Issue 5, pp. 785-99, 2013 (PubMed).

Weil, Parton, Herpers, Soetaert, Veenendaal, Xanthakis, Dobbie, Halstead, Hayashi, Rabouille, Davis: "Drosophila patterning is established by differential association of mRNAs with P bodies." in: Nature cell biology, Vol. 14, Issue 12, pp. 1305-13, 2012 (PubMed).