EdU Click-594

The number at the end of the product name specifies the excitation wavelength of the included fluorescent marker. Suitable excitation source are:

Our ROTI^®Kits for EdU mediated detection of cell proliferation provide a superior alternative to time-consuming and less sensitive bromodeoxyuridine (BrdU) assays. Similar to BrdU assays, our assays utilise a uracil derivative (in this case ethynyldeoxyuridine, EdU) which is incorporated into the replicating DNA. EdU is not detected via antibodies; instead fluorescent dyes are coupled directly via click reaction. This fast and elegant approach allows for direct labelling of replicating DNA with the desired fluorescent dye. Thus, the assay is perfectly adaptable to the present facilities and to the assay situation (e.g. for double or triple detection). Furthermore, the click reaction is highly selective and therefore prevents any unspecific labelling. The protocol includes only few steps and decreases the total amount of time.

Kits available for:

• Fluorescence microscopy (ROTI^®Kits for Imaging)
• Flow Cytometry (ROTI^®Kits for Flow Cytometry)
• High Throughput Screening (ROTI^®Kits for High Throughput Screening)

Advantages of the click chemistry based approach:

Schematic illustration of the click chemistry based EdU cell proliferation assay. A) Incubation of cells with EdU. EdU is a thymidine analogue and is incorporated in DNA during active DNA synthesis. B) After DNA synthesis, EdU was incorporated in the DNA strands. C) Detection of proliferating cells via click chemistry. This reaction is finished within 30 minutes, whereby a variety of different fluorescent dye azides can be used.

Advantages of the Click Assay:

• Highly reliable
• Easy handling
• Fast detection procedure (only 30 min.)
• Mild conditions, no DNA denaturation required
• Compatible with various dyes
• Modular system in click chemistry
• Compatible with multiplexing

The copper(I)-catalysed azide-alkyne cycloaddition (CuAAC) is the most prominent example of a group of reactions named click reactions. According to Sharpless’ definition, these reactions are characterised by high yields, mild reaction conditions, and by their tolerance of a broad range of functional groups.^[1-3] Typically, the reactions require simple or no workup or purification of the product. The most important characteristic of the CuAAC reaction is its unique bioorthogonality as neither azide nor terminal alkyne functional groups are generally present in natural systems.

References:
1 H. C. Kolb, M.G. Finn, K. B. Sharpless, Angew. Chem. Int. Ed. 2001, 40, 2004-2021.
2 C.W. Tornoe, C. Christensen, M. Meldal, J. Org. Chem. 2002, 67, 3057-3064.
3 V. V. Rostovtsev, L. G. Green, V. V. Fokin, K. B. Sharpless, Angew. Chem. 2002, 114, 2708-2711; Angew. Chem. Int. Ed. 2002, 41, 2596-2599.

The detection of cell proliferation is of utmost importance for monitoring of cell vitality, determining genotoxicity or evaluating anticancer drugs. Carl ROTH’s cell proliferation assays based on EdU (5-ethynyl-2'-deoxyuridine) provide a superior alternative to BrdU (bromodeoxyuridine) assays. Carl ROTH’s EdU Click assays are not antibody based and therefore do not require DNA denaturation for detection of the incorporated nucleoside. Instead, the EdU Click assay utilizes click chemistry for detection in a variety of dye fluorescent readouts. Furthermore, the streamlined detection protocol both reduces the total number of steps and significantly decreases the total amount of time.

References:
1 S. Vidak et al., Proliferation of progeria cells is enhanced by lamina-associated polypeptide 2α (LAP2α) through expression of extracellular matrix proteins, Genes & Development 2015, 29, 2022-2036.

EdU Click-488:
2 V. Fock et al., Trophoblast subtype-specific EGFR/ ERBB4 expression correlates with cell cycle progression and hyperplasia in complete hydatidiform moles. Human Reproduction 2015, 30, 4, 789-799.
3 K. Plessl et al., Expression pattern and function of Notch2 in different subtypes of first trimester cytotrophoblast. Placenta 2015, 36, 365-371.
4 G. Meinhardt et al., Wingless ligand 5a is a critical regulator of placental growth and survival. Scientific reports 2016, 6, 28127.
5 V. Ziegler, A. Albers, G. Fritz, Lovastatin protects keratinocytes from DNA damage-related pro-apoptotic stress responses stimulated by anticancer therapeutics. Biochimica et Biophysica Acta 2016, 1863, 1082-1092.

EdU Click-555:
6 S. Barberán, S. Fraguas and F. Cebrià, The EGFR signaling pathway controls gut progenitor differentiation during planarian regeneration and homeostasis. Development 2016, 143, 2089-2102.
7 M. Hennenberg et al., Cooperative effects of EGF, FGF, and FGF-ß1 in prostate stromal cells are different from responses to single growth factors. Life Sciences 2015, 123, 18-24.
8 D. Herrmann et al., Honokiol, a constituent of Magnolia species, inhibits adrenergic contraction of human prostate strips and induces stromal cell death. Prostate Int 2014, 2, 3, 140-146.