Molecular Probes™ Click-iT™ Plus EdU Alexa Fluor™ 488 Flow Cytometry Assay Kit

Click-iT™ Plus EdU Alexa Fluor™ 488 Flow Cytometry Assay Kit

Brand:  Molecular Probes™ C10632

Product Code. 15267156

Description

The Click-iT™ Plus EdU Alexa Fluor™ 488 Flow Cytometry Assay Kit provides a simplified, more robust assay for analyzing DNA replication in proliferating cells compared to traditional BrdU methods. Newly synthesized DNA is analyzed using the 488 nm laser of the flow cytometer. The Click-iT™ Plus formulation is compatible with standard fluorophores, including R-PE and R-PE tandems, as well as fluorescent proteins.

Multiplexable—compatible with R-PE (and tandems) and fluorescent proteins

Accurate—superior results compared to BrdU assays

Fast—results in as little as 60 minutes

View selection guide for all Click-iT™ EdU and Click-iT™ Plus EdU assays for flow cytometry.

Multiplexable
The Click-iT™ Plus formulation provides increased multiplexibility compared to the original Click-iT™ EdU Flow Cytometry assays. Click-iT™ Plus EdU assays can be used in conjunction with R-PE and R-PE tandems, as well as fluorescent proteins such as GFP and mCherry, without loss of the accuracy or speed of the original Click-iT™ EdU assay.

An Advanced Method Giving You Results Superior to BrdU
The most accurate method of proliferation analysis is direct measurement of DNA synthesis. Originally, this was performed through incorporation of radioactive nucleosides. This method was replaced by antibody-based detection of the nucleoside analog bromodeoxyuridine (BrdU). The Click-iT™ Plus EdU Flow Cytometry assay is a novel alternative to the BrdU assay. EdU (5-ethynyl-2´-deoxyuridine) is a thymidine analog that is incorporated into DNA during active DNA synthesis. Detection is based on click chemistry, which is a copper-catalyzed covalent reaction between an azide and an alkyne. In this application, the alkyne is found in the ethynyl moiety of EdU, while the azide is coupled to the Alexa Fluor™ dye. Standard flow cytometry methods are used for determining the percentage of S-phase cells in the population.

Mild Conditions Allow Use with Cell Cycle Dyes and Antibodies
The small size of the dye azide allows for efficient detection of the incorporated EdU using mild conditions, while standard aldehyde-based fixation and detergent permeabilization are sufficient for the Click-iT™ Plus detection reagent to gain access to the DNA. This is in contrast to BrdU assays that require DNA denaturation (using HCl, heat, or digestion with DNase) to expose the BrdU so that it can be detected with an anti-BrdU antibody. Sample processing for the BrdU assay can result in signal alteration of the cell cycle distribution, as well as destruction of antigen recognition sites when using the HCl method. In contrast, the easy-to-use Click-iT™ Plus EdU assay is compatible with cell cycle dyes. The Click-iT™ Plus EdU assay can also be multiplexed with antibodies against surface and intracellular markers, as well as conjugates labeled with standard fluorophores including R-PE, R-PE tandems, and fluorescent proteins (GFP and mCherry).

Quick and Simple Protocol
The Click-iT™ Plus EdU protocol is based on the aldehyde fixation and detergent permeabilization steps for immunohistochemical antibody labeling. However, EdU is compatible with other fixation/permeabilization agents including saponin and methanol. In just five steps you’ll be ready to analyze your cell proliferation data:

1. Treat cells with EdU.
2. Fix and permeabilize cells.
3. Detect S-phase cells with Click-iT™ Plus detection cocktail for 30 min.
4. Wash once.
5. Analyze.

Results can be seen in as little as 60 minutes in some circumstances, but we recommend 90 minutes for all applications.

Specifications

• Content And Storage: Contains EdU (5-ethynyl-2' -deoxyuridine), Alexa Fluor™ 488 picolyl azide, anhydrous dimethylsulfoxide (DMSO), Click-iT™ fixative, Click-iT™ saponin-based permeabilization and wash buffer, Copper Protectant, and Click-iT™ EdU buffer additive.
• Store at 2°C to 8°C
Dessicate and protect from light.
• For Use With (Equipment): Flow Cytometer
• Dye Type: Alexa Fluor™ 488
• Format: Tube(s)
• Product Line: Click-iT™
• Shipping Condition: Room Temperature

• Detection Method: Fluorescence
• Product Type: Flow Cytometry Assay Kit
• Emission: 488
• Green Features: Less hazardous
• Quantity: 50

Safety and Handling

Product Identifier

• Click-iT Plus EdU Alexa Fluor 488 Flow Cytometry Assay Kit

Signal Word

• Danger

Hazard Category

• LONG-TERM AQUATIC HAZARD Chronic 2
• Carcinogenicity Category 1
• Serious eye damage/eye irritation Category 1
• Germ cell mutagenicity Category 1B
• Germ cell mutagenicity Category 2
• Reproductive toxicity Category 2
• Skin sensitiser Category 1
• Specific target organ toxicity Category 2

Hazard Statement

• H317-May cause an allergic skin reaction.
• H318-Causes serious eye damage.
• H340-May cause genetic defects.
• H341-Suspected of causing genetic defects.
• H350-May cause cancer.
• H361-Suspected of damaging fertility or the unborn child.
• H371-May cause damage to organs.
• H412-Harmful to aquatic life with long lasting effects.

Precautionary Statement

• P201-Obtain special instructions before use.
• P202-Do not handle until all safety precautions have been read and understood.
• P261-Avoid breathing dust/fume/gas/mist/vapours/spray.
• P264-Wash hands thoroughly after handling.
• P270-Do not eat, drink or smoke when using this product.
• P273-Avoid release to the environment.
• P280-Wear protective gloves/protective clothing/eye protection/face protection.
• P302+P352-IF ON SKIN: Wash with plenty of water/soap.
• P305+P351+P338-IF IN EYES: Rinse cautiously with water for several minutes. Remove contact lenses, if present and easy to do. Continue rinsing.
• P308+P313-IF exposed or concerned: Get medical advice/attention.
• P310-Immediately call a POISON CENTER/doctor/.
• P332+P313-If skin irritation occurs: Get medical advice/attention.
• P391-Collect spillage.
• P501b-Dispose of contents/container in accordance with local/regional/national/international regulations.

Supplemental information

• MIXTURE LIST-Contains : 5-ethynyl-2idine, Paraformaldehyde, Sulfuric acid,copper(2+) salt

For Research Use Only. Not for use in diagnostic procedures.