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Invitrogen™ EnzChek™ Phospholipase A2 Assay Kit
Description
Includes
1 kit
This kits takes the stand alone assay for phospholipase A2, PLA2 and combines the necessary Ez and lipids to run two complete 96-well microplate assays that monitor activity in purified enzyme preparations and cell lysates. The EnzChek Phospholipase A2 Kit provides enough reagents for two microplates, using 200μL volumes in 96-well format to perform continuous fluorometric monitoring of PLA2 . This product offers an alternative to (B7701), (bis-BODIPY FL C11-PC) reagent, by providing an PLA2 selective substrate and one that is ratiometric, thereby lowering variations produced by instrumentation and assay conditions. Phospholipase A2 or PLA2 represents a family of enzymes that hydrolyze the sn-2 ester linkage of phospholipids. The activities of these enzymes play important roles in cardiovascular, inflammatory and nervous system disorders, and in cancers. The EnzChek Phospholipase A2 substrate provides sensitive and continuous rapid real-time monitoring of PLA2 enzyme activities. This unique substrate is selective for PLA2 and can be used either in a intensity or ratiometric based detection mode. In intensity based detection mode the PLA2 activity is monitored by the intensity increase of a single wavelength at approximately 515nm. In ratiometric analysis, which is based on the distinct fluorescence resonance energy transfer (FRET) emission of this substrate prior to and after cleavage, PLA2 is detected by changes in the emission intensity ratio at 515/575nm with excitation at ≈ 460nm. Either detection mode provides a simple method with low background, high sensitivity and high specificity for PLA2.
Enzyme and Protein Activity Assays, Lipase and Phopholipase Activity, Proteins, Expression, Isolation and Analysis
Order Info
Shipping Condition: Room temperature
Specifications
Specifications
| Color | Mixture, Green |
| Content And Storage | 1 kit. Store kit at -<20°, protected from light, desiccated |
| Detection Method | Fluorescence Intensity |
| For Use With (Application) | Phospholipase Assay |
| For Use With (Equipment) | Microplate Reader |
| Product Type | EnzChek Assay Kit |
| Dye Type | BODIPY™ FL |
| Emission | Ratiometric: 460 to 490/515, 575, Intensity-based: 460 to 490/515 |
| Filter Type | FITC (Fluorescein) Filter |
| Product Line | EnzChek |
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Frequently Asked Questions (FAQs)
The EnzChek Phospholipase A2 Assay Kit (Cat. No. E10217) can detect PLA2 at 0.05 U/mL or lower.
There are numerous factors that can interfere with FRET-based detection. Here are a few items to consider:
a) Is there any component in the reaction solution that can quench green fluorescence?
b) Is there any component in the reaction solution that autofluoresces in the green or red emission range?
c) Too high of a substrate concentration may result in diffusional FRET: Lower the final working concentration of the substrate.
d) Ensure adequate dispersal of the substrate during the reaction. BODIPY dyes are very hydrophobic, and aggregates of the green product, BODIPY FL, can form an excimer and emit in the red range.
Yes, phospholipase A2 from other species may be used as long as the phospholipase A2 can hydrolyze the sn-2 ester linkage of phospholipids and the enzymatic reaction can be done within a pH of >3 and <9.
Safety and Handling
For Research Use Only. Not for use in diagnostic procedures.
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