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Neutrophil Elastase on the ASK


Overview


Neutrophil elastase is a key protease involved in acute lung injury, acute respiratory distress syndrome, as well as many other inflammatory processes such as emphysema, cystic fibrosis, COPD, wound healing, rheumatoid arthritis, and ischemia-reperfusion. Neutrophil Elastase 680 FAST™ is a fluorescent agent that is selectively cleaved by elastase produced by activated neutrophils during acute inflammation. Neutrophil Elastase 680 FAST is optically silent upon injection and produces fluorescent signal only after cleavage by elastase.

Neutrophil Elastase 680 FAST is a member of a family of activatable fluorescent imaging agents comprising a novel architecture, termed F.A.S.T. (Fluorescent Activatable Sensor Technology) that confers an improved pharmacokinetic profile with a broader range of early imaging time points. This architecture also offers higher target specific signal with reduced background. Neutrophil Elastase 680 FAST can be used for imaging both in vitro and in vivo.  

Neutrophil Elastase 680 FAST enables imaging of neutrophil elastase activity in applications including:

  • Acute lung injury models  
  • Acute respiratory distress syndrome 
  • Emphysema 
  • Cystic Fibrosis 
  • COPD 
  • Wound Healing 
  • Rheumatoid Arthritis 
  • Ischemia-reperfusion


Products and catalog numbers


ProductCatalog NumberEx/Em wavelength (nm)Molecular weight (g/mol)Validated ExperimentsApplicationsStorage and Stability
Neutrophil Elastase 680 FASTNEV11169675/69343,000In vivo/Ex vivoAcute inflammation
Pulmonary Diseases
Technical Data Sheet


Using Neutrophil Elastase 680 FAST in vivo/ex vivo


The generally recommended procedure for in vivo imaging with Neutrophil Elastase 680 FAST is administration via intravenous injection and imaging 4-8 hours post injection. Earlier and later time points may be appropriate for some disease models, and the optimal imaging time point for any application should be determined empirically.

View instructions on setting up an in vivo mouse experiment with Neutrophil Elastase 680 FAST and imaging on an IVIS or FMT system.

ProductRoute of InjectionMouse Dose (25 g)Rat Dose (250 g)Blood t 1/2Tissue t 1/2Optimal imaging timeOptimal Re-injection Time (complete clearance)Route of Metabolism/ background tissueFMT& IVIS settings
Neutrophil Elastase 680 FASTIV4 nmol12-40 nmol3 h12 h3-6 h2 dBladder, liver, intestinesFMT 680/700
IVIS 675/720

neutrophilelast1.jpg
Figure 1: Paw inflammation was induced with carageenan (CG) injection. CG-injected mice were measured for changes in paw thickness, vascular leak (AngioSense® 750EX), neutrophil elastase activity (Neutrophil Elastase 680 FAST), and cathepsin activity (Cat B 680 FAST) by injecting these agents at two time points, 2h (early) and 24h (late), post CG injection. Tomographic (3D) paw imaging was performed on the FMT2500 5h after agent injection. All CG-induced responses were statistically significant at both time points (p < 0.01), revealing significant recruitment of inflammatory cells.


Frozen Tissue Protocol

We have validated Neutrophil Elastase 680 FAST for use with frozen tissue samples. Here is a brief protocol with a recommended concentration of agent to use:

  1. Freeze tumor (without agent) and section 5-10 µm by cryostat.  For lung samples, mice are challenged intranasally with 100 µg of LPS followed by intranasal instillation of 200 nM fMLP in 40 uL PBS 18 h later. Harvest lungs 5 h later. For specificity, co-incubate agent with Silvelestat inhibitor.
  2. Incubate with 1 uM Neutrophil Elastase 680 FASTat 37ºC for 10-30 min for tumors; 5 h for lungs.
  3. Wash 1x with PBS. 
  4. Mount with anti-fade reagent.
  5. Fluorescence microscopy filter: Cy5.5


Application notes and posters


  • Poster: A novel NIR dye for in vivo temporal tracking of labeled macrophages to sites of acute inflammation 
  • Poster: Near-Infrared Quantitative Fluorescence Imaging of Renin Activity in Kidney Tissue


Citations


Please visit our Citations Library for references using Neutrophil Elastase 680 FAST on the IVIS or on the FMT.