Collagens and elastins are primarily synthesized by fibroblasts. These molecules are principal components of extracellular matrices. Once outside the cell, collagen assembles into fibrils and fibers that provide mechanical strength to tissues. Elastin is secreted by cells, and also forms fibers which crosslink to create a flexible network of fibers and sheets. Col-F is a low molecular weight fluorescent probe that exhibits affinity for collagen and elastin. Col-F easily penetrates between cells and into tissues where it can then bind to collagen and elastin fibers via hydrogen bonds and noncovalent hydrophobic interactions. Although Col-F has shown binding affinity for collagen and elastin, because the labeling is non-specific in nature, other tissue types may also be labeled. Analyze samples using a fluorescence microscope.
Col-F Collagen Binding Reagent
The green fluorescence probe Col-F exhibits affinity to collagen and elastin. This nontoxic collagen-binding probe provides a simple and convenient tool for fluorescence three-dimensional imaging of intricate collagenous and elastic structures in fresh and frozen animal tissues. Analyze using a fluorescence microscope.
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1. When ready to use, reconstitute the 0.5 mg Col-F vial by adding 100 µL DMSO (this creates a stock at 6.8 mM).
2. Optional: Incubate samples with blocking buffer for 60 minutes.
3. Add Col-F to the media/buffer for each appropriate sample.
4. Incubate the samples with the dye solutions for 5-60 minutes at 37°C.
5. Wash samples twice with PBS (allow samples to incubate at room temp for 5-10 minutes in PBS during each wash step).
6. Once wash steps are complete, mount a coverslip on each slide. Samples are now ready to visualize with a fluorescence microscope capable of excitation at 490 nm, and emission at 515-520 nm.
Col-F Excitation (black line) and Emission (green line) Spectra (Figure 1).
Fluorescence microscopy imaging of a frozen murine tissue section containing a blood vessel stained with 1 µM Col-F for 30 minutes. Collagen and elastin present in the blood vessel wall stained green. Nuclei were counter-stained blue with DAPI. Image was captured using a Nikon TiE Deconvolution Microscope. Data courtesy of Dr. Mike Olin, University of Minnesota (Figure 2).
For more images, please see Col-F, a fluorescent probe for ex vivo confocal imaging of collagen and elastin in animal tissues. http://onlinelibrary.wiley.com/doi/10.1002/cyto.a.22264/full
• Target: Collagen and Elastin
• Excitation/Emission: 490 nm / 515-520 nm
• Method of Analysis: Fluorescence Microscope
• Types of Samples: Cell culture and Tissue
• Storage: ≤-20°C
• Shipping: Ships overnight (domestic), International Priority Shipping
Rogozhnikov D, O’Brien PJ, Elahipanah S, Yousaf MN. Scaffold Free Bio-orthogonal Assembly of 3-Dimensional Cardiac Tissue via Cell Surface Engineering. Sci Rep. Full text.
“To observe the secretion of ECM over time for the various assembled 2D and 3D tissues, the samples were treated with Col-F fluorescent probe (Immunochemistry Technologies, MN), which has an affinity for collagen and elastin.”
Hoyle NP, Seinkmane E, Putker M, Feeney KA, Krogager TP, Chesham JE, Bray LK, Thomas JM, Dunn K, Blaikley J, O’Neil, JS. Circadian actin dynamics drive rhythmic fibroblast mobilization during wound healing. Sci Transl Med. 2017.Nov 8;9(415). pii: eaal2774. doi: 10.1126/scitranslmed.aal2774. Abstract
“Collagen deposition was measured after 14 days by staining the sections for 3 hours with Col-F (ImmunoChemistry Technologies).”