Olympus Olympus MicroSuite Software Packages .
Laser Scanning Confocal Microscopy
Confocal Microscopy Image Gallery

Raccoon Uterus Fibroblast Cells (Pl 1 UT Line)

The Pl 1 UT cell line was derived from the uterine tissue of an adult female North American raccoon (Procyon lotor). The cells exhibit fibroblast-like morphological characteristics and grow adherently to glass and polymer surfaces in culture.

Pl 1 UT Cells

Studies have shown that Pl 1 UT cells are susceptible to an array of viruses, including herpes simplex virus, reovirus 3, and vesicular stomatitis (Ogden strain). The line is, therefore, commonly utilized in the propagation of such viruses for research purposes and has been particularly useful in investigations of feline and canine viral diseases. Pl 1 UT cells are known to be resistant to infection with poliovirus 2, coxsackieviruses A9 and B5, vaccinia, and adenoviruses 2 and 5.

Fibroblasts are a type of cell predominantly found in connective tissue, where they participate in the production of the glycosaminoglycans, fibers, and glycoproteins that together comprise the collageneous extracellular matrix. The cells are also very important for their role in wound healing, fibroblasts in the area near an injury multiplying and migrating to the fissure and subsequently producing significant amounts of tropocollagen (the precursor of collagen) to promote tissue repair. Considered the least specialized of the connective tissue cells, fibroblasts are capable of differentiating into other members of their cell family, such as adipocytes (fat cells), osteocytes (bone cells), and chondrocytes (cartilage cells). The conditions that trigger such differentiation appear to be both chemical and physical in nature, some research suggesting that signal proteins, the composition of the extracellular matrix, and the shape and attachments experienced by the cells affect the stable cell state. Fibroblasts are generally regarded as the simplest cells to grow in vitro and may be incited into differentiation by careful control of culture conditions.

The adherent monolayer culture of raccoon uterus fibroblasts presented in the digital image above was stained for F-actin with Alexa Fluor 488 conjugated to phalloidin, and for the mitochondrial network with MitoTracker Red CMXRos (pseudocolored yellow). Cell nuclei were counterstained with TO-PRO-3, a monomeric cyanine dye. Images were recorded with a 60x oil immersion objective using a zoom factor of 2.0 and sequential scanning with the 488-nanometer spectral line of an argon-ion laser, the 543-nanometer line from a green helium-neon laser, and the 633-nanometer line of a red helium-neon laser. During the processing stage, individual image channels were pseudocolored with RGB values corresponding to each of the fluorophore emission spectral profiles unless otherwise noted above.

Additional Confocal Images of Raccoon Uterus Fibroblast (Pl 1 UT) Cells

Raccoon Uterus Fibroblast Cells with BODIPY FL, MitoTracker Orange CMTMRos, and DRAQ5 - The mitochondrial and cytoskeletal filamentous actin networks were targeted in the culture of Pl 1 UT fibroblasts presented in this section with BODIPY FL conjugated to phallacidin (a mushroom toxin) and MitoTracker Orange CMTMRos, respectively. Nuclei in the cells were counterstained with the far-red fluorescent DNA probe DRAQ5.

Targeting F-Actin and Mitochondria in Pl 1 UT Cell Cultures - A triple fluorophore combination of MitoTracker Red CMXRos, BODIPY FL conjugated to phallacidin, and DRAQ5 (pseudocolored cyan) was used to label an adherent log phase culture of raccoon uterus cells for mitochondria, the filamentous actin network, and nuclear DNA, respectively. The cells were first treated with the MitoTracker probe in growth medium for one hour, washed and fixed with paraformaldehyde (prepared in growth medium), permeabilized, and blocked with bovine serum albumen. The cells were subsequently labeled with the conjugated phallotoxin and counterstained with the anthraquinone (DRAQ5) reagent.

Contributing Authors

Nathan S. Claxton, Shannon H. Neaves, and Michael W. Davidson - National High Magnetic Field Laboratory, 1800 East Paul Dirac Dr., The Florida State University, Tallahassee, Florida, 32310.