The 3T3 cell line was initiated in the early 1960s by George Todaro and Howard Green from the tissue of an albino Swiss mouse embryo. The cells, which are heavily utilized in biomedical research laboratories around the world, exhibit contact inhibited motility and tend to form confluent monolayers.

Testing has established that most variants of the initial 3T3 cell line are susceptible to polyoma and simian virus 40 (SV40). The cells are known to be negative for the mousepox virus and for the enzyme reverse transcriptase, an indication that retrovirus genomes have not been integrated into 3T3 cell DNA.

The vast majority of immortal cell lines in use when the 3T3 line was established were known to induce tumors in animal models. Accordingly, it was widely assumed among the scientific community that the characteristics of immortality and oncogenicity were synonymous with one another. Studies with 3T3 cells, however, helped dispel this belief. Research with the line clearly demonstrated that though the cells can be grown indefinitely, they do not cause tumors when injected into laboratory rodents. Further studies have suggested that for cell immortalization to occur, the natural shortening of telomeres must be surmounted, but that this development is not automatically linked to a cell’s ability to instigate tumor growth.

The embryonic Swiss mouse fibroblast cell culture presented in the digital image above was immunofluorescently labeled with mouse anti-alpha-tubulin primary antibodies followed by goat anti-mouse secondary antibodies conjugated to Alexa Fluor 405. In addition, the culture was stained with Texas Red conjugated to phalloidin and SYTOX Green, targeting the filamentous actin network and nuclei, respectively. Images were recorded in grayscale with a 12-bit digital camera coupled to either a Nikon E-600 or Eclipse 80i microscope equipped with bandpass emission fluorescence filter optical blocks. During the processing stage, individual image channels were pseudocolored with RGB values corresponding to each of the fluorophore emission spectral profiles.

Distribution of Golgi Bodies and Small Nuclear Ribonucleoproteins in 3T3 Cells - The 3T3 fibroblast cell culture in this section was fixed, permeabilized, washed, and blocked with 10-percent normal goat serum in phosphate-buffered saline prior to immunofluorescent labeling with rabbit primary antibodies to giantin, a protein resident in the Golgi complex of mammalian cells, and mouse primary antibodies to fibrillarin, a component of a nucleolar small nuclear ribonucleoprotein (SnRNP).

Mouse Embryo Fibroblast Cells Labeled with Three Fluorophores and a Phallotoxin - A monolayer culture of embryonic albino Swiss mouse fibroblasts was stained with MitoTracker Red CMXRos, DAPI, and Alexa Fluor 488 conjugated to phalloidin, which target the mitochondrial network, nuclear DNA, and filamentous actin, respectively.

Targeting Histones and Peroxisomal Membrane Proteins in 3T3 Cultures with Immunofluorescence - Nuclear histone proteins were targeted in the culture of Swiss mouse fibroblasts presented in this section with mouse anti-histone (pan) monoclonal antibodies, which were imaged with goat anti-mouse Fab fragments conjugated to Alexa Fluor 568 (labeling the nucleus). The specimen was simultaneously labeled for peroxisomes with Alexa Fluor 488 conjugated to goat secondary antibodies that target rabbit anti-PMP 70.

Peroxisome Organelles and the Cytoskeletal F-Actin Network in Mouse Embryo Fibroblasts - The culture of featured 3T3 cells was labeled for peroxisomes with Rhodamine Red conjugated to goat secondary antibodies that target rabbit anti-PMP 70 (peroxisomal membrane protein 70). The culture was additionally labeled with Alexa Fluor 488 conjugated to phalloidin and Hoechst 33258, targeting filamentous actin and the nucleus, respectively.

Distribution of Tubulin, Actin, and DNA in 3T3 Cell Cultures - A culture of embryonic Swiss mouse fibroblasts was immunofluorescently labeled with primary anti-tubulin mouse monoclonal antibodies followed by goat anti-mouse Fab fragments conjugated to Cy3. The fibroblast cells were also stained with DAPI, which preferentially binds to nuclear DNA, and Alexa Fluor 488 conjugated to phalloidin to target the F-actin network.

Swiss Mouse Embryo Fibroblast Cells with MitoTracker Red CMXRos, Alexa Fluor 488, and DAPI - The embryonic Swiss mouse fibroblast cells presented in this section were resident in a culture labeled with MitoTracker Red CMXRos and Alexa Fluor 488 conjugated to phalloidin, targeting the mitochondrial network and filamentous actin, respectively. The culture was counterstained for DNA in the cell nucleus with DAPI.

Visualizing Microtubule and Actin Cytoskeletal Frameworks in Monolayer Cultures - Immunofluorescence was utilized to label a culture of 3T3 cells, which was treated with mouse anti-alpha-tubulin primary antibodies followed by goat anti-mouse secondary antibodies conjugated to the cyanine dye Cy3. In addition, F-actin in the culture was targeted with Alexa Fluor 488 conjugated to phalloidin and cell nuclei were labeled with DAPI.

Immunofluorescently Labeling 3T3 Cells for Nuclear Pore Complex Proteins - The log phase culture of 3T3 cells illustrated in this section was fixed, permeabilized, blocked with 10-percent normal goat serum, and then treated with mouse anti-NPCP (nuclear pore complex proteins) primary antibodies followed by secondary antibodies conjugated to Alexa Fluor 568. The secondary cocktail contained Alexa Fluor 488 conjugated to phalloidin to label the filamentous actin network, and the nuclei were counterstained with Hoechst 33342.

Proximity of F-actin and the Mitochondrial Network in a Culture of Mouse Fibroblasts - SYTOX Orange and Alexa Fluor 488 conjugated to phalloidin were utilized to target nuclear DNA and filamentous actin, respectively, in a culture of embryonic Swiss mouse fibroblasts. In addition, mitochondria were labeled with MitoTracker Deep Red 633.

Double Labeled Embryonic Swiss Mouse Fibroblast Cells - The mitochondrial network and DNA in the cell nucleus were targeted in a culture of 3T3 embryonic Swiss mouse fibroblast cells with the popular probes MitoTracker Red CMXRos and Hoechst 33258, respectively.