Simultaneous imaging of GFP, CFP and collagen in tumors in vivo using multiphoton microscopy.
BACKGROUND
The development of multiphoton laser scanning microscopy has greatly facilitated the imaging of living tissues. However, the use of genetically encoded fluorescent proteins to distinguish different cell types in living animals has not been described at single cell resolution using multiphoton microscopy.


RESULTS
Here we describe a method for the simultaneous imaging, by multiphoton microscopy, of Green Fluorescent Protein, Cyan Fluorescent Protein and collagen in vivo in living tumors. This novel method enables: 1) the simultaneous visualization of overall cell shape and sub-cellular structures such as the plasma membrane or proteins of interest in cells inside living animals, 2) direct comparison of the behavior of single cells from different cell lines in the same microenvironment in vivo.


CONCLUSIONS
Using this multi-fluor, multiphoton technique, we demonstrate that motility and metastatic differences between carcinoma cells of differing metastatic potential can be imaged in the same animal simultaneously at sub-cellular resolution.
Sahai
Erik
E
Tumour Cell Biology Laboratory, Cancer Research UK London Research Institute, London WC2A 3PX, UK. Erik.Sahai@cancer.org.uk <Erik.Sahai@cancer.org.uk>
Wyckoff
Jeffrey
J
Philippar
Ulrike
U
Segall
Jeffrey E
JE
Gertler
Frank
F
Condeelis
John
J
D000818:Animals; D001709:Biotechnology / Q000379:methods*; D002277:Carcinoma / Q000378:metabolism / Q000473:pathology; D002462:Cell Membrane / Q000378:metabolism; D002465:Cell Movement; D003094:Collagen / Q000737:chemistry*; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000737:chemistry*; D008164:Luminescent Proteins; D051379:Mice; D008807:Mice, Inbred BALB C; D008853:Microscopy / Q000379:methods*; D008856:Microscopy, Fluorescence; D036641:Microscopy, Fluorescence, Multiphoton / Q000379:methods*; D009362:Neoplasm Metastasis; D009368:Neoplasm Transplantation; D009369:Neoplasms / Q000378:metabolism; D017785:Photons; D010957:Plasmids / Q000378:metabolism; D051381:Rats
C485182:Cyan Fluorescent Protein; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins; D003094:Collagen