Previous work has shown that radioiodinated phospholipid ether analogs with the iodine-125 substituted on the meta position of the aromatic ring readily localized in a variety of animal tumors. In an effort to ascertain the importance of such meta substitution, three phospholipid ether analogs with the iodine-125 in the para position were synthesized for evaluation as potential tumor-localizing imaging agents. 12-(p-Iodophenyl)dodecyl phosphocholine, l-O-[12-(p-iodophenyl)dodecyl]-1,3-propanediol-3-phosphocholine, and 1-O-[12-(p-iodophenyl)dodecyl]-2-O-methyl-3-rac-glycerophosphocholine were synthesized and labeled with iodine-125 via an isotope exchange procedure. Similar to previous results with the meta substituted analogs, tissue distribution studies with the three para analogs demonstrated tumor localization and retention of radioactivity at 24 h after i.v. injection. In all three cases, the para isomers showed greater tumor avidity than the meta isomers and clearance of the radiotracer from the tumor was much slower than the clearance from nontarget tissue. 12-(p-Iodophenyl)dodecyl phosphocholine afforded the greatest tumor-to-nontarget tissue ratio. For example, the tumor-to-blood and tumor-to-liver ratios at 24 h were 10.96 and 1.85, respectively. As a result of such selective tumor retention, it was possible to clearly delineate the tumor using γ-camera scintigraphy.