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Chromatofocusing Studies Involving a Monoclonal Fab'

Veterans Administration Medical Center, San Diego, California
University of California, San Diego, California

Correspondence: For reprints contact: Samuel E. Halpern, MD, Nuclear Medicine Service (115), Veterans Administration Medical Center, 3350 La Jolla Village Dr., San Diego, CA 92161.

ABSTRACT

Isoelectric focusing (IEF) of the Fab' derivative of murine monoclonal antibody ZCE-025 is known to detect at least six bands having isoelectric points (pl) ranging from 5.4 to 7.8. Chromato focusing was employed to separate these bands. Electrophoresis of the starting materials under nonreducing conditions indicated all of the materials to migrate as Fab'. The electrophoresis of urine samples obtained from Balb/c and nude mice 8 hr after the i.v. injection of the various ¹²⁵I bands revealed the low pl bands to migrate approximately as a ¹²⁵I-Fab'. The higher pl band activity was located in lower molecular weight regions. Serum samples taken at 8 hr postinjection from the above mice revealed a series of what appeared to be high molecular weight complexes and some low molecular weight species. Biodistribution studies in comparison Balb/c mice and nude mice revealed that the low pl ¹²⁵I-Fab' bands gave an organ and tumor uptake at 8 hr very similar to Fab', while the high pl ¹²⁵I-Fab' bands were rapidly excreted into the urine and feces and did not concentrate in the tumor. The data suggest that the population of molecules making up the Fab' of this antibody is heterogeneous and variably stable. Theoretically, some of the entities observed could be counter productive to successful radioimmunoimaging. It is also possible that some of the labeled molecules are associating in vivo with endogenous proteins that might, in some Mabs, affect the biodistribution of the radiopharmaceutical.