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Purification of Localizing Antibodies Formed against Microsomes of the N-2-Fluorenylacetamide-induced Rat Hepatoma¹

( Department of Biochemistry Research, Roswell Park Memorial Institute,³ Buffalo 3, New York)

Rabbit antibodies against the microsome fraction of the N-2-fluorenylacetamide-induced rat hepatoma were shown to localize in the tumorous livers (hepatoma and the remaining portion) and in normal livers when injected i.v. This was demonstrated by the radioiodine paired label technic. The localizing antibodies were purified by specific adsorption and elution. The purified product showed increased localization in the tumor livers and normal livers. It contained some components, presumably soluble complexes of hepatoma antigens and antibodies, that were separated from the 7 S antibody by gel filtration. The 7 S antibody, free of complexes, showed good localization. The complexes also showed some localization, but were present in too low a proportion to form an important contribution toward the total localization of the product before the gel filtration. The antiserum globulin used here was isolated by chromatography on DEAE-cellulose⁴ and was entirely 7 S -globulin. It was freed of anti-rat plasma antibodies by a solid adsorbent containing rat plasma proteins.

¹ Partly presented at the 55th Annual Meeting of the American Association for Cancer Research, Chicago, Ill., April, 1964. Supported in part by Contract AT-2651 from the U. S. Atomic Energy Commission.

⁴ The following abbreviations are used: DEAE-cellulose, diethylaminoethyl cellulose; GNS, globulin fraction from normal rabbit serum; G-anti-Hep-micro, globulin fraction from antisera to hepatoma microsomes; G-anti-rat-plasma, globulin fraction from antisera to rat plasma.

² This work was done during a leave of absence from the Department of Obstetrics and Gynecology, Osaka University Medical School, Osaka, Japan.

³ A unit of the New York State Department of Health.

Received 1/22/65.