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The Isolation and Characterization of a Ninth Component of Hemolytic Complement, C'3f

From the Laboratories of Microbiology, Howard Hughes Medical Institute, Miami, Florida

Abstract

Experiments are described which strongly indicate that nine fractions of guinea pig serum are required for the hemolytic action of C'. The new component, termed C'3f, reacts seventh in the hemolytic sequence, i.e., after C'3e but before C'3a, and produces an intermediate complex on the surface of sheep erythrocytes which is relatively stable. C'3f is difficult to separate completely from C'2 by column chromatography. Along with C'3d, potent C'3f reactivity is found in certain fractions of bovine plasma. A method is described for the measurement of C'3f in guinea pig, canine and human sera. It is depleted from the fluid phase by treatment with EAb C'1,4,2, 3c,3b,3e. About 70% of C'3f reactivity is lost during the absorption of serum with zymosan.

Initial studies indicate that C'3f is a macromolecule with a sedimentation constant of about 5.0. It has a half-life at 56°C of about 23 min. There is no major loss of C'3f reactivity when whole serum is treated with NH₄OH, cobra venom, phlorizin or salicylaldoxime. Purified C'3f is rapidly inactivated by trypsin. The major portion of C'3f is precipitated from serum when (NH₄)₂-SO₄ is added to yield a final concentration of about 2.2 M. As yet, there are no known co-factors or inhibitors of the hemolytic reactivity of C'3f.

Footnotes

¹ Present address: Department of Bacteriology, Osaka University Medical School, Osaka, Japan.

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