“…In order to rule out the possibility of nonspecific coprecipitation of label, cultures of monocytes from donors of C3 allotype SS, FS, and FF, respectively, were prepared, and after 2 wk in vitro cells were incubated in the presence of radiolabeled amino acids for 5 days. The media were then harvested, dialyzed, concentrated, and subjected to agarose electrophoresis and immunofixation in the presence of carrier serum of either identical or distinct C3 allotype (15) or in the absence of carrier serum. In each instance, the radiolabeled C3 produced in vitro was of monocyte donor type (Figs.…”
A B S T R A C T Human monocytes synthesized the third component of complement (C3) up to 5 wk in vitro. Evidence for net C3 synthesis was based on (a) incorporation of 14C-labeled amino acids into C3 protein, (b) identity of the allotype of C3 produced in vitro with that of the doner's serum C3, even in the presence of carrier C3 protein of a different allotype; (c) correspondence of electrophoretic mobility, size, and subunit structure of C3 protein produced in vitro with serum C3; (d) inhibition of C3 production with cycloheximide.Monocytes from two unrelated C3-deficient patients were studied under conditions that supported C3 synthesis by normal monocytes. Serum from each of the patients contained <1% of the normal C3 concentration, but their monocytes produced C3 at -25% of the normal rate when studied after 2 wk in vitro. The C3 produced in vitro by monocytes from one of the patients had the molecular weight of normal serum C3 and dissociated appropriately under reducing conditions. Monocytes from C3-deficient patients could not be distinguished from normals on the basis of morphology, rosetting with C3-coated erythrocytes, or rates of C2, and total protein synthesis.
“…In order to rule out the possibility of nonspecific coprecipitation of label, cultures of monocytes from donors of C3 allotype SS, FS, and FF, respectively, were prepared, and after 2 wk in vitro cells were incubated in the presence of radiolabeled amino acids for 5 days. The media were then harvested, dialyzed, concentrated, and subjected to agarose electrophoresis and immunofixation in the presence of carrier serum of either identical or distinct C3 allotype (15) or in the absence of carrier serum. In each instance, the radiolabeled C3 produced in vitro was of monocyte donor type (Figs.…”
A B S T R A C T Human monocytes synthesized the third component of complement (C3) up to 5 wk in vitro. Evidence for net C3 synthesis was based on (a) incorporation of 14C-labeled amino acids into C3 protein, (b) identity of the allotype of C3 produced in vitro with that of the doner's serum C3, even in the presence of carrier C3 protein of a different allotype; (c) correspondence of electrophoretic mobility, size, and subunit structure of C3 protein produced in vitro with serum C3; (d) inhibition of C3 production with cycloheximide.Monocytes from two unrelated C3-deficient patients were studied under conditions that supported C3 synthesis by normal monocytes. Serum from each of the patients contained <1% of the normal C3 concentration, but their monocytes produced C3 at -25% of the normal rate when studied after 2 wk in vitro. The C3 produced in vitro by monocytes from one of the patients had the molecular weight of normal serum C3 and dissociated appropriately under reducing conditions. Monocytes from C3-deficient patients could not be distinguished from normals on the basis of morphology, rosetting with C3-coated erythrocytes, or rates of C2, and total protein synthesis.
“…The C3 phenotypes were characterised electrophoretically by the method of Alper and Propp (1968) with minor laboratory modification. Alpha-l-antitrypsin(Pi) was examined by starch gel electrophoresis, using the conditions described by Cook (t975).…”
“…C3 Phenotyping. Fresh mouse sera were applied to agarose gels and subjected to high-voltage electrophoresis, as described by Alper and Propp (18,19). The patterns of electrophoresis of mouse serum proteins are similar to those of human serum, and the murine bands corresponding to human CS are easily identified.…”
Two electrophoretic variants of murine complement component 3 (C3) were detected by using high-voltage electrophoresis of fresh mouse serum in agarose gels. Most of the inbred strains tested were homozygous for the S allele (for the slow-migrating variant); only four out of 46 strains had the alternative F allele (fast variant). Pen-bred Swiss-Webster animals belonged to one of three phenotypes-S. F, or SF-and the genes responsible for this variation segregated in a strictly Mendelian manner. In three such crosses, with 52 offspring, C3 segregated with H-2 in 46 instances, corresponding to a recombination frequency of -0.12.
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