An examination of the structural and biological properties of three intravenous immunoglobulin preparations.
Three commercial preparations of immunoglobulin G prepared for administration by the i.v. route were tested for their physical integrity and in vitro biological activity. Size exclusion chromatography by HPLC in native and denaturing buffers together with SDS-PAGE analysis were used to determine whether covalent-bond cleavage had occurred as a result of procedures used in their preparation. C1 complement binding assays and measurements of competitive binding to an Fc receptor-bearing promonocyte cell line U937 were used to assess whether such changes had altered the capacity of these preparations to engage biological effector functions. A purified IgG1 myeloma protein was used as a reference standard. WinRho, an unmodified IgG, consisted almost wholly of monomeric IgG by HPLC size exclusion and showed no evidence of proteolytic fragments in denaturing buffers or on SDS-PAGE. Sandoglobulin, a product treated at pH 4 with pepsin, contained about 10% dimeric protein and, as revealed under denaturing conditions, about 2% fragments. Relative affinity of binding to U937 cells was similar to WinRho. C1 binding by Sandoglobulin showed normal activity with 50% inhibition at 2.8 nM. Gamimune, modified by partial reduction and alkylation, contained about 15% dimers. Between 20 and 30% of the preparation retained covalent interchain disulfides. Binding to U937 cells was two-fold weaker than the other preparations and binding to C1 was also diminished and modified. This accords well with previous reports of the deleterious effect of reduction and alkylation on Fc function.
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D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D003172:Complement C1 / Q000276:immunology; D003168:Complement Fixation Tests; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007074:Immunoglobulin G / Q000302:isolation & purification* / Q000627:therapeutic use; D007275:Injections, Intravenous; D009194:Myeloma Proteins / Q000302:isolation & purification; D011961:Receptors, Fc
D003172:Complement C1; D007074:Immunoglobulin G; D009194:Myeloma Proteins; D011961:Receptors, Fc