The content of free SH groups (about 0.24) and labile S-S bonds (about 0.64) per mole human IgG can be differentially determined by the reaction with 5,5′-dithio(2,2′-dinitro)benzoate (DTNB) for 30 min and 24 h, respectively. Highly significant linear correlations were found between the number of labile S-S and the percentage of IgG₁, and the number of free SH and the percentage of IgG₂ of the total IgG fraction. It is concluded that the IgG₁ molecule contains one S-S bond which is opened during the 24 h interaction with DTNB by a disulfide exchange reaction. This was also confirmed by the investigation of pure monomeric IgG₁. The splitting of this bond does not alter molecular weight, antigenic properties or antigen binding activity, but reduces significantly the complement binding activity of IgG. On the other hand, one free SH group could be found in the IgG₂ molecule. Changes have been observed in SH and S-S levels of total IgG in patients with various malignant diseases that are to be explained by corresponding changes of the percentages of IgG₁ and IgG₂.

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A reactive disulfide bond (SS)* was detected and characterized in IgG of humans, rats and mice by virtue of disulfide interchange with dithionitrobenzoate. (SS)* was found exclusively in human IgG1 and rat IgG2b. In human IgG1 (SS)* was identified as the upper one of the two interheavy bridges in the hinge, where it appears to take part in complement activation. The biological significance of (SS)* in IgG was underlined by the fact that no other serum proteins were found to exhibit a similar reactivity.

The thiol groups of human blood serum proteins were determined after 24 hours interaction with dithionitrobenzoicacid (DTNB) to an average of 538 ± 60 µmol/l serum. After treatment of the serum with [35S]DTNB , autoradiograms of the protein elpherograms revealed two main peaks: The first with 63% of total activity, in the albumin region, corresponding to 0.60 SH/mol, the second with 23% of total activity, in the 7-globulin range, corresponding to 2.2 SH/mol. After 30 minutes incubation with D TNB , or with p-chloromercuribenzoate (CMB), in freshly prepared pools of IgG only 0.2 SH/mol were found which is the expected value already known from the literature.Autoradiograms taken from serum protein elpherograms after interaction with [UC] CMB only show the main SH-peak in the albumin range. Thus ist is concluded that the SH-peak in the γ-globulin region after 24 hours incubation with [35S]DTNB is due to one highly labile S-S-bond which easily undergoes a disulfide exchange with DTNB .