A specific subclass of IgG copurifies and interacts with human HRG^P. (A) Western blot analysis of the IgG content of human HRG^P (400 ng), with a human IgG4κ myeloma (20 ng) being included as a positive control. (B) Presence of HRG and IgG in preparations of HRG^PID (0.5 μg) and copurified IgG isolated from HRG^P (IgG^HRG; 0.5 μg) as determined by Western blot analysis. (C) Analysis of the proportion of different IgG subclasses present in IgG^HRG and pooled normal human IgG preparations as determined by human IgG subclass specific ELISA. (D) Comparison of the relative amount of κ and λ L-chains present in pooled normal human IgG and IgG^HRG as determined by densitometry analysis of κ and λ L-chains on Western blots. (E) Analysis of the ability of HRG^PID to bind to different immobilized IgG preparations by ELISA using wells precoated with 2 μg/mL IgG^HRG, human IgG2κ or human IgG1κ myeloma proteins. (F) Effect of heparan sulfate (HS) and hemin on HRG^PID binding to immobilized IgG^HRG, as measured by ELISA, with wells being precoated with 2 μg/mL IgG^HRG and then analyzed for HRG^PID (2 μg/mL) binding under the different conditions. Error bars in panels E and F represent SEM (n = 3).