Immune complexes determination

An affinity sorbent based on WPA-PG carrying immobilized human IgG was applied to the isolation of the first component of the complement (Cl) from human serum and for its separation into subcomponents Clr, Cls and Clq by the one-step procedure [126,127]. Cl was quantitatively bound to the sorbent at 0 °C. The activities of subcomponents Clq and Clr2r2 in the unbound part of the serum were found to be 0.8% and 3.3% of the initial activities in serum. This fraction, therefore, could be used as a R1 reagent for determining the hemolytic activity of Cl. Apparently, the neighboring macromolecules of immobilized IgG resemble to some extent an immune complex, whereas Cl formation is facilitated due to the mobility of polymer chains with the attached IgG macromolecules (Cl is usually dissociated in serum by 30%). After activation of bound Cl by heating (30 °C, 40 min) the activated subcomponent Clr is eluted from the sorbent. Stepwise elution with 0.05 mol/1 EDTA at pH 7.4 or with 0.05 mol/1 EDTA + 1 mol/1 NaCl at pH 8.5 results in a selective and quantitative elution of the activated subcomponent Cls and subcomponent Clq. 

CH50 determinations can be used to analyze the total serum complement and are useful for monitoring immune complex diseases (Sullivan, 1989) activation of complement (Table 15.13) in the presence of autoantibodies is indicative of immune complex diseases and autoimmunity. The various components of the complement system (C3, C4) can also be measured to assess the integrity of the system. For instance, low serum concentrations of C3 and C4, with a concomitant decrease in CH50 may indicate activation of complement, while a low C4 alone is a sensitive indicator of reduced activation of the complement system. Since C3 is used as an alternate complement pathway, it usually measures high. Therefore, a low C3 with a normal C4 may indicate an alternate pathway of activation. 

Several approaches can be taken to determine the function of the different neutrophil Fey receptors during binding to immune complexes ... 

Table 19.2 Sensitivity determination with analyte-specific primary antibody in the form of a dendrimer complex or a double-antibody immune complex...

N4. Nezlin, R., Alarcon-Segovia, D., and Shoenfeld, Y., Immunochemical determination of DNA in immune complexes present in the circulation of patients with systemic lupus erythematosus. J. Autoimmun. 11, 489-493 (1998). 

Criteria for Determining Whether a Disease Is Caused by Immune Complexes. 7... 

The deposition of immune complexes along vascular basement membranes and the ensuing inflammatory response are the hallmark of immune complex disease. The distribution of deposition by and large determines the clinical features of the disease. In the human, factors governing the localization of immune complexes are incompletely understood, though it is presumed that blood flow physiology plays a role. Areas of relatively high-... 

Fig. 6. Immune complexes in sera from patients with disseminated gonococcal (GC) infection. Immune complexes were run in the staphylococci binding assay. Isotype-specific reagents and an anti-GC reagent were used to detect immune complex material. Standard deviations were determined on a panel of 27 normal sera.

G20. Gupta, R. C., and Kohler, P. F., Identification of HBsAg determinants in immune complexes from hepatitis B virus-associated vasculitis. J. Immunol. 132, 1223-1228 (1984). 

Macromolecular substances with multiple antigenic determinant [Ag] can be determined by agglutination of particles which are coated with specific antibodies [Pa-Ab Ag Ab-Pa]. This method can detect immunoglobulins, human placental lactogens, a-fetoprotein, etc. The sensitivity is approximately 10 (xg/liter. Alternatively, antigen-coated particles are used these particles agglutinate with antibodies. Thus, antibodies can be determined by this system. Cambiaso et al. reported automated determination of immune complexes by their inhibitory effect on the agglutination of IgG-coated particles by rheumatoid factor or Clq (C3). 

C3. Cambiaso, C. L., Riccomi, H. A., Sindic, C. J. M., and Masson, P. L., Particle counting immunoassay (PCIA). II. Automated determination of circulating immune complexes by inhibition of the agglutination activity of rheumatoid sera. J. Immunol. Methods 23, 29-50 (1978). 

Endoh T, Yagihashi A, Sasaki M, Watanabe N. Ceftizoxime-induced hemolysis due to immune complexes case report and determination of the epitope responsible for immune complex-mediated hemolysis. Transfusion 1999 39(3) 306-9. 

The immunoglobulins have been extensively studied by ITP (in serum and csf see Section 2.4.4) and in particular the subclasses of IgG have been studied (H8, HIO, Hll, Z3). An extension of this work has been the demonstration of soluble immune complex formation in vitro (H9), which has obvious implications, particularly for the assessment of immune complex diseases. Preparative work has involved the isolation of, for example, antibodies to pig lactate dehydrogenase (B21, PI) and IgD myeloma protein (Jl). ITP has also been applied in the field of en-zymology, not for the direct measurement of enzymes as proteins, but for the determination of enzyme reaction substrates and products, and hence has been of use in enzyme kinetics. This work is summarized in Table 1. 

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