Rheumatoid arthritis (RA) is an autoimmune connective tissue disease, associated 
with the presence of anti-citrullinated protein antibodies (ACPA). These 
antibodies have been found in approximately 70% of patients suffering from RA 
and they are currently used for diagnosis of RA. Although they exhibit an 
absolute need for citrulline for antibody reactivity, no precise cognate antigen 
for these antibodies has been determined. In this study, we analyzed the 
reactivity of ACPA to various citrullinated peptides by modified enzyme-linked 
immunosorbent assays, in order to determine the dependency of specific amino 
acids for antibody reactivity. A non-human protein (ovalbumin) and antigens 
directly related to RA were used as templates for synthesis of non-modified and 
citrullinated peptides, becoming potential target epitopes. Mainly peptides 
containing a Cit-Gly motif were recognized by ACPAs, while no particular amino 
acids N-terminal of citrulline were found to be essential for antibody 
reactivity. Moreover, ACPA reactivity was not restricted to antigens known to be 
associated with ACPA-positive RA alone, but also to proteins without relation to 
RA, primarily illustrating that any protein in theory can be turned into an RA 
autoantigen, by introducing Cit-Gly motifs. Knowledge about the interaction 
between ACPAs and their citrullinated targets is important for understanding 
autoimmune ACPA responses in RA, which are known to contribute to the 
pathophysiology.