In vitro cold activation of complement shown by an overestimation of totalcomplement 4: a study in patients with hepatitis C virus infection

Total complement 4 (C4) levels, when analysed on the Beckman Array nephelom eter, were found to increase in a number of serum specimens [predominantly from patients with hepatitis C virus (HCV) infection] after overnight stora ge at 4 degreesC. In order to investigate whether the phenomenon of in vitr o cold-dependent activation of complement (CDAC) was the explanation for th is increase, paired specimens were collected from 63 patients with HCV infe ction in tubes with no anticoagulant (serum) and in tubes containing EDTA ( which inhibits complement activation). C4 levels increased after overnight storage at 4 degreesC in 33 serum specimens (52%). In contrast, no increase in C4 levels was observed in any of the 63 EDTA specimens. Immunofixation of intact and activated C4 products confirmed that complement activation ha d taken place in the serum specimens in which C4 levels had increased after storage. There was a higher frequency of hepatitis C viraemia (P <0.0001), HCV antibody positivity (P <0.05) and the presence of rheumatoid factor (P <0.05) in the group of patients in whose serum samples CDAC had occurred ( n = 33) than in the other group (n = 30). As a result of our findings on C4 analysis in stored serum specimens, we would recommend potassium EDTA plas ma as the specimen of choice for complement analysis on the Beckman Array.