Association of activated properdin with complexes of properdin with C3.
An ELISA using antibody to properdin (P), followed by antibody to C3 to detect complexes of P with C3 (P-C3), detected low levels of P-C3 complexes in human serum and plasma samples. Incubating serum for 1 h at 37 degrees C increased the amount of P-C3 and diminished factor B hemolytic activity without altering total alternative pathway activity or C3 activity in serum. When P and C3 in incubated serum were analyzed by size exclusion HPLC, complexes of P-C3 were detected at retention times corresponding to molecular mass measuring in excess of 2 x 10(6) Da. Activation of serum with zymosan or cobra venom factor greatly increased the level of P-C3 and decreased alternative pathway hemolytic activity. Chromatography of proteins eluted from serum-treated zymosan detected a peak of P at 9.7 x 10(5) Da and a peak of P-C3 at 1.5 x 10(6) Da. Functional assays for activated properdin also revealed a peak of activity at 1.5 x 10(6) Da, congruent with the peak of P-C3. Native properdin was detected at 3.9 x 10(5) Da. When native properdin was added to properdin-depleted serum and incubated for 1 h at 37 degrees C, activated properdin was detected at the same position in the chromatograph as were P-C3 complexes. We conclude that incubation of serum at 37 degrees C produces complexes of P with C3, that exposure of serum to alternative pathway activators increases the amount of P-C3, and that generation of P-C3 complexes is associated with the presence of activated P.
Whiteman
L Y
LY
Department of Internal Medicine, Medical College of Virginia, Virginia Commonwealth University, Richmond, VA 23298.
Purkall
D B
DB
Ruddy
S
S
D002850:Chromatography, Gel; D003176:Complement C3 / Q000378:metabolism*; D003170:Complement Pathway, Alternative; D004546:Elapid Venoms / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D008970:Molecular Weight; D011414:Properdin / Q000378:metabolism*; D015054:Zymosan / Q000494:pharmacology
D003176:Complement C3; D004546:Elapid Venoms; C016310:cobra venom factor; D011414:Properdin; D015054:Zymosan