The association constants between C1q and C1r₂C1s₂ and between C1q and C1̄r₂C1̄s₂ were measured in solution using a new technique which employs sucrose gradient ultracentrifugation to estimate thermodynamic association constants. In this technique, zones of dilute, radioiodine-labeled C1q were sedimented through uniform concentrations of either C1r₂C1s₂ or C1̄r₂C1̄s₂. The zones remained intact, indicating that the dynamic equilibrium was rapid compared with the time of centrifugation. The observed increases in the sedimentation coefficients of the C1q zones were assumed to be directly proportional to the fraction of C1q bound in the dynamic equilibrium. Binding curves were constructed by performing the measurements at many C1r₂C1s₂ and C1̄r₂C1̄s₂ concentrations. The association constants were estimated from the midpoints of the binding curves and found to be 6.7 × 10⁷M⁻¹ for C1r₂C1s₂ binding to ¹²⁵I-C1q. After activation of the C1r₂C1s₂ the association constant decreased 10-fold to 7.1 × 10⁶M⁻¹. These association constants refer to solvent conditions of pH 7.35, 1 mM Tris, 5 mM Ca²⁺ and 150 mM NaCl, pH 7.35. Similar measurements were performed with the collagenous peptic fragment of C1q and both ¹²⁵I-C1r₂C1s₂ and ¹²⁵I-C1̄r₂C1̄s₂. The association constants were independent of the state of activation and both found to be about 2 × 10⁷M⁻¹. suggesting that most if not all of the interactions between C1q and C1r₂C1s₂ were confined to the collagenous portion of C1q.