Spontaneous C3 deposition in Crry-deficient and DAF-deficient mouse erythrocytes.

Crry-deficient mouse erythrocytes incur higher levels of spontaneous C3 deposition than DAF-deficient erythrocytes in vivo. Erythrocytes obtained from Crry/C3 knockout or DAF/C3 knockout mice were biotinylated and transfused into C57BL/6 wild-type recipients. Blood samples were taken at 5 minutes, 1 hour, and 3 hours, and erythrocytes were analyzed by 2-colored FACS, with PE staining for the transfused biotinylated knockout cells (quadrants i and ii) and FITC for C3 (quadrants ii and iv). Examples of the 2-colored FACS analysis are given in panels A and B (for DAF/C3 knockout and Crry/C3 knockout erythrocytes, respectively); examples were cells taken at 1 hour after transfusion. Percentages of the transfused knockout cells that were C3⁺ are calculated by the formula ii/(i+ii) × 100 and are given in panel C. The values for Crry/C3 knockout cells are significantly higher than those for DAF/C3 knockout cells (P < .05, Student t test). Percentages of the endogenous erythrocytes of the recipient mice that were C3⁺ are calculated by the formula iv/(iii+iv) × 100 and are given in brackets in panel C. No significant difference in such values is observed between mice that received Crry-deficient erythrocytes and mice that received DAF-deficient erythrocytes, suggesting that the difference observed with the transfused cells was specific and not an experimental artifact.