Table 2.
The table lists the antibody pairs used in the multiplex sets, the sources of the antibodies and the standards, dilutions and assay working range. MM; mouse monoclonal antibody, RP; rabbit polyclonal antibody, HRP; horseradish peroxidase (antibodies labelled in-house), TCC; terminal complement complex; C1Inh; C1 inhibitor; FB; Factor B; FH; Factor H; FHR125; Factor H related proteins 1, 2, and 5; CR1; complement receptor 1; CRP; C-reactive protein. A kind gift from SRdC – Prof S. Rodriguez de Cordoba, Madrid. ECACC: European Collection of Authenticated Cell Cultures, Hycult: http://www.hycultbiotech.com/; CompTech: http://www.complementtech.com/; R&D Systems: http://www.rndsystems.com/.
| Assay | Capture antibody | Detection antibody | Standard | Working range (ng/ml) | Sample dilution |
|---|---|---|---|---|---|
| C1q | MM anti-C1q mAb (WL02, Hycult) | MM anti-C1q (DJ01, Hycult)-HRP | C1q (in-house purified) | 32–1000 | 1:1000 |
| C3 | RP anti human C3 (in-house) | RP anti-C3 (in-house)-HRP | C3 (CompTech) | 32–1000 | 1:16,000 |
| C4 | RP anti-C4 (in-house) | RP anti-C4 (in-house)-HRP | C4 (CompTech) | 8–500 | 1:4000 |
| C5 | RP anti-C5 (in-house) | MM anti-C5 (2D5; in-house)-HRP | C5 (in-house purified) | 32–1000 | 1:200 |
| Factor B | MM anti-FB (JC1; in house) | MM anti-FB (MBI-5; in-house)-HRP | FB (in-house purified) | 64–1000 | 1:500 |
| Factor H | MM anti-FH (OX24; ECACC) | MM anti-FH (35H9; in-house)-HRP | FH (in-house purified) | 16–1000 | 1:3000 |
| C1inh | MM anti-C1inh (in-house) | RP anti-C1inh (in-house)-HRP | C1 inhibitor (Cinryze drug) | 4–100 | 1:16,000 |
| Properdin | MM anti-properdin (1.1.1; Gift of SRdC) | MM anti-properdin (12-14-2; gift of SRdC)-HRP | Properdin (CompTech) | 7–100 | 1:400 |
| FHR125 | MM anti-FHR125 (MBI125; in-house) | RP anti-FH (in-house)-HRP | FHR125 (in-house) | 4–250 | 1:4000 |
| TCC | MM anti-TCC (aE11, Hycult) | MM anti C8 (E2, in-house)-HRP | TCC (in-house purified) | 60–1000 | 1:50 |
| CR1 | RP anti-CR1 (in-house) | MM anti-CR1 (MBI35; in-house)-HRP | CR1 (in-house purified) | 16–500 | 1:2 |
| CRP | MM anti-CRP (R&D systems; DuoSet) | MM anti-CRP-biotin/avidin-HRP (R&D systems; DuoSet) | CRP (R&D systems; DuoSet) | 0.8–50 | 1:200 |