Human Complement C5a PicoKine® Quick ELISA Kit

C5a is a protein fragment released from complement component C5. This gene is mapped to 9q33.2. The protein encoded by this gene is the fifth component of complement, which plays an important role in inflammatory and cell killing processes. This protein is comprised of alpha and beta polypeptide chains that are linked by a disulfide bridge. An activation peptide, C5a, which is an anaphylatoxin that possesses potent spasmogenic and chemotactic activity, is derived from the alpha polypeptide via cleavage with a convertase. The C5b macromolecular cleavage product can form a complex with the C6 complement component, and this complex is the basis for formation of the membrane attack complex, which includes additional complement components. Mutations in this gene cause complement component 5 deficiency, a disease where patients show a propensity for severe recurrent infections. Defects in this gene have also been linked to susceptibility to liver fibrosis and to rheumatoid arthritis.

Reproducibility To assay reproducibility, three samples with differing target protein concentrations were assayed using four different lots. Lots Lot 1 (pg/ml) Lot 2 (pg/ml) Lot 3 (pg/ml) Lot 4 (pg/ml) Mean (pg/ml) Standard Deviation CV (%) Sample 1 93 102 97 91 95 5.51 5.8% Sample 2 268 256 271 260 263 12.88 4.9% Sample 3 1307 992 978 1024 1075 45.15 4.2% Lots Lot 1 (pg/ml) Lot 2 (pg/ml) Lot 3 (pg/ml) Lot 4 (pg/ml) Mean (pg/ml) Standard Deviation CV (%) Lots Lot 1 (pg/ml) Lot 2 (pg/ml) Lot 3 (pg/ml) Lot 4 (pg/ml) Mean (pg/ml) Standard Deviation CV (%) Sample 1 93 102 97 91 95 5.51 5.8% Sample 1 93 102 97 91 95 5.51 5.8% Sample 2 268 256 271 260 263 12.88 4.9% Sample 2 268 256 271 260 263 12.88 4.9% Sample 3 1307 992 978 1024 1075 45.15 4.2% Sample 3 1307 992 978 1024 1075 45.15 4.2%

Intra-Assay Precision Inter-Assay Precision Sample 1 2 3 1 2 3 n 16 16 16 24 24 24 Mean (pg/ml) 93 268 1307 87 339 1075 Standard deviation 3.90 17.15 90.18 5.30 25.42 73.10 CV (%) 4.2% 6.4% 6.9% 6.1% 7.5% 6.8% Intra-Assay Precision Inter-Assay Precision Sample 1 2 3 1 2 3 n 16 16 16 24 24 24 Mean (pg/ml) 93 268 1307 87 339 1075 Standard deviation 3.90 17.15 90.18 5.30 25.42 73.10 CV (%) 4.2% 6.4% 6.9% 6.1% 7.5% 6.8% Intra-Assay Precision Inter-Assay Precision Intra-Assay Precision Inter-Assay Precision Sample 1 2 3 1 2 3 Sample 1 2 3 1 2 3 n 16 16 16 24 24 24 n 16 16 16 24 24 24 Mean (pg/ml) 93 268 1307 87 339 1075 Mean (pg/ml) 93 268 1307 87 339 1075 Standard deviation 3.90 17.15 90.18 5.30 25.42 73.10 Standard deviation 3.90 17.15 90.18 5.30 25.42 73.10 CV (%) 4.2% 6.4% 6.9% 6.1% 7.5% 6.8% CV (%) 4.2% 6.4% 6.9% 6.1% 7.5% 6.8%

Description|Quantity|Volume Anti-tag Pre-coated 96-well Strip Microplate|1|12 strips of 8 wells Standard||2 Antibody A|1|3ml Antibody B|1|3ml Sample Diluent|1|15ml TBS-T Wash Buffer (25x) |1|12ml Color Developing Reagent (TMB) |1|10ml Stop Solution|1|10ml Adhesive Plate Sealers|2|Piece

Concentration (pg/ml) 0 31.2 62.5 125 250 500 1,000 2,000 O.D. 0.043 0.106 0.201 0.426 0.810 1.346 1.580 2.024 Concentration (pg/ml) 0 31.2 62.5 125 250 500 1,000 2,000 O.D. 0.043 0.106 0.201 0.426 0.810 1.346 1.580 2.024 Concentration (pg/ml) 0 31.2 62.5 125 250 500 1,000 2,000 Concentration (pg/ml) 0 31.2 62.5 125 250 500 1,000 2,000 O.D. 0.043 0.106 0.201 0.426 0.810 1.346 1.580 2.024 O.D. 0.043 0.106 0.201 0.426 0.810 1.346 1.580 2.024

Microplate Reader capable of reading absorbance at 450nm. Automated plate washer (optional) Pipettes and pipette tips capable of precisely dispensing 0.5 μl through 1 ml volumes of aqueous solutions. Multichannel pipettes are recommended for large amount of samples. Deionized or distilled water. 500ml graduated cylinders. Test tubes for dilution. Horizontal orbital microplate shaker capable of maintaining a speed of 500 rpm, amplitude 3mm.