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Fitc labeled cross reactive rabbit anti human c3c antibody

Manufactured by Agilent Technologies

The FITC-labeled cross-reactive rabbit anti-human C3c antibody is a laboratory reagent used for the detection and identification of the C3c component of the human complement system. This antibody is labeled with the fluorescent dye FITC, which allows for visualization and quantification of the target protein using appropriate detection methods.

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2 protocols using fitc labeled cross reactive rabbit anti human c3c antibody

1

Quantifying Streptococcal Complement Deposition

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Complement deposition on the streptococcal surface was assessed basically as described [10], with the differences of the use of an IgG depleted porcine anti-S. suis strain 10 serum as source of IgM and complement. Briefly, S. suis strain 10, 10∆ideSsuis, 10∆ideSsuis ∇ideSsuis_EcoRI, ∆ideSsuis∇ideSsuis_C195S were grown to an OD600 of 0.8. Then, 75µl of the respective cultures were centrifuged and incubated for 1h at 37°C on a rotator with 150 µl of a 1:2 dilution of an anti-S. suis strain 10 hyperimmune serum depleted of all IgG. Staining of C3 labeled bacteria was conducted with 250µl of a 1:150 diluted FITC-labeled cross-reactive rabbit anti-human C3c antibody (Dako, catalogue #F020102-2) for 1h at 4°C under constant rotation. As positive control, undiluted serum was used, as negative control, serum that had been heat inactivated at 56°C for 30 min was used. For samples with the addition of recombinant protein, 5µg of rIdeSsuis or rIdeSsuis_C195S were added to 10∆ideSsuis prior to incubation with serum. Samples were measured using BD, FACSFortessa and analyzed using FlowJoTM_V10 software.
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2

Streptococcal Complement Deposition Assay

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Deposition of complement on the streptococcal surface was assessed using flow cytometry assays as described previously [23 (link), 31 (link)]. Briefly, C3 deposition was investigated by incubating 2 × 106 CFU of S. suis in 50 μL of CDS for 30 min at 37 ℃ under rotation (8 rpm). As negative control CDS was incubated for 30 min at 56 ℃ to inactivate all complement factors. Staining of C3-labeled bacteria was conducted with 200 µL of a 1:150 diluted FITC-labeled cross-reactive rabbit anti-human C3c antibody (Dako, F020102-2, 3 g/L) for 1 h at 4 ℃. Samples were measured using BD FACS Fortessa and analyzed using FlowJoTM_V10 software. Results of complement binding assay are presented a fluorescence index (FI; percentage of positive bacteria multiplied by the geometric mean fluorescence intensity) in arbitrary units [32 (link), 33 (link)].
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