Goat anti human igg antibodies

Recombinant PLA2R-Ab was generated by cloning in 293 T cells. Goat anti-human IgG antibodies (Jackson Immuno Research, USA) were labeled with Eu3+. The optimal coating concentration of PLA2R antigen was approximately 5 μg/mL. The addition of a fluorescence intensifier enhanced the original fluorescence by 1 million times and improved the sensitivity and the range of detection during TRFIA for measuring anti-PLA2R-IgG. AutoDELFIA1235 (Perkin Elmer, USA) was used to read Eu3+ fluorescence in microtiter wells.
The measurement range of anti-PLA2R-IgG by TRFIA was 0.02–340 mg/L. The intra-assay and inter-assay coefficients of variation (CV) of anti-PLA2R-IgG by TRFIA were 3.2% and 5.6%, respectively.

Cells were maintained in RPMI 1640 medium (Invitrogen, Paisley, UK) or in IMDM, supplemented with 2 mmol/L l‐glutamine, 100 U/mL penicillin, 100 μg/mL streptomycin (PAA Laboratories, Pasching, Austria), and 10% fetal calf serum (Life Technologies, Carlsbad, CA).
The following murine mAbs were raised in our laboratory: negative control antibody VIAP (against calf intestine alkaline phosphatase), 8–301 (against ct‐CD45), CD63‐11C9 (CD63), and VIT200 (CD45). Anti‐mouse CD45.2‐APC (clone 104) was obtained from eBioscience (Hatfield, UK). Goat anti‐mouse IgG and goat anti‐human IgG antibodies were purchased from Jackson Immunoresearch (Newmarket, UK). Oregon Green‐conjugated Goat anti‐mouse IgG was acquired from Life Technologies. The OKT3 (CD3) antibody for T‐cell activation was obtained from Jansen‐Cilag (Vienna), anti‐CD28 was supplied by Caltag Laboratories (Burlingame, CA).
ct‐CD45‐Ig fusion proteins (full length, D1 and D2 constructs) were generated in our laboratory 5. CTLA4‐Ig (Belatacept) was used as a control Ig fusion protein and was purchased from Bristol Myers Squibb (New York, NY). Recombinant ct‐CD45 (aa 584–1281, from Saccharomyces cerevisiae) was purchased from Merck Millipore (Darmstadt, DE). Human recombinant IL‐2 was acquired from PeproTech (London, UK).

The concentration of IgG was determined using ELISA according to the procedure described previously [8 (link)]. Goat anti-human IgG antibodies (Jackson ImmunoResearch, Europe Ltd., Ely, UK) and serum IgG standard from 0.2–12.5 ng per 100 µL (Jackson ImmunoResearch) were used for analysis.