Cell proliferation elisa brdu kit

At day 21 after immunization, splenocytes (5 × 10⁵/well) were separated from PTL- or DMSO-treated EAE mice and cultured in 96-well plates with complete medium. Cells were treated without or with different doses of the MOG_35–55 peptide at 37°C in 5% CO₂ for 72 h. The proliferation rate was measured using the colorimetric Cell Proliferation ELISA BrdU Kit (Roche, Basel, Switzerland).
At day 21 after immunization, splenocytes (5 × 10⁵/well) were separated from EAE mice and cultured in 96-well plate with complete medium. Cells were activated with MOG_35–55 peptide (20 μg/ml), cultured without or with different doses of PTL at 37°C in 5% CO₂ for 72 h. The proliferation rate was measured using the colorimetric Cell Proliferation ELISA BrdU Kit (Roche, Basel, Switzerland).
Ex vivo T-cell recall assays were performed on day 21 after immunization. Splenocytes (5×10⁵) were cultured in 96-well with 100 µl of RPMI 1640 medium with or without 20 µg/ml of peptide MOG35–55 peptide or a non-relevant peptide (ovalbumin OVA323–339; Sigma-Aldrich, Missouri, USA). After 72 h, the supernatants were collected and the IFN-γ and IL-17A were measured by using ELISA kits (BioLegend, CA, USA) following the manufacturer’s instructions.

The cells were seeded in a 96-well plate and incubated overnight. Cell proliferation was measured using a Cell Proliferation BrdU ELISA kit (Roche Diagnostics Ltd., Burgess Hill, West Sussex, UK) according to manufacturer’s instructions. Ultimately, the absorbance was recorded at 450 nm using a microplate reader (Bio-Rad Laboratories Inc., Hercules, CA, USA). The experiment was performed in triplicate.