Cytomegalovirus neutralization assay using different sera measured by direct cell counting in bright field in 96-well plate

  1. The ARPE epithelial host cells are seeded in 96-well microplate and incubated for 24 hours to allow the cells to adhere
  2. The cells are then infected by cytomegalovirus and incubated with titrations of different sera for another 24 hours
  3. Finally, the cells are fixed and stained with primary anti-viral antibodies and Horse Radish Peroxidase (HRP) secondary
  4. After staining, the cells are imaged and analyzed using the Celigo image cytometer to count the number of infected cells

Bright field-based antibody neutralization assay using HRP

The example image below shows the low number of infected cells with high serum concentration in bright field using HRP

Bright field-based antibody neutralization assay - high serum

Bright field image of HRP labeled infected cells with high serum

The example image below shows the high number of infected cells with low serum concentration in bright field using HRP

Bright field-based antibody neutralization assay - low serum

Bright field image of HRP labeled infected cells with low serum

The number of infected cells are counted and dose response curves are generated for 4 different sera, where the IC50 values are calculated. The Celigo is able to output the counted results directly to EXCEL for analysis

dilution-dependent neutralization plot

Plate-view of imaged and analyzed wells, and serum dilution-dependent neutralization plot in respect to number of HRP-labeled infected cells

The Celigo Image Cytometer automates antibody neutralization assays: