Antibody-Dependent Cellular Phagocytosis Assay

For the ADCP assays^{41 (link),42 (link)}, biotinylated antigens were coupled to FluoSphere NeutrAvidin beads (ThermoFisher). Immune complexes were formed by incubating 10 ul of antigen coupled beads with 10 ul of 1:100 diluted plasma samples at 37 °C for 2 hours. THP-1 monocytes (25,000 cells per well) were then added to the beads then incubation at 37 °C for 16 hours. Following incubation, cells were fixed with 4% para-formaldehyde and analyzed on an iQue analyzer (Intellicyt).

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Bartsch Y.C., Chen J.W., Kang J., Burns M.D., St Denis K.J., Sheehan M.L., Davis J.P., Edlow A.G., Balazs A.B., Yonker L.M, & Alter G. (2022). BNT162b2 induces robust cross-variant SARS-CoV-2 immunity in children. NPJ Vaccines, 7, 158.

Publication 2022

FluoSphere NeutrAvidin beads iQue analyzer

Antigen Cells Formaldehyde Immune complexes Monocytes Neutravidin Plasma

Corresponding Organization :

Other organizations : Ragon Institute of MGH, MIT and Harvard, Massachusetts General Hospital, Reproductive Science Center

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Variable analysis

independent variables

Biotinylated antigens coupled to FluoSphere NeutrAvidin beads (ThermoFisher)
Plasma samples diluted at 1:100

dependent variables

Immune complexes formed between antigen coupled beads and plasma samples
THP-1 monocyte activation after incubation with immune complexes

control variables

Incubation temperature of 37 °C
Incubation time of 2 hours for immune complex formation
Incubation time of 16 hours for THP-1 monocyte activation
THP-1 monocyte seeding density of 25,000 cells per well
4% para-formaldehyde used for cell fixation

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

Annotations

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