This study investigates the impact of pathogenic bacterial infection on the antigen presentation pathway in immune cells. It demonstrates how infected immune cells exhibit impaired MHC-I-mediated antigen presentation compared to uninfected controls.
Start with an immune cell culture infected with pathogenic bacteria that form intracellular inclusion bodies.
The bacterial infection impairs the antigen presentation pathway in the immune cells.
Take an uninfected culture as a control.
Next, add a model protein antigen and incubate.
Immune cells internalize, process, and present the model antigen-derived peptides on MHC-I molecules.
After incubation, wash the cells to remove any unprocessed antigen.
Fix the cells, then add a quenching agent to neutralize the fixative.
Introduce the engineered CD8+ T cells and incubate.
These cells recognize the antigen fragment displayed on the MHC-I molecule, become activated, and secrete an enzyme that serves as a marker for MHC-I-mediated antigen presentation.
Next, add a substrate that reacts with the enzyme to produce a colorimetric signal.
Measure the absorbance. A reduced absorbance in the infected cell culture compared to the uninfected culture indicates impaired MHC-I antigen presentation efficiency due to bacterial infection.
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