I am planning an experiment to determine if the expression of a certain cytokine (lets say IL-6) is dependent on the expression of another cytokine (lets say IL-11) in endothelial cells (in vitro). I was thinking of using a neutralization antibody against IL-11, and than measure the expression of IL-6 using ELISA. My question is how do I confirm that the IL-11 has actually been neutralized by the antibody?
Pudiera ser en un modelo de cáncer cervical que secreta grandes cantidades de IL-6, y valorar la concentración posterior a ser tratado con tu bloquador.
Espero que te sea útil la información.
Saludos
Alejandra
Regarding the secretion of IL6, there is a model that is widely used, and which concerns the activity of IL-17A.
The IL-17A induced IL-6 secretion (or IL8) by primary cells such as synoviocytes, endothelial cell lines or tumor cell lines (CHA, GUI, VER, TUMT ...).
Induced IL-6 secretion was measured by ELISA. The action of IL17A is validated by the addition of anti-IL17A neutralizing antibodies which blocks the secretion of induced IL6.
Neutralizing antibodies act by blocking the interaction between IL17A and its receptor.
It can also act aggregating molecule 17A: in this case there can be an inhibition of secretion, but sometimes this aggregation may have the opposite effect, by potentiating the action of IL-17A, and by significantly increasing the secretion of IL6: hence the notion of neutralizing antibody versus antibody facilitating
You can do a calibration curve using one known cytokine effect on your cells as your output (in this case, IL-6). When your output does not change increasing the antibody concentration, you will be sure all soluble IL-11 is neutralized.
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