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The following message was posted to: PharmPK
I need some advice concerning Caco-2 experiments. We use a Caco-2 kit
BD (3-day model) to profile compounds for absorption characteristics.
To check the quality of the cell layer we use TEER measurements (all
before we load the wells with test compounds, we have one well reserved
lucifer yellow as layer integrity test and after the permeability
experiment we check all layers with lucifer yello again. So far so good.
However, I have the impression, that the well-to-well reproducibility is
rather high (0.8 to 1.3 E-6 cm/sec for lucifer yellow). From literature
know that the 3-day model is much "leakier" than the other models.
Nevertheless, we want to keep up with this model, since it requires much
less time resources.
Now my question: Is it advisable to incubate the unknown compounds
with lucifer yellow as internal reference standard
is this a bad idea? Does anybody uses internal reference standards, and
yes, what compounds do you use?
(We cannot assess radioactive labelled componds).
All suggestions are wellcome!
Thank you very much in advance!
gmunder str. 37-37a
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Although we have not used the 3-day model, when we run the 21-day Caco
analyses we routinely mix the test compound with lucifer yellow as an
internal control. This has proven to be beneficial several times in
determining if the permeability data should be trusted and if a
compound is potentially toxic to the cells thereby giving a false high
permeability. The addition of lucifier yellow to control compounds
does not seem to affect the overall interpretation of their cell
I hope this helps.
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