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What is the regulatory position of the fm value?

The FDA and EMA guidelines consider metabolism to be a significant pathway when it contributes to 25% or more of the drug’s overall elimination. When the CYP contribution is ~25% (In-vitro fm value > 25 %) or unknown, in vivo studies using...

Can I add additional matrix proteins such as laminin and collagen type IV into a RAFT Culture with the addition of some basement membrane extract such as Matrigel or Geltrex?

We have tested adding up to 5%v/v Matrigel™ into the RAFT™ Collagen Mixture. It should be noted that this requires that the absorbers be left in contact with the hydrogel for 30 minutes instead of 15 minutes (step 5 of the standard protocol). All...

What are the Storage and thaw-refreeze conditions for SilensomesTM HLM?

The Silensomes™ HLM and associated Control SilensomesTM HLM should be stored at or below -70°C. CYP450 enzyme activities in Silensomes™ HLM and associated Control Silensomes™ HLM are stable for up to 5 years when stored at -70°C and can be...

Can I add an additional layer of matrix proteins e.g. laminin or fibronectin at the end of the RAFT Process?

It is possible to add a layer of laminin or fibronectin at the end of an experiment after the RAFT™ Process has taken place. However, if cells have been embedded in the RAFT™ Culture, it is important to make sure that the culture is not left to dry,...

How can we approach the in-vitro fm value for low clearance compounds when using Silensomes HLM?

The in-vitro fm value of low clearance compounds can be measured by optimizing or extending the incubation time with certain limit. In addition, the appearance of metabolites can be measured instead of the disappearance of the parent compound.

How can I create co-cultures using the RAFT3D Cell Culture System?

You can create co-cultures in different ways, which depend on the structure of the model you are trying to make. Below are three examples of co-cultures: 1. Two cell types mixed together inside one RAFT™ Culture: For this, it is possible to mix...

What is the benefit of using Silensomes instead of the classical testing methods?

Compared to classical phenotyping protocols relying on reversible, as well irreversible, inhibitors Silensomes™ HLM offer a superior performance and high level of quality. The mechanism based inhibitors used to make Silensomes™ HLM are irreversible,...

What is the intra-plate and inter-plate reproducibility of the RAFT System?

In terms of culture thickness, the intraplate variability is 10%; while the interplate variability is 8% (data obtained from measurements on 107 separate 96-well or 24-well insert plates).

How reproducible is the fm data for Silensomes HLM?

The fm value consistency depends on consistency in the analytical method used to calculate the disappearance or appearance. If the disappearance of the test compound is lower than 5 or 10% over the incubation time such as with low clearance compound,...

How does the in-vitro fm data predict the in-vivo fm value?

The Silensomes™ HLM do not need to incorporate a Relative Activity Factor and therefore directly predict the CYP contribution to drug metabolism. In-vitro fm values calculated using Silensomes™ HLM are directly correlated to in-vivo fm values using...