CacoReady Frequently Asked Questions (FAQs)
Contents
1. The use of a manifold for aspiration is highly recommended Please consult the Instruction for Use provided with the product manual This is not recommended Although it has been performed and has worked well However care must be taken not to disturb the Caco 2 cells barrier during aspiration and re use The Costar reservoir plates Cat No 3395 are for use during the media exchange to help support the top plates transwells The plates with cat No 3326 can also be used for the same function however care must be taken not to damage the transwells We have no data on the genomic expression profile However we have checked different barriers markers among passages i e Lucifer Yellow TEER junctions proteins and PgP The manufacturing of CacoReady plates are carried out using cells from passages 55 to 70 The PgP is well expressed at the passages that are utilized for manufacturing of the CacoReady plates compared to other passages used Thus it is not overexpressed but is present along with certain other transporters pH ranges are 6 5 apical to 7 4 basal 11 12 13 14 Which compounds are recommended as positive and negative controls for permeability studies What protein material is recommended for use in preparation of cell culture medium Any data available for the 2 hour incubation with 5 mM D glucose added to the transport buffer How do you move from well to well without a2. 10 CacoReady Frequently Asked Questions FAQs What is the composition of the CacoReady Shipping Media What is the optimum time for liquidifying the CacoReady shipping media Can CacoReady plates be tilted or kept at an angle during aspiration Which aspiration technique is recommended for washing the plates Can LY Lucifer Yellow be performed on the same well after transport study Is Costar reservoir plate Cat No 3395 absolutely necessary for CacoReady 24 well during media exchange Are there any data available on Caco 2 cells genomic expression profile What cell passages are used for the production of CacoReady plates Is PgP overexpressed at the passages used for manufacturing the CacoReady plates What is the optimal pH for using the CacoReady plates The CacoReady shipping media is a proprietary reagent that becomes solid at room temperature and provides two functions a stabilizes the Caco 2 cells during shipping and b preserves the integrity of the cells junctions The optimal time for liquidifying the CacoReady shipping media prior to the exchange with the fresh media is between 3 to 4 hours at 37 C Leaving the CacoReady plates in the shipping media at 37 C for more than 4 hours may compromise the cells junctions Tilting or keeping the CacoReady plates in an angle during aspiration is not recommended as this action may damage the cells barrier integrity
3. llowing the electrodes to have contact with the air Has CacoReady product been validated with polyester PET membrane Propanolol and Antipyrin can be used as the positive controls and LY and or Manitol as the negative controls Inactivated Calf serum should be used for preparation of the cell culture medium Papps of reference compounds are a Digoxin 1 23x10 b Colchicin 1 15x10 It all depends on the equipment used for reading the Teer In our case the electrode does not read until it is inside the well and the read button is manually pressed The user manual recommends that the button should not be pressed while the electrode is in contact with air Only press the button when the electrode is inside the well The CacoReady has not been validated with PET membrane The main difference between the polyester membrane and polycarbonate membrane is the density of the pores Nominal Pore Density Polycarbonate PC 1x10 pores cm Polyester PET 4x10 pores cm
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