I am testing a model of BBB and I have detected its integrity (the ability to prevent what needs to be prevented from permeation. I want now to detect its permeability i.e. it permeats what is needed to be permeated.
I want a substance that normally passes the BBB and in the same time can be imaged by confocal or multiphoton microscopy.
I would choose test solutes that have in vivo data and represent different transendothelial pathways/mechanisms. See Garberg et al. (2004) In vitro models for the blood-brain barrier. Toxicology In Vitro 19: 299-334 & Raub et al. (2006) Early preclinical evaluation of brain exposure in support of hit identification and lead optimization. In Optimizing the ”Drug-Like” Properties of Leads in Drug Discovery, Biotechnology: Pharmaceutical Aspects Series, R.T. Borchardt, C.R. Middaugh, eds; Springer, pp. 355-410. Good luck!
Dear Karim
If you just want to test integrity, in a setup using flourescence for detection, I would use (sodium) flourescein or flourescently labelled dextrans. See this nice review by Sunders & Colleagues for inspiration
Front Neurosci. 2015 Oct 29;9:385. doi: 10.3389/fnins.2015.00385. eCollection 2015.
BR Birger
Thomas J Raub
Thanks alot. but I want a fluorescent material, that can be detected by fluorescent microscope.
Birger Brodin
Thanks for your reply but I have already tested the integrity. I want to test the permeability ( the ability to pass what needed to be passed).
in another words, something that is fluorescent and, in physiological conditions, passes the blood brain barrier.
Dear Karim
Integrity (measured in transport studies) and permeability is same-same, from the flux of a compound you calculate the permeability. (Sodium) flouresceine would pass in low rates under physiological conditions , in higher rates in certain pathologies. see also here Article Assessment of Permeability in Barrier Type of Endothelium in...
The trick is then to corrrelate the in vitro values to in vivo equivalents, permeabilities will often differ by a scaling factor
If you are looking for a compound with a high in vivo uptake, and flourescent, it may be more troublesome, since most compounds taken up into the brain would be less than 500 Da and addition of a fluorophore may increase size substantially and change physchem properties of the parent molecule.
We wold therefore normally use radiolabelled substrates or HPLC analysis for this part. Compounds could be diazepam (passive), oxycodon (carrier) and similar where in vivo data exists, as outlined by Thomas.
BR Birger
Birger Brodin
thanks a lot for the clarification. I may try sodium fluoresceine ..
you can do the permeability assay the same we did in our lab: https://refubium.fu-berlin.de/bitstream/handle/fub188/13435/Josephine_Labus.diss.pdf?sequence=1&isAllowed=y
page 24+25 :)
Sonja Häckel
thanks a lot but I want a substance that normally passes the in-vivo BBB and in the same time can be imaged by confocal or multiphoton microscopy.
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