Tools For Measuring Free Zinc (pZn) In The Brain

Frederickson, Christopher

Abstract

We will deliver to basic and clinical neuroscientists the tools they need to measure """"""""free"""""""" (rapidly- exchangeable) zinc in biological fluids. As it turns out, the health of brain tissue in vivo or in vitro is just as sensitive to the free zinc concentration, or """"""""pZn,"""""""" as it is to the free proton concentration, or """"""""pH."""""""" We have developed the first, practical pZn Meter, a fluorimetric instrument, with both in-cuvette and optical fiber probe measuring modalities. Our customers will include: (i) neuroscientists interested in synaptic and intracellular zinc signaling, (ii) clinicians treating victims of stroke, head trauma, cardiac arrest, and seizures, where zinc is now understood to be one of the worst of the toxins injuring brain cells, and (iii), biologists growing or studying cells in vitro who want to monitor (and control) the pZn of their media. Collectively, these markets represent about 80,000 customers, with consumables of ~ $500/yr (scientists) to ~$10,000/yr (clinicians). In Phase I, we built prototype, fluorimetric zinc measurement tools, and we used them to establish the value for physiological pZn in the brain (7.7 +/- 0.3). In Phase II we shall capitalize on our knowledge of the physiological value of pZn and our access to proprietary zinc-sensing fluorescent molecules and: (i) complete development of a solution-based fluorimetric pZn Meter, """"""""tuned"""""""" to measure physiological pZn, (ii) develop and optimize cuvettes with the zinc-sensing fluorescent molecules attached to the walls, for """"""""semi-automatic"""""""" pZn determinations, (iii) develop an evanescent wave illumination for the cuvette, so that the fluorimetric zinc assays can be done even with """"""""dirty"""""""" solutions in the cuvette, (iv) adapt the evanescent wave+ surface assay to an optical fiber """"""""zintrode,"""""""" (v) perfect the attachment and sensing chemistry on zintrodes so that they can be used for medium-term (days-weeks) in situ recording of intracranial pZn. ? ?