Device and method for mitochondrial membrane potential assessment

A method of making a microfluidic sensor device comprising:

• patterning a working electrode and a separate reference electrode on a substrate;
• applying a first layer of PDMS over the substrate so as to define respective openings over the working electrode and the reference electrode;
• applying a membrane selective to tetraphenylphosphonium (TPP+) ions the first layer of PDMS over the opening of the working electrode;
• applying a PVC membrane to the first layer of PDMS over the opening of the reference electrode;
• applying a second layer of PDMS over the first layer of PDMS so as to define respective openings over the membrane selective to tetraphenylphosphonium (TPP+) ions and the PVC membrane;
• and
• applying a third layer of PDMS over the second layer of PDMS, the third layer of PDMS having openings corresponding to the openings in the second layer of PDMS.

A microfluidic sensor device comprising:

• a substrate having patterned thereon at least one Ag/AgCl electrode;
• an inner chamber overlying the at least one Ag/AgCI electrode;
• a membrane selective to tetraphenylphosphonium (TPP+) ions disposed on one side of the first chamber;
• and
• a sensing chamber overlying the membrane.

A method of making a microfluidic sensor device comprising:

• patterning a plurality of contact electrodes on a substrate at different test regions;
• forming a silver/silver chloride electrode at each test region, the silver/silver chloride electrode electrically connected to at least one contact electrode;
• forming a first PDMS layer comprising a plurality of microfluidic channels containing a hole therein;
• applying the first PDMS layer to the substrate so that the silver/silver chloride electrodes are surrounded by the holes in the first PDMS layer;
• applying membranes selective to tetraphenylphosphonium (TPP+) ions to the first PDMS layer over each hole;
• and
• applying second and third PDMS layers over the first PDMS layer, wherein the second and third PDMS layers have respective holes corresponding to the locations of the holes in the first PDMS layer.

A method for analyzing mitochondrial bioenergetics comprising:

• providing a microfluidic device having first chamber containing an electrode therein and a second chamber separated from the first chamber by an ion selective membrane;
• loading the second chamber with a sample containing mitochondria;
• measuring the voltage using the electrode;
• and determining the membrane potential based at least in part on the measured voltage.

A method of for analyzing mitochondrial bioenergetics comprising:

• providing a microfluidic device having at least one chamber therein with a volume between 0 μl and about 100 μl;
• loading the at least one chamber with a sample containing mitochondria;
• and
• measuring the mitochondrial membrane potential ΔΨm of the sample.