Table 1 |.

Troubleshooting table

Step	Problem	Possible reason	Solution
1A(xi)	Patterned sizes are out of the tolerance	The exposure time and development time are not optimized	Spin SU-8 3050 on a dummy Si wafer and perform the dose test to find out the correct parameters
1A(xiv)	The etching depth did not reach the target of 430 ± 10 μm	Residual SU8 resist remained in the holes because of insufficient development time in Step1A(xii), or the etch rate is changed because of buildup of polymers in the chamber	Use a Filmetric with a smaller beam size (≤10 μm) to measure thicknesses in the holes. If SU8 remains in the holes, etch it away with oxygen plasma. Alternatively, run an oxygen plasma cleaning and then continue etching the wafer until it reaches the target depth
1A(xv)	SU8 is not etched	The plasma tool setting is not optimized to remove the hard-baked SU8	Increasing the plasma power, chamber temperature and oxygen flow rate will increase the SU8 ashing rate
1A(xix)	The alignment error is out of the tolerance	The alignment marks are contaminated, and/ or there are particles on the backside	Carefully wipe down the alignment marks using wet cleanroom papers with acetone followed by a nitrogen blow-dry until all the alignment marks can be seen clearly
1A(xxiii)	Some of the hollow microchannels are blocked or not opened yet	The etching time is not long enough, or contamination and particles exist in some holes	Etch until the clogged channels are opened. Keeping the fabrication environment clean and avoiding contamination from particle sources are important
1A(xxix)	The bore size is smaller than the target size after PECVD oxide deposition	Over-deposited SiO2 layer	Remove the coated oxide layer in hydrofluoric acid solution and redeposit the PECVD oxide layer (Step 1A(xxix)) to get the target bore size
9 and 13	The animal is agonal breathing	Anesthesia is too deep, or the flow rate is not strong enough to circulate enough air	Lower the isoflurane concentration or increase the air flow rate
20	No electric current (absence of convulsions)	Electrical issue, chip blocked or loss of contact between the chip and the skin	Check wires and connections, clean the chip through vigorous pipetting of water, use a new chip or maintain firm pressure on the chip against the skin
25A(vi)	Low gene expression	Not enough plasmid was delivered, or the plasmid was not transfected deeply enough	Increase the plasmid concentration, or increase the number of pulses or the voltage