AC impedance spectroscopy was used to investigate copper plated-through-holes (PTH) in glass fiber reinforced epoxy substrates (FR4) at different stages of temperature-humidity-bias driven electrochemical migration failure. The test vehicles consisted of PTHs arranged in an alternate anode and cathode configuration in FR4 boards. The test vehicles were subjected to accelerated stress testing [130 °C, 85 % relative humidity (RH) and 100 V DC] to initiate a specific sub-surface electrochemical failure mode referred to as conductive anodic filament (CAF) formation. The test vehicle variations included a control with no exposure, test vehicle with electrical insulation failure (resistance change to 1 MΩ during the test) and test vehicle with initiation of CAF to compare the impedance characteristics at different stages of electrochemical migration failure. AC impedance measurements were carried out on the test vehicle variations at ambient and in an in situ humidity chamber as a function of RH at 14–85 %. The prior exposure condition of the test vehicles was found to have a strong effect on the impedance characteristics at different RH values. The accelerated testing results and impedance characteristics of the PTHs in epoxy-glass fiber substrates are discussed.