Understanding the effect of physical structure and the role of interfaces is critical for gaining insight into the optoelectronic properties of conjugated polymers and their behavior in semiconductor devices such as organic light-emitting diodes and photovoltaic cells. We have developed an in situ neutron reflection measurement that allows the direct relationship between film photoluminescence and structure to be studied. In addition, we have found that by judicious deuteration of the conjugated polymers, the polymer/indium tin oxide (ITO) interface can be probed. Critically for both poly[2-(2-d17-ethylhexyloxy)-5-methoxy-1,4-phenylenevinylene] and poly[9,9′-(2-d17-ethylhexyl)-2,6-fluorene] of thickness of order 140–150 nm on ITO, we found that a thermally stable low-density layer of 20 Å thickness was present between the polymer film and the ITO. The presence of the low-contact layer means that measurements involving these two families of polymers directly deposited onto ITO may need re-evaluating, and suggests why poly(3,4-ethylenedioxythiophene) poly(styrenesulfonate) may be so beneficial for polymer light-emitting diodes.