Nylon 6 capillary-channeled polymer (C-CP) fibers used as the stationary phase in high-performance liquid chromatography protein separations have been investigated. A difference in chromatographic behavior between two (presumably identical) nylon 6 C-CP fibers, designated simply as nylon 6A and nylon 6B, reveals unknown differences in their chemical and physical nature as proteins are less retained on the latter. The possible differences are evaluated through several test methods, including Fourier transform infrared spectroscopy (FTIR), differential scanning calorimetry (DSC), mechanical testing, MALDI-TOF mass spectrometry, an organic reaction with ninhydrin, and dilute solution viscometry. Nylon 6B is shown to be a larger perimeter, stronger fiber, yet in a 200 mm, 0.8 mm i.d. column having an interstitial fraction of similar to 0.6, it has an available surface area of similar to 189 cm(2), while nylon 6A is almost double that at similar to 374 cm(2). It is revealed that when in amorphous form, the base polymers have very comparable FTIR and DSC characteristics. When in their solid, extruded fiber form there are significant differences found in crystallinity, tensile strength, molecular weight distribution, and end group density. Nylon 6A is a lower molecular weight fiber, which results in similar to 1.3x more amine end groups than nylon 6B. It is the end group density that ultimately determines the separation qualities. These results imply that both a difference in base nylon 6 molecular weight and also the fiber extrusion process have a significant influence on the performance of nylon 6 C-CP fiber separation of proteins via ion exchange chromatography. (C) 2012 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 128: 1257-1265, 2013