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Recent advances in the fabrication and classification of amorphous carbon (a-Carbon) thin films play an active part in the field of surface materials science. In this paper, a pulsed laser deposition (PLD) technique through controlling experimental parameters, including deposition time/temperature and laser energy/frequency, has been employed to examine the substrate effect of amorphous carbon thin film fabrication over SiO2 and glass substrates. In this paper, we have examined the structural and magnetoresistance (MR) properties of these thin films. The intensity ratio of the G-band and D-band (ID/IG) were 1.1 and 2.4, where the C(sp2) atomic ratio for the thin films samples that were prepared on glass and SiO2 substrates, were observed as 65% and 85%, respectively. The MR properties were examined under a magnetic field ranging from −9 T to 9 T within a 2-K to 40-K temperature range. A positive MR value of 15% was examined at a low temperature of 2 K for the thin films grown on SiO2 substrate at a growth temperature of 400 °C using a 300 mJ/pulse laser frequency. The structural changes may tune the magnetoresistance properties of these a-Carbon materials. These results were demonstrated to be highly promising for carbon-based spintronics and magnetic sensors.