Surface chemistry is a key parameter in the choice of proper materials for electrochemical detection. It has been previously shown that the presence of oxygen containing groups (OCGs) on the surface of graphene oxide (GO) can be both effective and detrimental. This poses a question when GO materials are used as electrochemical platforms for biosensing. In this work, we study how the surface chemistry of graphene oxide nanocolloids (GONCs) affects the impedimetric biosensing of ochratoxin A (OTA), in terms of immobilization of biorecognition element and detection step. OCGs on GONCs were tuned by applying increasing reduction potentials from −0.3 V to −1.2 V, resulting in GONC platforms with decreasing amounts of oxygen functionalities. It was discovered that the sensitivity of biosensing is correlated to the residual amount of OCGs on GO surface. For a more detailed investigation, three representative materials, namely unreduced GONCs, as well as GONCs reduced at potentials of −0.8 V and −1.2 V were chosen. Results were compared in terms of calibration sensitivity, selectivity and reproducibility of the impedimetric response. GONCs reduced at −1.2 V have shown the best electroanalytical response for the impedimetric detection of OTA. These findings are anticipated to contribute to the design of novel biosensors, whereby an optimized platform is employed for the immobilization of the biorecognition element.