Chemically modified mesoporous silica nanoparticles (MSNs) are of interest due to their chemical and thermal stability with adjustable morphology and porosity; therefore, it was aimed to develop and compare the MCM-41 MSNs functionalised with imidazole groups (MCM-41-Im) to unmodified (MCM-41-OH) and primary amine functionalised (MCM-41-NH2) MSNs for experimental gene delivery. The results show efficient transfection of the complexes of the plasmid and either MCM-41-NH2 or MCM-41-Im. Furthermore, following transfection of HeLa cells using MCM-41-Im, an enhanced GFP expression was achieved consistent with the noticeable DNase1 protection and endosomal escape properties of MCM-41-Im using carboxyfluorescein tracer.