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Human leukocyte antigen G (HLA-G) is a non-classical HLA class I protein with various immunosuppressive functions. Besides its profound effect to induce fetal tolerance, HLA-G has been also found to enhance graft acceptance. The aim of the study was to analyse the association between HLA-G 14 bp insertion/deletion polymorphism, soluble HLA-G level and kidney graft outcome in the Slovak population. We investigated 69 kidney transplant recipients aged 27–65 years. Out of this group, 37 recipients developed acute rejection, confirmed by biopsy, and 32 patients had stable allograft function. Plasma was obtained from recipients at 1 day before transplantation and analyzed by ELISA. Genotyping of HLA-G polymorphism was performed by PCR. Significantly higher pre-transplantation levels of sHLA-G were found in the group with stable allograft function in comparison to group with acute rejection (P = 0.0409). In the homozygous −14/−14 recipients with stable allograft function, significantly higher values of sHLA-G were determined in comparison to the recipients with acute rejection (P = 0.0052). The study revealed an association between 14 bp deletion polymorphism and soluble HLA-G level that is proportional to kidney graft acceptance. It is suggested that pre-transplantation levels of soluble HLA-G should be monitored as additional marker to predict kidney graft outcome.