Background The laboratory diagnostics of boreliose is based on detection of specific IgG and IgM antibodies by ELISA test followed by confirmation test of Line-blot. Material and methods Testing of the presence of specific anti-Borrelia antibodies was performed in the group of 568 forest workers. The ELISA tests were performed in serum, saliva, and dry spot of blood. The confirmation tests Line blot were performed in serum then the prevalence of particular antibodies against Borrelia was calculated The ELISA test consists complete extract from Borrelia whereas the Line-blot test consists antigens from B. afzelii, B. burdorferi, and B. garinii. Results The samples from 201 (35%) forest workers have revealed the pos. results in ELISA in IgG class and 171 (30) results from this group were confirmed by Line-blot. Only in 40 (7) samples have revealed the pos. results in Line-blot but were neg. in ELISA tests. The main antigen was VIsE (77%) in IgG class. It has been shown that 119 (21%) samples have revealed pos. results in the IgM class in ELISA tests, and from this group 59 (10%) cases were confirmed by Line-blot. Antibody against antigen OspC revealed the highest prevalence (71%) in IgM class. The correlation between samples from sera and mucosal transudate have shown r = 0.736 in the case of IgG and r = 0.162 in the case of IgM. Conclusion The frequency, of anti-Borrelia antibodies occurrence was very high and appeared in 42% of forest workers. It has revealed that the correlation of results between serum and mucosal transudate was not satisfactory in the case of ELISA test. In contrary, the correlation of results between serum and dry spot of blood was very high and should be consider as alternative material for boreliose diagnosis.