The aim was to evaluate if the Dried Blood Spot (DBS)-technique can be used to analyse C-peptide. S-C-peptide and paired whole blood clotted on filters, dried, punched out and eluted were sampled from 198 healthy subjects. Six subjects with S-C-peptide values outside the reference range were excluded. A conversion formula using log-DBS-C-peptide was generated in a subset of 156 (∼80%) subjects with predictions made using also storage time (eluates) and age of subjects: (log S-C-peptide = 1.696 + 1.367 log DBS-C-peptide + 0.058 (storage time/month) + 0.014 (age/10 years). This formula was cross validated into the original population. Using Bland-Altman plots, mean difference between converted log DBS-C-peptide and log S-C-peptide at baseline was 0 and limits of agreements were -0.18 to +0.18. Mean difference between converted log DBS-C-peptide values after six months and log S-C-peptide value from baseline was -0.01 and limits of agreement were -0.20-0.19. The lowest value detected with the DBS-technique corresponded to serum C-peptide 0.44 nmol/L. We concluded that DBS-C-peptide can be used as a first line screening test to monitor normal beta cell function. C-peptide on filters remained stable for six months.