Background: Measurement of serum creatinine (SCr) and urine creatinine (UCr) is regularly used in clinical and research settings. For small animal experiments and for studies in which sample collection is spare (i.e. neonatal cohorts), measuring SCr and UCr using tiny amounts of sample (as low as 10 mcl) would maximize exploration and minimize iatrogenic blood loss. Methods: We performed an evaluation in six healthy adults to determine differences between SCr and UCr values in different methodologies and storage environments and time. Study was conducted using 20 mcl of sample. Analyses were done using two-way repeated measures of ANOVA. Results: Scr values showed no significant differences between LC/MS vs. Jaffe. However, the SCr using LC/MS method was lowest when measured immediately compared to other time points (F = 7.2; P< 0.001). Similarly, Jaffe measurements showed changes in the mean differences over time; however, these were not significant. UCr values were consistently higher using LC/MS than Jaffe (F = 19; P< 0.01), and UCr changed over time (F = 8.7; P < 0.02). In addition, the interaction term for method and time was also significant (F = 5.8; P < 0.04) which reflects the stability of the Jaffe measurements over time whereas the LC/MS measurements declined; especially after being frozen for 1 year (P < 0.001). Conclusion: UCr measured by Jaffe is lower than samples measured by LC/MS. UCr measurements by LC/MS vary more over time, mostly due to the sample measured after 1 year; therefore, storage of urine for more than 90 days measured by LC/MS may provide altered results. © 2014 Wiley Periodicals, Inc.