Candidate reference method for determining creatinine in serum: method development and interlaboratory validation.
We describe a "high-performance" liquid chromatographic (HPLC) method for accurately determining creatinine in serum. After prechromatographic precipitation of protein, we performed isocratic ion-exchange chromatography with ultraviolet detection (234 nm). Analytical results showed linearity up to 1770 mumol/L, a detection limit of 22 mumol/L, an average analytical recovery of 101%, and a CV ranging from 3% to 11%. We used certified human serum (National Institute of Standards and Technology), and additional lyophilized serum pools also assayed by definitive isotope-dilution mass spectrometry, to validate the accuracy of the HPLC method. In addition, the isocratic HPLC results showed close agreement with those obtained with a step-gradient HPLC method. We also compared the isocratic HPLC method with alkaline picrate and enzymatic methods. Our findings with samples from nonuremic, uremic, and diabetic ketoacidotic patients confirmed the positive bias previously reported with the alkaline picrate method. Interlaboratory transferability of the method was demonstrated with various commercial instruments and analytical columns. We evaluated column stability and possible interference from endogenous or exogenous compounds. On the basis of our analytical findings, we recommend the isocratic HPLC method as a candidate Reference Method for determining creatinine in serum.
Rosano
T G
TG
Department of Pathology and Laboratory Medicine, Albany Medical Center, NY 12208.
Ambrose
R T
RT
Wu
A H
AH
Swift
T A
TA
Yadegari
P
P
D002624:Chemistry, Clinical / Q000592:standards; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D003404:Creatinine / Q000097:blood*; D006801:Humans; D012015:Reference Standards
D003404:Creatinine