The present study shows that interlaboratory non-equivalence in SCr assays in the Netherlands was still substantial in 2009, notwithstanding the recent international creatinine restandardization effort. The high variability was largely explained by the ongoing use of Jaffe assays for measuring SCr. Compared to the enzymatic assays, the Jaffe assays had a significantly larger bias, especially for SCr levels in the lower range (reference value range 52–115 μmol/L). Although relative bias decreased when SCr reference values were higher, imprecision remained high. It was of course to be expected that Jaffe methods lead to a positive bias compared to the ID-/GC-MS method, and that adjustment for this bias would occur when using the appropriate MDRD equation. This has caused the downgrading of patients to a lower CKD category relevantly more often when a Jaffe technique instead of an enzymatic technique was used, especially in categories >45 ml/min/1.73 m2 (up to 79%). In contrast, the use of an enzymatic technique more often resulted in upgrading of the CKD stage, which may be explained by the differences in bias: the Jaffe technique provided higher values of SCr, whereas the enzymatic technique provided slightly lower values.
Ever since SCr is assessed in clinical practice its accuracy has been debated [1, 7, 19]. Although, SCr measurements are routinely performed, it is one of the most variable laboratory tests . The increasing use of eGFR in clinical practice has renewed the interest on the shortcomings of the SCr methodology [1, 21–24]. Since SCr is the most important variable in the renal function estimation equations, calibration of the creatinine assays is necessary to reduce bias in these formulas. This even lead to a modification of the factor used in the MDRD-equation (from 186 to 175 for IDMS traceable creatinine). However, the way this calibration was obtained has been regularly criticized in literature, due to the fact that the formulas were modified after having recalibrated the Jaffe creatinine to an IDMS traceable enzymatic method, having deleted the intercepts since these were not statistically significant.
The substantial bias and between laboratory variance as we found in our study, has been shown in various other studies in which data from proficiency testing (PT) and EQA scheme programs were evaluated [7, 25, 26]. A European trueness verification study of SCr also showed large interlaboratory variability before the matrix-based SRM 967 standard was available . In our study we would have expected a significantly reduced interlaboratory CV due to global restandardization on SRM 967. However, despite the European IVD directive with stricter regulations, no improvements compared to earlier studies, in which a method group SD of 2.6-11 μmol/L and a median CV of 5% at a SCr concentration of 74 μmol/L, had been reached . The failure to realize amelioration of interlaboratory non-equivalence is explained by the fact that standardization does not correct for analytical non-specificity problems, as is the case with the Jaffe method [28, 29]. These non-specificity problems concern the measurement of many endogenous and exogenous interfering substances such as protein, glucose and ketones when SCr is measured using a Jaffe method [28–31]. Despite many attempts to improve the performance characteristics of the Jaffe reaction, non-specificity remained . This leads to overestimation of the true SCr concentration. Calibration traceability cannot solve this problem nor substitute for improvement of suboptimal routine methods.
Although the enzymatic assay to measure SCr is not free of interference from various substances, it has a better specificity than the Jaffe technique . This was recently confirmed in a multicentre study evaluating IDMS-traceable enzymatic creatinine assays. It showed that the majority of enzymatic methods reached the acceptable total analytical error of 8% for SCr values as low as 36 μmol/L, when adequately calibrated against IDMS, improving the traceability and standardization of creatinine .
Moreover, upgrading in CKD stage as we observed when enzymatic assays were used in the MDRD formula may be less relevant in routine clinical practice than the downgrading to a lower CKD stage as occurs when a Jaffe assay was used to measure SCr. E.g. a patient whose eGFR is 57 ml/min/1.73 m2 (CKD stage 3a) or 62 ml/min/1.73 m2 (CKD stage 2) when a Jaffe respectively an enzymatic assay is used to examine SCr, probably have similar risks regarding end-stage renal disease, all-cause and cardiovascular mortality. From studies comparing the prognosis associated with the two most commonly used equations to estimate GFR (the MDRD and the Chronic Kidney Disease Epidemiology collaboration equation, CKD-EPI) during a long follow-up (≥7.5 years) we know that individuals reclassified from CKD stage 3a (eGFR 45–60 ml/min/1.73 m2) to no CKD had lower mortality risk than those not reclassified. Moreover, these participants had an equal risk to those classified as no CKD by both formulas [33, 34].
Based on the large batch of evidence in literature showing that alkaline picrate methods are inferior methods to measure creatinine, it is time for laboratories to substitute the alkaline picrate method by enzymatic methods. Moreover, if an increasing number of laboratories apply enzymatic techniques, the number of vendors of commercial enzymatic assays will increase, leading to more competition, which will ultimately reduce the costs of these assays. To bring this in a broader perspective, more accurate and precise measurements of SCr will lead to a reduction of the source of error in GFR estimates and thus errors in the staging of renal failure. Considering the number of patients that are misclassified in this study when using an alkaline picrate technique, clinical laboratories should also consider the implications for overall health costs, since patients are referred based on creatinine based estimates of GFR .
Although this study is a theoretical analysis, it is one of the few illustrating the consequences of variations in SCr measurements on GFR estimation and CKD staging for the individual patient. Since the majority of Dutch laboratories is included and we have a large heterogeneous cohort of patients in which we tested our model, we are able to give a good reflection of the consequences it might have for daily clinical practice. Moreover, we have studied creatinine values on 11 different levels against a strong reference method; and the samples used, were all recent instead of remote samples, which are frequently used in other studies. Selection bias may have occurred, since laboratories with too few analyses in the external quality control program were excluded for further analysis in our patient cohort. Moreover, we applied the MDRD formula in a patient cohort with an age range from 19–106 years. This may have introduced bias since, the MDRD has only been validated for patients from 19–70 years, and underestimates the GFR in patients >70 years. However, in clinical practice, laboratories automatically report eGFR’s each time a creatinine is measured, also in patients older than 70 years, and clinical decision making is often based on these estimates.