Analytical performance of an immunoassay to measure proenkephalin

Clin Biochem. 2018 Aug:58:72-77. doi: 10.1016/j.clinbiochem.2018.05.010. Epub 2018 May 18.


Background: Endogenous opioids, enkephalins, are known to increase with acute kidney injury. Since the mature pentapeptides are unstable, we evaluated the performance of an assay that measures proenkephalin 119-159 (PENK), a stable peptide formed concomitantly with mature enkephalins.

Methods: PENK assay performance was evaluated on two microtiterplate/chemiluminescence sandwich immunoassay formats that required 18 or 1 h incubation times. PENK concentration was measured in plasma from healthy individuals to establish a reference interval and in patients with varied levels of kidney function and comorbidities to assess the association with measured glomerular filtration rate (mGFR) using iothalamate clearance.

Results: Assay performance characteristics in plasma were similar between the assay formats. Limit of quantitation was 26.0 pmol/L (CV = 20%) for the 1 h assay and 17.3 pmol/L (CV = 3%) for the 18 h assay. Measurable ranges were 26-1540 pmol/L (1 h assay) and 18-2300 pmol/L (18 h assay). PENK concentrations are stable in plasma stored ambient to 10 days, refrigerated to at least 15 days, and frozen to at least 90 days. Results were comparable in paired SST serum and EDTA plasma. Age and sex were not associated with PENK concentrations in healthy individuals (reference interval: 36-97.5 pmol/L). Plasma PENK concentration correlated with mGFR. In a multivariate model PENK concentration, age, sex and transplant status were significant predictors of mGFR, and 49% of predicted GFR values fell within 30% of the mGFR.

Conclusions: Both assay formats are accurate and precise for measuring clinically relevant PENK concentrations. The association of PENK concentration with mGFR is influenced by gender, age, and history of kidney transplantation. Future studies will determine if blood PENK can be used clinically to estimate GFR and/or detect AKI.

Keywords: Assay validation; Biomarkers; Enkephalin; Pro-enkephalin; Proenkephalin; penKid.

MeSH terms

  • Animals
  • Antibodies, Monoclonal, Murine-Derived / chemistry*
  • Enkephalins / blood*
  • Enzyme-Linked Immunosorbent Assay / methods
  • Female
  • Humans
  • Male
  • Mice
  • Protein Precursors / blood*
  • Sensitivity and Specificity


  • Antibodies, Monoclonal, Murine-Derived
  • Enkephalins
  • Protein Precursors
  • proenkephalin