Neurofilament Light Chain and Glial Fibrillary Acidic Protein as early prognostic biomarkers after out-of-hospital cardiac arrest.

AIMS: Neurofilament Light Chain (NfL) and Glial Fibrillary Acidic Protein (GFAP) are proteins released into the bloodstream upon hypoxic brain injury. We evaluated the biokinetics and examined the prognostic performance of serum NfL and GFAP in comatose out-of-hospital cardiac arrest (OHCA) patients. Furthermore, we compared the prognostic performance to that of serum Neuron Specific Enolase (NSE). METHODS: This is a sub-study of the "Targeted temperature management for 48 vs 24 hours" (NCT01689077) trial. NfL and GFAP serum values from 82 patients were examined in blood samples collected at 24, 48 and 72 hours (h) after reaching target temperature of 33 ± 1 °C. This temperature was reached within a median of 281-320 minutes after intensive care unit admission. GFAP was analysed at 48 and 72 h. The neuroprognostic performance of NfL and GFAP was evaluated after 6 months follow-up. RESULTS: NfL and GFAP values were significantly higher in patients with a poor outcome (Cerebral Performance Category (CPC) score 3-5) vs. good outcome (CPC 1-2). NfL 24 h: 1371.5 (462.0; 2125.1) vs. 24.8 (14.0; 61.6). GFAP 48 h: 1285.3 (843.9; 2236.7) vs. 361.2 (200.4; 665.6) (both p < 0.001). Both biomarkers were promising markers of poor functional outcome at 24 and 48 h respectively: NfL 24 h: AUROC 0.95 (95% CI: 0.91-1.00). GFAP 48 h: AUROC 0.88 (95% CI: 0.81-0.96). NfL and GFAP both predicted outcome better than NSE at 48 h (both p < 0.01). At 72 h NfL but not GFAP outperformed NSE (p = 0.01). CONCLUSION: Serum NfL and GFAP may be strong biomarkers of poor functional outcome after OHCA from an early timepoint.

Neurofilament light chain: serum reference intervals in Danish children aged 0-17 years.

Elevated levels of neurofilament light chain (NfL) in the blood is an unspecific biomarker for damage to neuronal axons. The measurement of NfL levels in the blood can provide useful information for monitoring and prognostication of various neurological disorders in children, but a reference interval (RI) is needed before the clinical implementation of the biomarker. We aimed to establish a RI for children aged 0-17 years. Serum samples from 292 healthy reference subjects aged 0.4-17.9 years were analysed by a single-molecule array (Simoa®) established for routine clinical use. Non-parametric quantile regression was used to model a continuous RI, and a traditional age-partitioned non-parametric RI was established according to Clinical and Laboratory Standard Institute (CLSI) guideline C28-A3. Furthermore, we investigated the effect of hemolysis on assay performance. The traditional age-partitioned non-parametric RI for the age group <3 years was 3.5-16.6 ng/L and 2.1-13.9 ng/L in the age group ≥3 years, respectively. The continuous RI showed an age-dependent decrease in median NfL levels in the first three years of life which was also evident in the age-partitioning of the traditional RI. We found no difference between sexes and no impact of hemolysis on the NfL test results. This study establishes a pediatric RI for serum NfL and lays the groundwork for its future use in clinical practice.

Serum GFAP - pediatric reference interval in a cohort of Danish children.

OBJECTIVES: Glial fibrillary acidic protein (GFAP) in blood is an emerging biomarker of brain injury and neurological disease. Its clinical use in children is limited by the lack of a reference interval (RI). Thus, the aim of the present study was to establish an age-dependent continuous RI for serum GFAP in children. METHODS: Excess serum from routine allergy testing of 391 children, 0.4-17.9 years of age, was measured by a single-molecule array (Simoa) assay. A continuous RI was modelled using non-parametric quantile regression and presented both graphically and tabulated as discrete one-year RIs based on point estimates from the model. RESULTS: Serum GFAP showed a strong age-dependency with declining levels and variability from infants to adolescents. The estimated median level decreased 66 % from four months to five years of age and another 65 % from five years to 17.9 years of age. No gender difference was observed. CONCLUSIONS: The study establishes an age-dependent RI for serum GFAP in children showing high levels and variability in the first years of life.

Instability of uracil in whole blood might affect cancer treatment with fluoropyrimidines.

BACKGROUND AND AIMS: Measurement of plasma uracil is used before cancer treatment with fluoropyrimidines to determine if patients tolerate a full dose. Incorrect preanalytical handling may cause falsely elevated concentration and result in suboptimal cancer treatment. We aimed to examine the stability of uracil in whole blood stored at room temperature (RT) and the effect of centrifugation temperature. MATERIALS AND METHODS: EDTA tubes (6x4 mL) were collected from 25 healthy volunteers. Five samples were stored 0, 1.5, 2, 3, and 4 h at RT and centrifuged at 4 °C. The sixth sample was centrifuged at RT after 1.5 h. Uracil was measured using an in-house LC-MS/MS method. RESULTS: Storage of whole blood at RT followed by centrifugation at 4 °C caused a rapid increase in uracil concentration. Already after 1.5 h, the mean change (20.5 % (95 % CI: 11.9-29.2 %)) exceeded the maximum permissible difference. Centrifugation at RT instead of 4 °C after 1.5 h resulted in a smaller increase (7.0 % (95 % CI: 0.7-13.4 %)), although not statistically significant (p = 0.0527). CONCLUSION: Uracil was unstable in samples processed according to current recommendations. Our data indicates better stability when centrifugation is performed at RT compared with 4 °C but further research into this is necessary.

Serum GFAP - reference interval and preanalytical properties in Danish adults.

OBJECTIVES: Glial fibrillary acidic protein (GFAP) is a promising biomarker that could potentially contribute to diagnosis and prognosis in neurological diseases. The biomarker is approaching clinical use but the reference interval for serum GFAP remains to be established, and knowledge about the effect of preanalytical factors is also limited. METHODS: Serum samples from 371 apparently healthy reference subjects, 21-90 years of age, were measured by a single-molecule array (Simoa) assay. Continuous reference intervals were modelled using non-parametric quantile regression and compared with traditional age-partitioned non-parametric reference intervals established according to the Clinical and Laboratory Standards Institute (CLSI) guideline C28-A3. The following preanalytical conditions were also examined: stability in whole blood at room temperature (RT), stability in serum at RT and -20 °C, repeated freeze-thaw cycles, and haemolysis. RESULTS: The continuous reference interval showed good overall agreement with the traditional age-partitioned reference intervals of 25-136 ng/L, 34-242 ng/L, and 5-438 ng/L for the age groups 20-39, 40-64, and 65-90 years, respectively. Both types of reference intervals showed increasing levels and variability of serum GFAP with age. In the preanalytical tests, the mean changes from baseline were 2.3% (95% CI: -2.4%, 6.9%) in whole blood after 9 h at RT, 3.1% (95% CI: -4.5%, 10.7%) in serum after 7 days at RT, 10.4% (95% CI: -6.0%, 26.8%) in serum after 133 days at -20 °C, and 10.4% (95% CI: 9.5%, 11.4%) after three freeze-thaw cycles. CONCLUSIONS: The study establishes age-dependent reference ranges for serum GFAP in adults and demonstrates overall good stability of the biomarker.