Sepsis, a life-threatening organ dysfunction triggered by a dysregulated host response to infection [1], remains a global challenge with significant morbidity and mortality [2]. Early detection and prompt treatment of high-risk septic patients are crucial for improving outcomes [3].

Presepsin, a fragment of the CD14 protein identified as a soluble subtype, emerges as a promising biomarker for sepsis diagnosis and prognosis [4-7]. Secreted by lipopolysaccharide-recognizing activated monocytes, presepsin offers advantages over existing markers like procalcitonin and C-reactive protein, reportedly releasing earlier and reaching peak concentrations faster [4, 8, 9]. Presepsin’s potential as a prognostic marker has recently gained traction, particularly for coronavirus disease-2019 (COVID-19) patients. Studies have demonstrated its ability to enhance outcome prediction when combined with clinical indices, exceeding the accuracy of clinical indices alone [10]. Presepsin levels in moderate COVID-19 cases were significantly higher than in mild cases, suggesting its value in guiding treatment and prognosis decisions [11]. Moreover, presepsin levels have shown promise in predicting septic acute kidney injury [12].

Rapid turnaround times are crucial for sepsis diagnostic biomarkers. However, situations arise where limited laboratory resources necessitate test transfer to reference laboratories. In such cases, the sample stability during transport for Sysmex HISCL-5000 presepsin testing becomes a critical factor. This study aims to evaluate the stability of samples under such transport conditions.

Presepsin’s potential extends beyond sepsis diagnosis, demonstrating promise as a prognostic marker in diverse patient populations. For example, a study of 343 patients undergoing transcatheter aortic valve implantation found a high baseline presepsin level to be an independent predictor of one-year mortality, with an odds ratio of 2.2 [13]. Similarly, increased presepsin concentrations were reported to be associated with 30-day in-hospital mortality and disease severity in 211 emergency department patients with community-acquired pneumonia [14]. Moreover, a prospective cohort study of 108 gastric cancer patients revealed that presepsin levels on postoperative days 3, 5, and 7 served as more reliable predictors of postoperative infectious complications to C-reactive protein, white blood cell, and neutrophil levels [15].

Limited laboratory resources in healthcare facilities often require specimen transfer to referral laboratories. This study, to the best of our knowledge, is the first study to specifically evaluate sample stability for HISCL presepsin testing during transfer. While EDTA plasma remained stable for up to five days, serum tended to increase after 24 hours. This finding is somewhat divergent from the HISCL presepsin package insert, which indicates plasma stability for up to two days and serum instability exceeding 10% after four hours of refrigeration. Our observation 8% increase in serum presepsin at 24 hours suggests potential influence from factors such as cellular hemolysis or enzymatic interactions, like cathepsin D. This difference highlights the unsuitability of serum as a preferred sample for referral testing [6]. We chose to evaluate EDTA plasma due to its prevalence in laboratories for CBC tests and other analyses, but heparin can also be considered as an alternative anticoagulant for presepsin measurement.

To assess the impact of mixing on presepsin concentration, samples were mixed using the built-in mixing platform of the CBC analyzer. Compared to manual invert mixing, CBC platform mixing significantly increased presepsin levels, with the highest mean value observed at 16 mixing cycles. However, after this initial rise, no further increase in presepsin concentration was observed. This finding aligns with a previous study that examined NT-proBNP and presepsin changes in whole blood under different mixing methods [16]. While NT-proBNP levels remained unaffected, whole blood presepsin measured with the PATHFAST device significantly increased with CBC platform mixing compared to other methods. Similarly, our study, measuring presepsin in EDTA plasma after 30 minutes of centrifugation, also observed elevated levels following CBC platform mixing. Therefore, in a clinical setting where CBC is performed first using EDTA whole blood, followed by centrifugation for presepsin measurement in plasma, an increase in presepsin level is expected. Consequently, it is recommended to use separate specimens for CBC and presepsin testing to ensure accurate results.

Transporting whole blood under cold storage, followed by centrifugation at the referral laboratory, emerged as a viable alternative to transporting EDTA plasma for HISCL presepsin testing. This approach has the potential to significantly reduce the workload for healthcare providers, especially in smaller clinics, without compromising test accuracy. Our study demonstrated that whole blood transported within 12 hours yielded test results comparable to those obtained from EDTA plasma, suggesting the feasibility of ordering HISCL presepsin as a whole blood test even in a setting lacking immediate centrifugation capabilities.

Furthermore, we compared the presepsin values measured by HISCL and PATHFAST, two widely used assays, using 39 specimens. The excellent correlation coefficient of 0.9844 (similar to the 0.979 reported previously [17]) and the strong Spearman rank correlation of 0.9525 confirm a high degree of agreement between the two methods. This finding strengthens the case of centralized HISCL presepsin testing, enabling consistent and reliable results compared to point-of-care devices.

This study has some limitations. We primarily focused on evaluating specimen stability and comparing results across different test methods. As a result, we did not delve into specific diseases, measure other sepsis biomarkers, or perform specific bacterial identifications. Further research is warranted to elucidate the clinical utility of these biomarker combinations.

In summary, our findings offer valuable insights into presepsin stability for referral testing. EDTA plasma exhibited stability for up to five days under refrigeration, making it a suitable candidate for transport. Conversely, serum showed elevated presepsin levels at 24 hours, rendering it unsuitable for referral purposes. Notably, when transferred as EDTA whole blood, plasma remained stable for up to 12 hours following centrifugation. However, given the significant increase in presepsin observed with CBC platform mixing, we recommend separate specimen collection for presepsin testing to ensure accurate results.