The effects of storage temperatures, repeated freeze-thaw cycles, or delays in separating plasma or serum from blood samples are largely unknown for heat shock protein 27 (HSP27). We evaluated (1) the imprecision of the HSP27 assay used in this study; (2) the in vitro stability of HSP27 in blood samples stored at 4℃ for up to 6 hr with immediate and delayed serum/plasma separation from cells; and (3) the in vitro stability of HSP27 in blood samples stored at -80℃ after repeated freeze-thaw cycles. The ELISA to detect HSP27 in this study showed a within-run CV of <9% and a total CV of <15%. After 4-6 hr of storage at 4℃, HSP27 concentrations remained stable when using serum tubes irrespective of sample handling, but HSP27 concentrations decreased by 25-45% when using EDTA plasma tubes. Compared with baseline HSP27, one freeze-thaw cycle had no effect on serum concentrations. However, plasma concentrations increased by 3.1-fold after one freeze-thaw cycle and by 7.3-fold after five freeze-thaw cycles. In conclusion, serum is an appropriate biological sample type for use in epidemiological and large-scale clinical studies.
*Enzyme-Linked Immunosorbent Assay ; Freezing ; HSP27 Heat-Shock Proteins/*blood ; Humans ; Protein Stability ; Reproducibility of Results ; Specimen Handling ; Temperature ; Time Factors

Objective: To assess the prognostic value of the systemic immune-inflammation index (SII) in patients with vulvar cancer. Methods: Data of 130 consecutive patients who underwent primary surgical resection for vulvar cancer at the Medical University of Vienna between 1999 and 2018 was retrospectively analyzed. The SII was defined as platelets × neutrophils/lymphocytes as previously described. Its prognostic value on disease-specific survival (DSS) and overall survival (OS) was evaluated by univariate log-rank tests and multivariable cox regression models. Prediction accuracy was assessed by receiver operating characteristics curves and Youden's J statistics. A HosmerLemeshow test was performed to confirm the model's goodness of fit. Results: A pre-therapeutic high serum SII (>866.4) was associated with advanced International Federation of Gynecology and Obstetrics (FIGO)-stage. In univariate survival analysis, a high SII was associated with both DSS (p<0.001) and OS (p=0.001). A multivariate cox regression model confirmed the prognostic value of SII regarding DSS (p<0.001) and OS (p=0.014) independently from patients' age and FIGO stage. Conclusions Pretherapeutic SII may serve as a promising predictor for survival in patients with vulvar cancer. After clinical validation, the SII may be used to improve both pretreatment patient risk stratification and patient counseling.