Influence of variations in sample handling on SELDI‐TOF MS serum protein profiles for colorectal cancer

Abstract

Sample handling can have a profound effect on serum protein profiles, challenging results obtained with archived sera under non‐standardized sample collection. Here, we evaluate the influence of variations in sample handling on previous serum protein profiles for colorectal cancer (CRC) (Engwegen et al.,. World J. Gastroenterol. 2006, 12, 1536–1544). Sera were prospectively obtained from individuals with an indication for colonoscopy (n = 150: 65 controls, 52 adenomatous polyps, 29 CRC, 4 unknown), as well as from normal volunteers (n = 8). Protein profiles were acquired by SELDI‐TOF MS on CM10 chips at pH 5. We assessed the influence of storage temperature, type of collection tube, coagulation temperature and freeze‐thaw cycles on the serum protein profile. Several peptides occurred only in samples stored at –20°C, indicating proteolytic degradation during storage. One was a previous CRC biomarker candidate, an N‐terminal albumin fragment (m/z 3087), and two others complement C3f and a fragment thereof (m/z 2022 and 1863). Overall differences in protein profiles were also seen for different collection tubes, coagulation temperature and freeze‐thaw cycles. However, three of five of our previously defined CRC biomarker candidates are stable to variations in the sample handling protocol, justifying their further validation in prospective studies.

Influence of sample storage duration on serum protein profiles assessed by surface-enhanced laser desorption/ionisation time-of-flight mass spectrometry (SELDI-TOF MS)

“…has been described previously by our group (22). We observed the N-terminal albumin fragment to be positively correlated with duration of storage (1.4 years) at -308C.…”

“…The susceptibility of detected proteins to sample handling issues has been discussed in previous studies (8,22,31). A number of these proteins have, however, also been reported as potential cancer markers (10,11,21).…”

“…Clinical studies generally exceed these storage durations, since study sera either originate from sample banks (18)(19)(20), or were collected prospectively over a period of years (21). We have previously investigated the effects of long-term storage (0-16 months) using a larger sample size (ns150) (22). However, even this extended storage interval is usually exceeded by clinical proteomics studies.…”

Approaching clinical proteomics: current state and future fields of application in fluid proteomics

“…Differences between samples frozen for 1-3 months at -208C or -808C have not been observed in several studies (16,26,34). However, this may be due to the short storage times because another research group reported that most differences became apparent after about 5 months of storage (35). One peptide noted to be altered by storage at low temperature was a biomarker previously identified as a candidate for colorectal cancer, an N-terminal albumin fragment (m/z 3087).…”

Diagnostic and prognostic significance of serum apolipoprotein C-I in triple-negative breast cancer based on mass spectrometry

“…False discovery can exert a distinct influence on the serum proteome, potentially leading to significant differences between results, even with comparable patient populations and sample types under study. [22][23][24][25][26] New proteomic technologies that promote large-scale sample screening and facilitate identification of proteins associated with disease and treatment are developing rapidly. 27,28 Mass spectrometry (MS), a powerful proteomics tool, has evolved to a high-throughput level, allowing rapid and accurate analysis of several thousand proteins in a single study.…”