RESUMEN
Human pancreatic polypeptide (HPP) is a 36 amino acid peptide hormone that plays a role in the bidirectional communication between the digestive system and the brain. HPP measurements are used to assess vagal nerve function following sham feeding and to detect gastroenteropancreatic-neuroendocrine tumors. These tests have historically been conducted by radioimmunoassays, but liquid chromatography-tandem mass spectrometry (LC-MS/MS) has several advantages such as improved specificity and elimination of radioactive molecules. Here, we present our LC-MS/MS method. Initially, samples were immunopurified and subjected to LC-high resolution accurate mass tandem mass spectrometry (HRAM-MS/MS) to identify circulating forms of the peptide in human plasma. We identified 23 forms of HPP, including several glycosylated forms. The most abundant peptides then were used for targeted LC-MS/MS measurements. LC-MS/MS performance for precision, accuracy, linearity, recovery, limit of detection, and carryover met our acceptance criteria based on CLIA regulations. Additionally, we observed the expected physiological rise in HPP in response to sham feeding. Our results indicate that HPP measurement by LC-MS/MS produces clinically equivalent results to our established immunoassay when several peptides are monitored, making it a suitable replacement. The measurement of peptide fragments, including modified species, might have additional clinical value.
Asunto(s)
Polipéptido Pancreático , Espectrometría de Masas en Tándem , Humanos , Cromatografía Liquida/métodos , Espectrometría de Masas en Tándem/métodos , Péptidos , Inmunoensayo/métodosRESUMEN
BACKGROUND: Squamous cell carcinoma antigen (SCCA) is a glycoprotein biomarker for squamous cell carcinoma (SCC). SCCA elevations have also been noted in other conditions. The purpose of this study was to evaluate the analytical and clinical performance of an automated SCCA homogenous immunofluorescent assay (BRAHMS KRYPTOR). METHODS: Reference intervals were determined using 119 samples from healthy donors. To assess clinical performance, samples were collected from patients with cervical (n = 12), head and neck (n = 23), lung (n = 14), or cutaneous (n = 11) SCC in addition to hepatocellular carcinoma, psoriasis, or atopic dermatitis. RESULTS: Upper 95th percentile sex-specific reference intervals were 2.00 µg/L for males and 1.67 µg/L for females. Intra- and inter-assay CVs were less than 5%. Comparison of the BRAHMS KRYPTOR to an ELISA SCCA immunoassay exhibited a correlation coefficient of 0.8809. The mean sensitivity for all SCC positive patients was 23.3%. With the exception of psoriasis (58.6%) specificity exceeded 95% for the non-SCC populations. CONCLUSION: The BRAHMS KRYPTOR SCCA assay showed good analytical performance and acceptable overall clinical specificity. Consistent with previous studies, the sensitivity of SCCA for SCC was low. In the absence of other robust circulating markers, SCCA remains an imperfect yet useful tool in the evaluation of SCC.