Measurement of Vasoactive Intestinal Peptide using a Competitive Fluorescent Microsphere Immunoassay or ELISA in human blood samples

• Published in: Journal of immunological methods Vol. 300; no. 1; pp. 63 - 73
• Main Authors: Song, Eun Young, VanDunk, Cassandra, Kuddo, Thea, Nelson, Phillip G.
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 01.05.2005; Elsevier
• Subjects: Adult; Binding, Competitive; Biological and medical sciences; Buffers; ELISA; Enzyme-Linked Immunosorbent Assay - methods; Enzyme-Linked Immunosorbent Assay - standards; Enzyme-Linked Immunosorbent Assay - statistics & numerical data; Fluorescent Microsphere Immunoassay; Fluoroimmunoassay - methods; Fluoroimmunoassay - standards; Fluoroimmunoassay - statistics & numerical data; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Humans; Microspheres; Molecular immunology; Reference Standards; Reproducibility of Results; Sensitivity and Specificity; Techniques; Vasoactive Intestinal Peptide - analysis; Vasoactive Intestinal Peptide - blood; Vasoactive Intestinal Peptide - standards; VIP; LOD; PBS; Fluorescent Microsphere Immunoassay; Ab; FMI; PACAP; S.D; RIA; RIC; LOQ; PHI; BSA; cFMI; PE; EIA; VIP; CGRP; ELISA; Human; Vasoactive intestinal peptide; ELISA assay; Blood; Immunological method
• ISSN: 0022-1759; 1872-7905
• DOI: 10.1016/j.jim.2005.02.009

The concentration of Vasoactive Intestinal Peptide (VIP) as measured by recycling immunoaffinity chromatography (RIC) has been reported to be elevated in the blood of patients with autism as compared with normal subjects. In this study, we have developed a “Competitive Fluorescent Microsphere Immunoassay” (cFMI) in which VIP competes with biotinylated VIP in binding to polyclonal antibodies on microspheres. The results were obtained using the Luminex 100 system. We measured VIP in serum, plasma, and material eluted from dried blood spots on filter paper with both the cFMI and an ELISA procedure. We found that a purification procedure was necessary for obtaining useful results from plasma and serum, however, a preincubation step was required with the blood eluates. This newly developed cFMI was more sensitive (2.5 vs. 20.0 pg/ml), and more reproducible than the ELISA. To get accurate measurements of VIP in eluted material high sensitivity is especially important. Thus, the cFMI using the Luminex system has definite advantages over a conventional ELISA including the possibility that samples can be assayed at higher dilutions. We have determined that the VIP concentrations of serum, plasma, and dried blood spot eluate specimens as measured with the cFMI assay system were similar to those measured with ELISA. Thus, the new cFMI using Luminex system may be useful for detection of VIP in human blood samples.