title: Improved sensitivity of colorimetric compared to chemiluminescence ELISAs for cytokine assays.
abstract: Cytokines are often measured using ELISAs and chemiluminescence (CMIL) is reported to exhibit increased sensitivity compared to colorimetric (COL) assays. CMIL also has a wider dynamic detection range. We sought to directly compare ELISAs for measuring human TNF and IL-8 using CMIL or COL. CMIL substrates with glow fluorescence were obtained from 4 different commercial sources while the COL substrate was TMB. ELISAs for TNF and IL-8 were run under identical conditions and the standard curve extended from 0.5 to 4000 pg/ML. The COL substrate demonstrated a sigmoid shaped curve when plotted on a log-linear scale while the CMIL continued to increase up to the highest concentration. Both substrates were modeled most accurately by a 4 parameter equation with R values > 0.99. The standard curves for both the IL-8 and TNF demonstrated a lower limit of detection (LLD) for the COL comparable to the CMIL detection system. To precisely define the LLD quadruplicate blanks were run and the mean plus 4 standard deviations were used. By these criteria, the COL assay routinely had a LLD of < 1.5 pg/ML which was better than any of the CMIL substrates. Our data demonstrate the COL assays have the same or better sensitivity than CMIL and are significantly less expensive.
authors.affiliation: Department of Pathology, University of Michigan Medical School, Ann Arbor, Michigan 48109-0602, USA.
authors.forename: Javed
authors.initials: J
authors.lastname: Siddiqui
authors.forename: Daniel G
authors.initials: DG
authors.lastname: Remick
mesh_terms: D003124:Colorimetry; D016207:Cytokines / Q000032:analysis*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods* / Q000706:statistics & numerical data; D006801:Humans; D007202:Indicators and Reagents; D016209:Interleukin-8 / Q000032:analysis; D008163:Luminescent Measurements; D012680:Sensitivity and Specificity; D019809:Streptavidin; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis
chemical_list: D016207:Cytokines; D007202:Indicators and Reagents; D016209:Interleukin-8; D014409:Tumor Necrosis Factor-alpha; D019809:Streptavidin