Development of a sensitive enzyme-linked immunosorbent assay for eotaxin and measurement of its levels in human blood
| Autor: | Hiroshi Teraoka, Motoji Kitaura, Koichi Hirai, Atsushi Morita, Osamu Yoshie, Mikio Tamaki, Kazuyuki Sasakura, Toshiharu Nakajima, Yasushi Taniguchi, Shino Kikuoka, Tetsuo Tsuji, Kenichi Shimosako |
|---|---|
| Rok vydání: | 1999 |
| Předmět: |
Chemokine CCL11
Eotaxin Chemokine Chemotactic Factors Eosinophil medicine.medical_treatment Immunology Enzyme-Linked Immunosorbent Assay Sensitivity and Specificity law.invention law medicine Humans Immunology and Allergy Bovine serum albumin medicine.diagnostic_test biology Chemistry respiratory system Molecular biology Recombinant Proteins Recombinant Eotaxin Cytokine Chemokines CC Immunoassay Recombinant DNA biology.protein Cytokines Quantitative analysis (chemistry) |
| Zdroj: | Journal of Immunological Methods. 226:159-167 |
| ISSN: | 0022-1759 |
| DOI: | 10.1016/s0022-1759(99)00066-6 |
| Popis: | The CC chemokine eotaxin is potent eosinophil-selective chemoattractant, and it is thought that the function of eotaxin is closely related to the recruitment of eosinophils in certain inflammatory reactions. In order to learn more about the biological role of this molecule, we have developed a new sandwich ELISA method to measure human eotaxin using two monoclonal anitibodies and purified recombinant eotaxin as a standard. The minimal detectable concentration of eotaxin in this assay was 1.5 pg/ml, and the working range was 3.1--200 pg/ml with low CVs (< 10%). Both within- and between-run precision levels were less than 6.7% of the CVs. The dilution curves of two serum and two spiked plasma samples showed good linearity and the recovery range was 92.8--103.3%. No cross-reactivity was found with other similar chemokines. MCP-1, MCP-2, MCP-3, MCP-4, eotaxin-2 and RANTES. This assay was sensitive enough to measure the circulating eotaxin levels of healthy volunteers. However, the eotaxin levels in serum samples (mean+/-SD; 68.6+/-13.4 pg/ml, n=15) were significantly higher than those in matched plasma samples (19.2+/-5.4 pg/ml) separated from blood collected in tubes containing EDTA. Kinetic studies revealed that the eotaxin levels in serum markedly increased depending on the elapsed time before separation from blood cells, but such changes in EDTA-plasma were negligible up to 4 h at 25 degrees C. Our new ELISA is an accurate and useful method for quantifying human eotaxin in blood and demonstrates that the process of preparing blood samples affects the measurement of the eotaxin levels. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|
Full Text from ScienceDirect