| Autor: |
Seto Y; Department of Chemistry, Faculty of Engineering, Gunma University, 1-5-1 Tenjin-cho, Kiryu, Gunma, 376-8515, Japan. yoshiakiseto@eiken.co.jp, Abe K, Itoh M, Sawai H |
| Jazyk: |
angličtina |
| Zdroj: |
Bioconjugate chemistry [Bioconjug Chem] 2002 Mar-Apr; Vol. 13 (2), pp. 303-8. |
| DOI: |
10.1021/bc015554l |
| Abstrakt: |
A bioluminescent enzyme immunoassay (BLEIA) for 2',5'-oligoadenylate 5'-triphosphate (pppA2'p5'A2'Ap5'A, 2-5A) was developed using a conjugate of 2-5A and firefly luciferase as a probe. The conjugate was prepared from a 2-5A derivative bearing a thiol group at the 2'(or 3') end, and streptavidin (SA) bearing maleimide via a linker-arm and biotinylated luciferase. The thiol group of the 2-5A derivative was relatively stable under aerobic conditions and remained 100% and 56% intact at 25 degrees C after 1 and 96 h, respectively, without dimerization by aerobic oxidation. The thiol-modified 2-5A was coupled with the SA-maleimide conjugate, followed by complex formation with biotinylated firefly luciferase. BLEIA using this conjugate allowed for the direct analysis of 2-5A in a range of 5-500 fmol (0.1-10 nM in a 50 microL sample) and in a reaction time of 15 min. The measurement of microquantities of 2-5 A from various sources is easy to perform by this BLEIA, because no radioisotope labeled 2-5A was required as a probe. |
| Databáze: |
MEDLINE |
| Externí odkaz: |
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