Bioluminescent detection probe for tick-borne encephalitis virus immunoassay.
To facilitate the detection of the tick-borne encephalitis virus (TBEV), the causative agent of one of the most severe human neuroinfections, we have developed an immunoassay based on bioluminescent hybrid protein 14D5a-Rm7 as a detection probe. The protein containing Renilla luciferase as a reporter and a single-chain variable fragment (scFv) of murine immunoglobulin to TBEV as a recognition element was constructed, produced by bacterial expression, purified, and tested. Both domains were shown to reveal their specific biological properties-affinity to the target antigen and bioluminescent activity. Hybrid protein was applied as a label for solid-phase immunoassay of the antigens, associated with the tick-borne encephalitis virus (native glycoprotein E or extracts of the infected strain of lab ticks). The assay demonstrates high sensitivity (0.056 ng of glycoprotein E; 10(4)-10(5) virus particles or 0.1 pg virions) and simplicity and is competitive with conventional methods for detection of TBEV.
Institute of Biophysics, Siberian Branch, Russian Academy of Sciences, Krasnoyarsk, 660036, Russia.
Ludmila P
LP
Burakova
Alexander N
AN
Kudryavtsev
Galina A
GA
Stepanyuk
Ivan K
IK
Baykov
Vera V
VV
Morozova
Nina V
NV
Tikunova
Maria A
MA
Dubova
Victor N
VN
Lyapustin
Valeri V
VV
Yakimenko
Ludmila A
LA
Frank
D000818:Animals; D004669:Encephalitis Viruses, Tick-Borne / Q000302:isolation & purification*; D004675:Encephalitis, Tick-Borne / Q000821:virology*; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D049410:Luciferases, Renilla / Q000737:chemistry* / Q000235:genetics; D049408:Luminescent Agents / Q000737:chemistry* / Q000378:metabolism; D008163:Luminescent Measurements / Q000379:methods*; D051379:Mice; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D057127:Single-Chain Antibodies / Q000737:chemistry* / Q000235:genetics; D013987:Ticks
D049408:Luminescent Agents; D011993:Recombinant Fusion Proteins; D057127:Single-Chain Antibodies; D049410:Luciferases, Renilla