Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving luciferase
Patent
1999-04-30
2000-05-09
Leary, Louise N.
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving luciferase
435 4, C12Q 166
Patent
active
060602619
DESCRIPTION:
BRIEF SUMMARY
TECHNICAL FIELDS
The present invention relates to a luminescence method for luciferin/luciferase system, which stabilizes a luminescent reaction and a luciferase assay reagent. The luminescence method for luciferin/luciferase system and the luciferase assay reagent, provided by the present invention, can be applied to all assay systems using a luciferase enzyme as a reporter and signal.
BACKGROUND
In a luminescent reaction with luciferin/luciferase in vitro, conventionally, the luminescence pattern thereof is observed in a flash state. Therefore, it is impossible to measure the luminescent reaction accurately without using a device having a specific mechanism of injecting a reagent. However, K. V. Wood et al. has established a luminescence system having a relatively long half-life period of luminescence as long as 5 minutes. (Wood, K. V. Recent advantage and prospects for beetle luciferase as genetic reporters. In: Bioluminescence and Chemiluminescence current status. Proceeding of the VIth International Symposium on Bioluminescence and Chemiluminescence, Cambridge, September 1990. P.543. Ed. by P. Stanley and L. J. Kricka.). Owing to this, it has become possible to accurately measure a luminescence unit in a luciferin/luciferase system with a luminometer or a liquid scintillation counter which does not have an automatic reagent-injecting device. So, a reporter assay method wherein luciferase generated as a reporter in a culture cell is measured at a high sensitivity has been widely used. However, when the reporter assay method is used in a High Throughput screening for developing a medication, the luminescence half-life period of 5 minutes is insufficient. For example, for measuring many samples by using 96 wells, luminescent reagents must be added several times and measurement must be carried out each time. Like this, restrictions on operation and measurement exist.
Further, luciferin contained as a luminescent substrate in a luminescent reagent is apt to be oxidized in a solution during storage, to form an oxyluciferine. It is well-known that the oxyluciferin inhibits the enzyme reaction of a luciferase. Conventional luminescent reagents containing luciferin as a luminescent substrate have a big problem that a luminescent activity deteriorates due to the oxidation of the luciferin. This problem prevents the further expansion of uses for the assay using luciferase as a reporter. Therefore, the development of a new stabilization method is expected.
It is an object of the present invention to provide a luminescence method for luciferin/luciferase system, which extends the half-life period of a luminescent reaction, and a luciferase assay reagent.
It is an another object of the present invention to a luminescence method for luciferin/luciferase system, which prevents the deterioration of a luminescent activity and gives a stable luminescence reaction, and a Luciferase assay reagent.
DISCLOSURE OF THE INVENTION
According to the present invention, there is provided a luminescence method for luciferin/luciferase system, which comprises reacting luciferin solution containing an increased amount of dissolved carbon dioxide with luciferase.
According to the present invention, further, there is provided a luminescence method for luciferin/luciferase system, wherein the pH of the luciferin solution is adjusted in the range of from 5.0 to 8.5.
According to the present invention, further, there is provided a luminescence method for luciferin/luciferase system, wherein the amount of the dissolved carbon dioxide in the luciferin solution is increased by adding a dry ice.
According to the present invention, further, there is provided a luminescence method for luciferin/luciferase system, wherein the amount of the dissolved carbon dioxide in the luciferin solution is increased by introducing a carbon dioxide gas.
According to the present invention, further, there is provided a luciferase assay reagent which comprises a luciferin solution containing an increased amount of dissolved carbon dioxide.
According to the
REFERENCES:
patent: 5618682 (1997-04-01), Scheirer
patent: 5866348 (1999-02-01), Scheirer
Ryufuku Masayuki
Takeuchi Hideyuki
Tanaka Hozumi
Leary Louise N.
Toyo Ink Mfg. Co. Ltd.
LandOfFree
Luminescence method for luciferin/luciferase system and luminesc does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Luminescence method for luciferin/luciferase system and luminesc, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Luminescence method for luciferin/luciferase system and luminesc will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-1063776