Process and kit to initiate a luminiferous reaction

Details Process and kit to initiate a luminiferous reaction Process and kit to initiate a luminiferous reaction

1997-08-18

1998-06-30

Snay, Jeffrey

Chemistry: analytical and immunological testing

Optical result

With fluorescence or luminescence

422 61, 435810, 435 8, G01N 2176

Patent

active

057733031

DESCRIPTION:

BRIEF SUMMARY
TECHNICAL FIELD

The invention relates to a starter kit to initiate luminiferous reaction by oxidation of luminescent molecules in analytical tests.


PRIOR ART

Light-emitting chemical reactions have various uses in analytical tests. In clinical analysis, they have found wide use, in particular in immunoassays, in protein blotting and in DNA samples. The most successful tests include those in which the luminescent molecules employed are isoluminol derivatives and acridinium derivatives which give off energy in the form of light quanta in alkaline solution with consumption of peroxide. Particularly in the case of isoluminol derivatives, this reaction is subject to very slow kinetics. In order that a measurable amount of light is released in a reasonable time, catalysts are added. These catalysts mostly contain complexed iron. Beside potassium ferricyanide or ferrocyanide, compounds such as microperoxidase, hemin, hematin or enzymes such as catalase and horseradish peroxidase are described as suitable in the literature. In particular, catalase and microperoxidase have already been successfully used in commercial systems. In the case of isoluminol derivatives, ABEI (aminobutylethylisoluminol) or ABENH (aminobutylethylnaphthalenedicarboxylic hydrazide) have gained acceptance, since in contrast to luminol they can be coupled without problems to haptens or proteins without noticeable loss of quantum yield.
To measure the luminescence, suitable reagent solutions are pipetted into a solution containing the label in unknown amount in a measuring apparatus and the luminescence is immediately measured. The kinetics of the photoreactions of both classes of substance proceed in a few seconds under optimized conditions.
With luminescence becoming known in the literature, to initiate luminescence the procedure was originally used was to pipette three solutions: sodium hydroxide solution, catalyst and finally dilute hydrogen (1983)!. With increasing commercialization of luminescence in immunoassays, a simplification of the light production was achieved by use of an alkaline peroxide solution (DE 3439742). The alkaline peroxide solution described is sufficiently stable for use in immunoassays within a given period of time after preparation, so that after addition of the catalyst the luminescence reaction is produced by injection of the alkaline peroxide solution into the measuring chamber. In an optimized variant which has been used commercially by the applicant for some years, the light yield can be increased in that first the alkaline peroxide solution is pipetted and then the catalyst starts the photo-reaction. The light yield is thereby again markedly increased. In both variants, the concentration of the sodium hydroxide solution used is very high (about 1M), as the light yield increases with increasing concentration. The combination of the two solutions of catalyst and peroxide is usually described as a starter kit.
A major problem of the starter kits used for light production is their restricted shelf-life. This has the consequence that in no case can they remain uncooled in an automatic measuring system for several days. For this reason, the solutions of a starter kit should if possible be completely used up on the same day or else stored in a refrigerator in the meantime at 2.degree.-8.degree. C. Stability investigations have shown that the usual liquid solutions can only be used for a maximum of 14 days even under optimum storage conditions. After this time, particularly due to the known light and temperature sensitivity of hydrogen peroxide in alkaline solution, on the one hand a marked decrease in the luminescence signal and on the other hand an increase in the nonspecific reagent blank value are to be observed. Thus the noise/signal ratio becomes markedly worse with increasing aging of the usual solutions, which markedly restricts their use in routine operation or in an automatic immunoassay analyzer.
Starter solutions are therefore also not offered for sale in ready-to-use form in commercial test systems, but

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Jones, P. et al "Kinetics and mechanism of catalysis by ferrihemes in the chemiluminogenic oxidation of luminol by hydrogen peroxide" Chemical Abstracts, vol. 107, No. 5 (1987) abstract No. 39496u.
Frew, J.E. et al "Assay of same clinically important reductants by a chemiluminescence delay technique" Chemical Abstracts, vol. 103, No. 25 (1985) abstract No. 210411f.
Schroeder et al., Analytical Chemistry, ,"Chemiluminescence Yields and Detection Limits of Some Isoluminol Derivatives in Various Oxidation Systems", vol. 50, No. 8, 1114-1120, Jul. 1978.

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