Kits pertaining to analyte determination
View Patent ↗This invention pertains to methods, kits and/or compositions for the determination of analytes by mass analysis using unique labeling reagents or sets of unique labeling reagents. The labeling reagents can be isomeric or isobaric and can be used to produce mixtures suitable for multiplex analysis of the labeled analytes.
1. A kit comprising:
a) a set of two or more isobaric and/or isomeric reagents for the labeling of analytes, wherein all reagents of the set comprise the formula:
or a salt thereof, wherein;
i) RG is a reactive group that is an active ester wherein the alcohol moiety of the active ester is:
wherein X′ is O or S;
ii) Z is O, S, NH or NR 1 ;
iii) each j is the same or different and is selected from the group consisting of: H, R 1 , OR 1 , SR 1 , NHR 1 , N(R 1 ) 2 , fluorine, chlorine, bromine and iodine;
iv) one of A 1 , A 2 , A 3 , A 4 and A 5 is W, wherein W is selected from the group consisting of: NH, N—R 2 , P—R 2 , O and S; and
v) the remainders of A 1 , A 2 , A 3 , A 4 and A 5 that isn't W have the formula C(J) 2 , where J is as defined above;
vi) each R 1 is the same or different and is an alkyl group comprising one to eight carbon atoms which may optionally contain a heteroatom or a substituted or unsubstituted aryl group wherein the carbon atoms of the alkyl and aryl groups independently comprise linked hydrogen and/or fluorine atoms; and
R 2 is an amino alkyl, hydroxy alkyl, thio alkyl group or a cleavable linker that cleavably links the reagent to a solid support wherein the amino alkyl, hydroxy alkyl or thio alkyl group comprises one to eight carbon atoms, which may optionally contain a heteroatom or a substituted or unsubstituted aryl group, and wherein the carbon atoms of the alkyl and aryl groups independently comprise linked hydrogen, and/or fluorine atoms; and
wherein each reagent of the set is optionally isotopically enriched by replacing one or more atoms of the reagent with a corresponding heavier atom isotope; and
b) one or more other reagents, containers, enzymes, buffers or instructions.
2. The kit of claim 1 , wherein the kit comprises a proteolytic enzyme.
3. The kit of claim 1 , wherein the active ester is an N-hydroxysuccinimide ester.
4. The kit of claim 1 , wherein each reagent of the set is independently linked to a solid support through a cleavable linker.
5. The kit of claim 1 , wherein all reagents of the set are isomeric.
6. The kit of claim 1 , wherein all reagents of the set are isobaric.
7. The kit of claim 1 , wherein the set comprises one or more of the following four reagents:
8. The kit of claim 1 , wherein the set comprises one or more of the following four reagents:
9. The kit of claim 1 , wherein the set comprises one or more of the following four support bound reagents:
wherein:
a) E is a solid support;
b) F is a cleavable linker linked to the solid support; and
c) G is an amino alkyl, hydroxy alkyl or thio alkyl group, cleavably linked to the cleavable linker wherein the amino alkyl, hydroxy alkyl or thio alkyl group comprises one to eight carbon atoms, which may optionally contain a heteroatom or a substituted or unsubstituted aryl group, and wherein the carbon atoms of the alkyl and aryl groups independently comprise linked hydrogen, deuterium and/or fluorine atoms.
10. The kit of claim 1 , wherein each reagent of the set is isotopically enriched with one or more heavy atom isotopes.
11. The kit of claim 10 , wherein each heavy atom isotope is present in at least 80% isotopic purity.
12. The kit of claim 10 , wherein each heavy atom isotope is present in at least 93% isotopic purity.
13. The kit of claim 10 , wherein each heavy atom isotope is present in at least 96% isotopic purity.