Assays for amphetamine and methamphetamine using stereospecific reagents

• US 7,037,669 B2
• Filed: 03/22/2004
• Issued: 05/02/2006
• Est. Priority Date: 03/22/2004
• Status: Active Grant

• Current Assignee: Siemens Healthcare Diagnostics Incorporated (Siemens AG)
• Original Assignee: Dade Behring, Inc. (Siemens AG)
• Inventors: Liu, Hshiou-ting, Zheng, Yi Feng, Yamout, Khaled A., Berger, Donald E. Jr., Hu, Mae W.
• Primary Examiner(s): Le, Long V.
• Assistant Examiner(s): Haq, Shafiqul
• Application Number:
  US10/805,813
• Publication Number:
  US 20050208603A1
• Time in Patent Office: 771 Days
• Field of Search: 435/961, 435/975, 435/7.1, 435/7.9, 435/188, 530/388.9, 530/389.8, 530/403, 530/405, 530/402, 436/546, 436/544, 564/336, 424/175.1, 549/443, 549/444, 548/526
• US Class Current: 435/7.9
• CPC Class Codes: C12N 9/0006 acting on CH-OH groups as donors (1.1)
• CPC Class Codes: G01N 33/946 CNS-stimulants, e.g. cocaine, amphetamines
• CPC Class Codes: Y10S 435/961 including a step of forming, releasing, or exposing the antigen or forming the hapten-immunogenic carrier complex or the antigen per se
• CPC Class Codes: Y10S 435/975 Kit

Abstract

Methods, compositions and kits are disclosed. Enzyme conjugates of Formula I may be employed in assays for the determination of an amphetamine and/or a methamphetamine. Immunogenic conjugates of Formula I may be employed to prepare antibodies for an amphetamine and/or for a methamphetamine for use in assays for the determination of an amphetamine and/or a methamphetamine. The enzyme conjugates may also be employed to screen antibodies for use in such methods.

Claims

1. A compound of the formula:

wherein;

R¹ is H, lower alkyl, a protecting group,

R² is H, lower alkyl, a protecting group,

L is —

(CH₂)_t—

X—

(CH₂)_v—

Y—

wherein X is C(O) or SO₂, Y is a bond, S or —

NR³ wherein R³ is H or lower alkyl, W is O, S, or NH, and t is an integer from 1 to 6 and v is an integer from 2 to 6,

Z is H, a protecting group, a poly(amino acid), a non-poly(amino acid) label moiety, a non-poly(amino acid) immunogenic carrier, or a functional group excluding thiol,

n is 1 when Z is other than a poly(amino acid) or, when Z is a poly(amino acid), n is an integer between 1 and the molecular weight of the poly(amino acid) divided by 500;

and salts thereof.

2. A compound according to claim 1 wherein R¹ is H and R² is H.

3. A compound according to claim 1 wherein R¹ is H and R² is lower alkyl.

4. A compound according to claim 3 wherein lower alkyl is methyl.

5. A compound according to claim 1 wherein Z is a poly(amino acid).

6. A compound according to claim 5 wherein said poly(amino acid) is an enzyme or a protein immunogenic carrier.

7. A compound of the formula:

wherein;

R¹′

is H, lower alkyl, a protecting group,

R²′

is H, alkyl, a protecting group,

W′

is O, S or NR³ wherein R³ is H or lower alkyl,

X′

is C(O) or SO₂,

Y′

is bond, S or —

NR³ wherein R³ is H or lower alkyl,

Z′

is H, a protecting group, a poly(amino acid), a non-poly(amino acid) label moiety, a non-poly(amino acid) immunogenic carrier, or a functional group,

t′

is an integer from 1 to 6 and v′

is an integer from 2 to 6,

n′

is 1 when Z′

is other than a poly(amino acid) or, when Z′

is a poly(amino acid), n′

is an integer between 1 and the molecular weight of the poly(amino acid) divided by 500;

and salts thereof.

8. A compound according to claim 7 wherein R¹′

is H and R²′

is H.

9. A compound according to claim 7 wherein R¹′

is H and R²′

is methyl.

10. A compound according to claim 7 wherein Z′

is a poly(amino acid).

11. A compound according to claim 7, wherein said stereoisomeric mixture comprises at least 90% of one stereoisomeric form over the other.

12. A compound according to claim 11 wherein said stereoisomer has the formula:

13. A compound of the formula:

wherein;

R¹″

is H, lower alkyl, a protecting group,

R²″

is H, lower alkyl, a protecting group,

W″

is O, S, or NR³′

wherein R³′

is H or lower alkyl,

Y″

is bond, S or —

NR³′

wherein R³′

is H or lower alkyl,

Z″

is H, a protecting group, a poly(amino acid), a non-poly(amino acid) label moiety, a non-poly(amino acid) immunogenic carrier, or a functional group,

t″

is an integer from 1 to 6 and v″

is an integer from 2 to 6,

n″

is 1 when Z″

is other than a poly(amino acid) or, when Z″

is a poly(amino acid), n″

is an integer between 1 and the molecular weight of the poly(amino acid) divided by 500;

and salts thereof.

14. A compound according to claim 13 wherein R¹″

is H and R²″

is H.

15. A compound according to claim 13 wherein R¹″

is H and R²″

is methyl.

16. A compound according to claim 13 wherein Z″

is an enzyme.

17. A compound according to claim 16 wherein said enzyme is glucose-6-phosphate dehydrogenase.

18. A compound according to claim 13 wherein Z″

is a protein immunogenic carrier or a non-poly(amino acid) immunogenic carrier.

19. An antibody raised against a compound according to claim 18.

20. A reagent system comprising a compound according to claim 16, an antibody for amphetamine and/or an antibody for methamphetamine.

21. A reagent system comprising an antibody according to claim 19 and an enzyme conjugate of an amphetamine and/or an enzyme conjugate of methamphetamine.

31. A method for determining amphetamine and/or methamphetamine in a sample suspected of containing amphetamine and/or methamphetamine, said method comprising:

(a) providing in combination in a medium;

(i) said sample and

(ii) a reagent system according to claim 20; and

(b) examining said medium for the presence of a complex comprising said amphetamine and said antibody for amphetamine and/or a complex of said methamphetamine and said antibody for methamphetamine, the presence thereof indicating the presence of said amphetamine and/or methamphetamine in said sample.

32. A method according to claim 31 wherein said examining comprises measuring signal from said enzyme, the amount thereof being related to the presence of said amphetamine and/or methamphetamine in said sample.

33. A method according to claim 32 wherein said method is a homogeneous method and said medium is examined for the amount of said signal.

34. A method according to claim 32 wherein said method is a heterogeneous method and said complex, if present, is separated from said medium and said medium or said complex is examined for the amount of said signal.

35. A method for determining amphetamine and/or methamphetamine in a sample suspected of containing amphetamine and/or methamphetamine, said method comprising:

(a) providing in combination in a medium;

(i) said sample and

(ii) a reagent system according to claim 21; and

(b) examining said medium for the presence of a complex comprising said amphetamine and said antibody for amphetamine and/or a complex of said methamphetamine and said antibody for methamphetamine, the presence thereof indicating the presence of said amphetamine and/or methamphetamine in said sample.

36. A method according to claim 35 wherein said examining comprises measuring signal from said enzyme, the amount thereof being related to the presence of said amphetamine and/or methamphetamine in said sample.

37. A method according to claim 36 wherein said method is a homogeneous method and said medium is examined for the amount of said signal.

38. A method according to claim 36 wherein said method is a heterogeneous method and said complex, if present, is separated from said medium and said medium or said complex is examined for the amount of said signal.

22. A compound of the formula:

wherein;

R¹′

″

is H, lower alkyl, a protecting group,

R²′

″

is H, lower alkyl, a protecting group,

W″

is O, S, or NR³′

″

wherein R³′

″

is H or lower alkyl,

Y′

″

is a bond, S or —

NR³′

″

wherein R³′

″

is H or lower alkyl,

Z′

″

is H, a protecting group, a poly(amino acid), a non-poly(amino acid) label moiety, a non-poly(amino acid) immunogenic carrier, or a functional group,

t′

″

is an integer from 1 to 6 and v′

″

is an integer from 2 to 6,

n′

″

is 1 when Z′

″

is other than a poly(amino acid) or, when Z′

″

is a poly(amino acid), n′

″

is an integer between 1 and the molecular weight of the poly(amino acid) divided by 500;

and salts thereof.

23. A compound according to claim 22 wherein R¹′

″

is H and R²′

″

is H.

24. A compound according to claim 22 wherein R¹′

″

is H and R²′

″

is methyl.

25. A compound according to claim 22 wherein Z′

″

is an enzyme.

26. A compound according to claim 25 wherein said enzyme is glucose-6-phosphate dehydrogenase.

27. A compound according to claim 22 wherein Z′

″

is an antigen or a non-poly(amino acid) immunogenic carrier.

28. An antibody raised against a compound according to claim 27.

29. A reagent system comprising a compound according to claim 25, an antibody for amphetamine and/or an antibody for methamphetamine.

30. A reagent system comprising an antibody according to claim 28 and an enzyme conjugate of an amphetamine and/or an enzyme conjugate of methamphetamine.

39. A method for determining amphetamine and/or methamphetamine in a sample suspected of containing amphetamine and/or methamphetamine, said method comprising:

(a) providing in combination in a medium;

(i) said sample and

(ii) a reagent system according to claim 29; and

(b) examining said medium for the presence of a complex comprising said amphetamine and said antibody for amphetamine and/or a complex of said methamphetamine and said antibody for methamphetamine, the presence thereof indicating the presence of said amphetamine and/or methamphetamine in said sample.

40. A method according to claim 39 wherein said examining comprises measuring signal from said enzyme, the amount thereof being related to the presence of said amphetamine and/or methamphetamine in said sample.

41. A method according to claim 40 wherein said method is a homogeneous method and said medium is examined for the amount of said signal.

42. A method according to claim 40 wherein said method is a heterogeneous method and said complex, if present, is separated from said medium and said medium or said complex is examined for the amount of said signal.

43. A method for determining amphetamine and/or methamphetamine in a sample suspected of containing amphetamine and/or methamphetamine, said method comprising:

(a) providing in combination in a medium;

(i) said sample and

(ii) a reagent system according to claim 30; and

(b) examining said medium for the presence of a complex comprising said amphetamine and said antibody for amphetamine and/or a complex of said methamphetamine and said antibody for methamphetamine, the presence thereof indicating the presence of said amphetamine and/or methamphetamine in said sample.

44. A method according to claim 43 wherein said examining comprises measuring signal from said enzyme, the amount thereof being related to the presence of said amphetamine and/or methamphetamine in said sample.

45. A method according to claim 44 wherein said method is a homogeneous method and said medium is examined for the amount of said signal.

46. A method according to claim 44 wherein said method is a heterogeneous method and said complex, if present, is separated from said medium and said medium or said complex is examined for the amount of said signal.

47. A method for determining amphetamine and/or methamphetamine in a sample suspected of containing amphetamine and/or methamphetamine, said method comprising:

(a) providing in combination in a medium;

(i) said sample,

(ii) an antibody for amphetamine,

(iii) an antibody for methamphetamine,

(iv) a compound of the formula;

wherein;

R¹′

is H, lower alkyl, a protecting group,

R²′

is H, lower alkyl, a protecting group,

W′

is O, S, or NR³′

wherein R³′

is H or lower alkyl,

X′

is C(O) or SO₂,

Y′

is bond, S or —

NR³′

wherein R³′

is H or lower alkyl,

Z′

is an enzyme,

t′

is an integer from 1 to 6 and v′

is an integer from 2 to 6,

n′

is an integer between 1 and the molecular weight of said enzyme divided by 500; and

(b) examining said medium for the presence of a complex comprising said amphetamine and said antibody for amphetamine and/or a complex of said methamphetamine and said antibody for methamphetamine, the presence thereof indicating the presence of said amphetamine and/or methamphetamine in said sample.

48. A method according to claim 47 wherein said examining comprises measuring signal from said enzyme, the amount thereof being related to the presence of said amphetamine and/or methamphetamine in said sample.

49. A method according to claim 48 wherein said method is a homogeneous method and said medium is examined for the amount of said signal.

50. A method according to claim 48 wherein said method is a heterogeneous method and said complex, if present, is separated from said medium and said medium or said complex is examined for the amount of said signal.

51. A method according to claim 47 wherein said enzyme is glucose-6-phosphate dehydrogenase.

52. A method for determining amphetamine and/or methamphetamine in a sample suspected of containing amphetamine and/or methamphetamine, said method comprising:

(a) providing in combination in a medium;

(i) said sample,

(ii) a conjugate of an enzyme and an amphetamine analog and/or a conjugate of an enzyme and a methamphetamine analog,

(iii) an antibody for amphetamine, said antibody being raised against a compound of the formula;

wherein;

R¹′

is H and R²′

is H,

W′

is O, S, or NR³′

wherein R³′

is H or lower alkyl,

X′

is C(O) or SO₂,

Y′

is bond, S or —

NR³′

wherein R³′

is H or lower alkyl,

Z′

is a protein immunogenic carrier or a non-poly(amino acid) immunogenic carrier,

t′

is an integer from 1 to 6 and v′

is an integer from 2 to 6,

n′

is an integer between 1 and the molecular weight of said antigen or said immunogenic carrier divided by 500; and

/or

(iv) an antibody for methamphetamine, said antibody being raised against a compound of the formula;

wherein;

R¹′

is H and R²′

is methyl,

W′

is O, S, or NR³′

wherein R³′

is H or lower alkyl,

X′

is C(O) or SO₂,

Y′

is bond, S or —

NR³′

wherein R³′

is H or lower alkyl,

Z′

is a protein immunogenic carrier or a non-poly(amino acid) immunogenic carrier,

t′

is an integer from 1 to 6 and v′

is an integer from 2 to 6,

n′

is an integer between 1 and the molecular weight of said antigen or said immunogenic carrier divided by 500; and

(b) examining said medium for the presence of a complex comprising said amphetamine and said antibody for amphetamine and/or a complex of said methamphetamine and said antibody for methamphetamine, the presence thereof indicating the presence of said amphetamine and/or methamphetamine in said sample.

53. A method according to claim 52 wherein said examining comprises measuring signal from said enzyme, the amount thereof being related to the presence of said amphetamine and/or methamphetamine in said sample.

54. A method according to claim 53 wherein said method is a homogeneous method and said medium is examined for the amount of said signal.

55. A method according to claim 53 wherein said method is a heterogeneous method and said complex, if present, is separated from said medium and said medium or said complex is examined for the amount of said signal.

56. A method according to claim 52 wherein said enzyme is glucose-6-phosphate dehydrogenase.

57. A kit comprising in packaged combination:

(i) an antibody for amphetamine,

(ii) an antibody for methamphetamine,

(iii) a compound of the formula;

wherein;

R¹′

is H, lower alkyl, a protecting group,

R²′

is H, lower alkyl, a protecting group,

W′

is O, S, or NR³′

wherein R³′

is H or lower alkyl,

X′

is C(O) or SO₂,

Y′

is bond, S or —

NR³′

wherein R³′

is H or lower alkyl,

Z′

is an enzyme,

t′

is an integer from 1 to 6 and v′

is an integer from 2 to 6,

n′

is an integer between 1 and the molecular weight of said enzyme divided by 500.

58. A kit according to claim 57 wherein said enzyme is glucose-6-phosphate dehydrogenase.

59. A kit comprising in packaged combination:

(i) a conjugate of an enzyme and an amphetamine analog and/or a conjugate of an enzyme and a methamphetamine analog,

(ii) an antibody for amphetamine, said antibody being raised against a compound of the formula;

wherein;

R¹′

is H and R²′

is H,

W′

is O, S, or NR³′

wherein R³′

is H or lower alkyl,

X′

is C(O) or SO₂,

Y′

is bond, S or —

NR³′

wherein R³′

is H or lower alkyl,

Z′

is a protein immunogenic carrier or a non-poly(amino acid) immunogenic carrier,

t′

is an integer from 1 to 6 and v′

is an integer from 2 to 6,

n′

is an integer between 1 and the molecular weight of said antigen or said immunogenic carrier divided by 500; and

/or

(iii) an antibody for methamphetamine, said antibody being raised against a compound of the formula;

wherein;

R¹′

is H and R²′

is methyl,

W″

is O, S, or NR³′

wherein R³′

is H or lower alkyl,

X′

is C(O) or SO₂,

Y′

is bond, S or —

NR³′

wherein R³′

is H or lower alkyl,

Z′

is a protein immunogenic carrier or a non-poly(amino acid) immunogenic carrier,

t′

is an integer from 1 to 6 and v′

is an integer from 2 to 6,

n′

is an integer between 1 and the molecular weight of said antigen or said immunogenic carrier divided by 500.