Method for interpreting tandem mass spectrometry data for clinical diagnosis

• US 7,229,834 B2
• Filed: 09/13/2005
• Issued: 06/12/2007
• Est. Priority Date: 01/30/1999
• Status: Expired due to Term

• Current Assignee: PerkinElmer Genetics, Inc. (Revvity, Inc.)
• Original Assignee: PerkinElmer Genetics, Inc. (Revvity, Inc.)
• Inventors: Chace, Donald H.
• Primary Examiner(s): Cole; Monique T.
• Application Number:
  US11/225,243
• Publication Number:
  US 20060128027A1
• Time in Patent Office: 637 Days
• Field of Search: 702/22, 702/23, 702/27, 436/173, 436/86
• US Class Current: 436/86
• CPC Class Codes: C12Q 1/32 involving dehydrogenase
• CPC Class Codes: G01N 2800/02 Nutritional disorders
• CPC Class Codes: G01N 33/6815 containing sulfur, e.g. cysteine, cystine, methionine, homocysteine
• CPC Class Codes: G01N 33/6848 Methods of protein analysis involving mass spectrometry
• CPC Class Codes: G01N 33/6893 related to diseases not provided for elsewhere
• CPC Class Codes: G01N 33/92 involving lipids, e.g. cholesterol , lipoproteins, or their receptors steroid hormones G01N33/743
• CPC Class Codes: H01J 49/0036 Step by step routines describing the handling of the data generated during a measurement
• CPC Class Codes: Y10T 436/24 Nuclear magnetic resonance, electron spin resonance or other spin effects or mass spectrometry

Abstract

A method for interpreting data that is produced after a group of amino acids and acylcarnitines are derivatized from blood spots taken from newborn babies and scanned by a tandem mass spectrometer. Concentration levels of each metabolite, which are directly proportional to the butyl ester fragment after derivatization, are compared to threshold flags for determining a significance of any deviation of the metabolite relative to the flag threshold. The threshold flags are diagnostic limits to the data retrieved from each blood spot. The data includes metabolite concentrations and molar ratios of metabolites with other metabolites. Samples are labeled normal for a disease if the concentration of any of the metabolite concentrations or molar ratio concentration do not deviate from the flag threshold, but, in contrast, the sample must be further evaluated if a value is elevated or deficient to some degree. Thus, as each metabolite fragments at a different mass to charge value (m/z), corresponding data is compared to the respective flag thresholds for determining a next course of action that must be taken to ultimately assist a physician in the diagnosis of a genetic disorder resulting from an elevation or deficiency of the metabolite particular for that disorder.

Claims

1. A method comprising steps of:

acquiring mass spectral data from a sample that has been derived from the blood of a patient, wherein said data includes values comprised of a propionyl carnitine concentration (C3) and a molar ratio of propionyl carnitine and acetyl carnitine (C3/C2) or a molar ratio of propionyl carnitine and palmitoyl carnitine (C3/C16);

comparing said values, respectively, with a C3 flag threshold and a C3/C2 flag threshold or a C3/C16 flag threshold;

identifying whether or not there is an elevation of any of said values above any of said flag thresholds; and

interpreting said sample as being normal for propionic acidemia, provided there is no said elevation of any of said values.

2. The method of claim 1, wherein:

in the acquiring step, said data includes values comprised of a propionyl carnitine concentration (C3), and a molar ratio of propionyl carnitine and acetyl carnitine (C3(C2) and a molar ratio of propionyl carnitine and palmitoyl carnitine (C3/C16); and

in the comparing step, said C3, C3/C2 and C3/C16 are compared with a C3 flag threshold, a C3/C2 flag threshold and a C3/C16 flag threshold, respectively.

3. The method of claim 1 further comprising:

flagging the sample for re-analysis provided any of the following occur;

i. said C3 is equal to or greater than said C3 flag threshold;

ii. said C3/C16 is greater than or equal to said C3/C16 flag threshold and said C3/C2 is greater than said C3/C2 flag threshold and said C3 is greater than about 2.5 μ

M;

or

iii. said C3 is greater than about 4 μ

M, and either said C3/C16 is greater than said C3/C16 flag threshold or said C3/C2 is greater than said C3/C2 flag threshold.

4. The method of claim 3 further comprising:

performing a re-analysis of said sample or a new sample from the same patient to obtain mean values.

5. The method of claim 4 further comprising:

initiating a follow-up protocol based on follow-up criteria utilizing said mean values.

6. The method of claim 1 further comprising:

flagging the sample for re-analysis provided any of the following occur;

i. said C3 is greater than about 9.0 μ

M;

or

ii. said C3 is greater than about 7.0 μ

M and said C3/C2 is greater than said C3/C2 flag threshold or said C3/C16 is greater than said C3/C16 flag threshold.

7. The method of claim 6 further comprising:

performing a re-analysis of said sample or a new sample from the same patient to obtain priority mean values.

8. The method of claim 7 further comprising:

initiating a follow-up protocol based on follow-up criteria utilizing said priority mean values.

9. The method of claim 1, wherein said C3 flag threshold, said C3/C2 flag threshold, and said C3/C16 flag threshold are, respectively, at least about 3.4 μ

M, 0.21, and 1.12.

10. The method of claim 1, wherein said C3 flag threshold, said C3/C2 flag threshold, and said C3/C16 flag threshold are, respectively, at least about 4.6 μ

M, 0.29, and 1.54.

11. The method of claim 1, wherein said C3 flag threshold, said C3/C2 flag threshold, and said C3/C16 flag threshold are, respectively, about 5.0 μ

M, 0.30, and 1.75.

12. The method of claim 1, wherein the sample was derived from a dry blood spot on filter paper.

13. The method of claim 12, wherein the sample was derivatized.

14. The method of claim 1, wherein:

in the acquiring step, said data was acquired using a tandem mass spectrometer.

15. The method of claim 1, wherein:

the sample was derived from a dry blood spot on filter paper;

the sample was derivatized; and

in the acquiring step, said data was acquired using a tandem mass spectrometer.

16. The method of claim 15, wherein said C3 flag threshold, said C3/C2 flag threshold, and said C3/C16 flag threshold are, respectively, at least about 3.4 μ

M, 0.21, and 1.12.

17. The method of claim 15, wherein said C3 flag threshold, said C3/C2 flag threshold, and said C3/C16 flag threshold are, respectively, at least about 4.6 μ

M, 0.29, and 1.54.

18. The method of claim 15, wherein said C3 flag threshold, said C3/C2 flag threshold, and said C3/C16 flag threshold are, respectively, about 5.0 μ

M, 0.30, and 1.75.

19. A method comprising steps of:

acquiring mass spectral data from a sample that has been derived from the blood of a patient, wherein said data includes values comprised of a glutaryl carnitine concentration (C5DC) and optionally a molar ratio of glutaryl carnitine and palmitoyl carnitine (C5DC/C16);

comparing said values, respectively, with a C5DC flag threshold and optionally a C5DC/C16 flag threshold;

identifying whether or not there is an elevation of any of said values above any of said flag thresholds; and

interpreting said sample as being normal for glutaric acidemia, provided there is no said elevation of any of said values.

20. The method of claim 19 wherein:

in the acquiring step, said data includes values comprised of a glutaryl carnitine concentration (C5DC) and a molar ratio of glutaryl carnitine and palmitoyl carnitine (C5DC/C16); and

in the comparing step, said C5DC and C5DC/C16 are compared with a C5DC flag threshold and a C5DC/C16 flag threshold, respectively.

21. The method of claim 19 further comprising:

flagging the sample for re-analysis provided any of the following occur;

i. said C5DC is greater than said C5DC flag threshold;

or

ii. said C5DC/C16 is greater than said C5DC/C16 flag threshold and said C5DC is greater than about 0.14 μ

M.

22. The method of claim 21 further comprising:

performing a re-analysis of said sample or a new sample from the same patient to obtain mean values.

23. The method of claim 22 further comprising:

initiating a follow-up protocol based on follow-up criteria utilizing said mean values.

24. The method of claim 19 further comprising:

flagging the sample for re-analysis provided any of the following occur;

i. said C5DC is greater than about 0.4 μ

M;

or

ii. said C5DC is greater than about 0.2 μ

M and said C5DC/C16 is greater than about 0.2 μ

M.

25. The method of claim 24 further comprising:

performing a re-analysis of said sample or a new sample from the same patient to obtain mean values.

26. The method of claim 24 further comprising:

initiating a follow-up protocol based on follow-up criteria utilizing said mean values.

27. The method of claim 19, wherein said C5DC flag threshold and said C5DC/C16 flag threshold are, respectively, at least about 0.13 μ

M and 0.04.

28. The method of claim 19, wherein said C5DC flag threshold and said flat C5DC/C16 flag threshold are, respectively, at least about 0.18 μ

M and 0.06.

29. The method of claim 19, wherein said C5DC flag threshold and said C5DC/C16 flag threshold are, respectively, about 0.17 μ

M and 0.12.

30. The method of claim 19, wherein the sample was derived from a dry blood spot on filter paper.

31. The method of claim 30, wherein the sample was derivatized.

32. The method of claim 19 wherein:

in the acquiring step, said data was acquired using a tandem mass spectrometer.

33. The method of claim 19, wherein:

the sample was derived from a dry blood spot on filter paper;

the sample was derivatized; and

in the acquiring step, said data was acquired using a tandem mass spectrometer.

34. The method of claim 33, wherein said C5DC flag threshold and said C5DC/C16 flag threshold are, respectively, at least about 0.13 μ

M and 0.04.

35. The method of claim 33, wherein said C5DC flag threshold and said C5DC/C16 flag threshold are, respectively, at least about 0.18 μ

M and 0.06.

36. The method of claim 33, wherein said C5DC flag threshold and said C5DC/C16 flag threshold are, respectively, about 0.17 μ

M and 0.12.

37. A method comprising steps of:

acquiring mass spectral data from a sample that has been derived from the blood of a patient, wherein said data includes values comprised of a leucine+isoleucine concentration (leu+ile) and, a valine concentration (val) and optionally one or more of a molar ratio of leucine+isoleucine and phenylalanine (leu+ile/phe), and a molar ratio of leucine+isoleucine and alanine (leu+ile/ala);

comparing said values, respectively, with a leu+ile flag threshold and a val flag threshold and optionally one or more of a leu+ile/phe flag threshold and a leu+ile/ala flag threshold;

identifying whether or not there is an elevation of any of said values above any of said flag thresholds; and

interpreting said sample as being normal for maple syrup urine disease (MSUD), provided there is no said elevation of any of said values.

38. The method of claim 37, wherein:

in the acquiring step, said data includes values comprised of a leucine+isoleucine concentration (leu+ile), a molar ratio of leucine+isoleucine and phenylalanine (leu+ile/phe), and a valine concentration (val); and

in the comparing step, said leu+ile, leu+ile/phe, and val are compared with a leu+ile flag threshold, a leu+ile/phe flag threshold, and a val flag threshold, respectively.

39. The method of claim 37 further comprising:

flagging the sample for re-analysis provided any of the following occur;

i. said leu+ile is greater than about 400 μ

M;

ii. said leu+ile is greater than about 350 μ

M and said val is greater than said val flag threshold;

iii. said leu+ile is greater than said leu+ile flag threshold and said leu+ile/ala is greater than said leu+ile/ala flag threshold;

or

iv. said leu+ile/phe is greater than said leu+ile/phe flag threshold and said val is greater than said val flag threshold.

40. The method of claim 39 further comprising:

performing a re-analysis of said sample or a new sample from the same patient to obtain mean values.

41. The method of claim 40 further comprising:

initiating a follow-up protocol based on follow-up criteria utilizing said mean values.

42. The method of claim 37 further comprising:

flagging the sample for re-analysis provided any of the following occur;

i. said leu+ile is greater than about 500 μ

M;

or

ii. said leu+ile is greater than about 400 μ

M and said leu+ile/phe is greater than said leu+ile/phe flag threshold and said leu+ile/ala is greater than said leu+ile/ala flag threshold.

43. The method of claim 42 further comprising:

performing a re-analysis of said sample or a new sample from the same patient to obtain mean values.

44. The method of claim 43 further comprising:

initiating a follow-up protocol based on follow-up criteria utilizing said mean values.

45. The method of claim 37, wherein said leu+ile flag threshold, said val flag threshold, said leu+ile/phe flag threshold, and said leu+ile/ala flag threshold are, respectively, at least about 229 μ

M, 215 μ

M, 4.36, and 1.19.

46. The method of claim 37, wherein said leu+ile flag threshold, said val flag threshold, said leu+ile/phe flag threshold, and said leu+ile/ala flag threshold are, respectively, at least about 291 μ

M, 288 μ

M, 5.53, and 1.54.

47. The method of claim 37, wherein said leu+ile flag threshold, said val flag threshold, said leu+ile/phe flag threshold, and said leu+ile/ala flag threshold are, respectively, about 325 μ

M, 300 μ

M, 8.0, and 2.25.

48. The method of claim 37, wherein the sample was derived from a dry blood spot on filter paper.

49. The method of claim 48, wherein the sample was derivatized.

50. The method of claim 37, wherein:

in the acquiring step, said data was acquired using a tandem mass spectrometer.

51. The method of claim 37, wherein:

the sample was derived from a dry blood spot on filter paper;

the sample was derivatized; and

in the acquiring step, said data was acquired using a tandem mass spectrometer.

52. The method of claim 51, wherein said leu+ile flag threshold, said val flag threshold, said leu+ile/phe flag threshold, and said leu+ile/ala flag threshold are, respectively, at least about 229 μ

M, 215 μ

M, 4.36, and 1.19.

53. The method of claim 51, wherein said leu+ile flag threshold, said val flag threshold, said leu+ile/phe flag threshold, and said leu+ile/ala flag threshold are, respectively, at least about 291 μ

M, 288 μ

M, 5.53, and 1.54.

54. The method of claim 51, wherein said leu+ile flag threshold, said val flag threshold, said leu+ile/phe flag threshold, and said leu+ile/ala flag threshold are, respectively, about 325 μ

M, 300 μ

M, 8.0, and 2.25.

69. The method of claim 48 further comprising:

flagging the sample for re-analysis provided any of the following occur;

i. said C8is greater than about 1.0 μ

M;

ii. said C8is greater than about 0.5 μ

M and said C8/C16 is greater than about 0.35;

or

iii. said C6 is greater than about 0.3 μ

M and said C10;

1 is greater than about 0.3 μ

M.

70. The method of claim 69 further comprising:

performing a re-analysis of said sample or a new sample from the same patient to obtain mean values.

71. The method of claim 70 further comprising:

initiating a follow-up protocol based on follow-up criteria utilizing said mean values.

55. A method comprising steps of:

acquiring mass spectral data from a sample that has been derived from the blood of a patient, wherein said data includes values comprised of a leucine+isoleucine concentration (leu+ile) and a molar ratio of leucine+isoleucine and alanine (leu+ile/ala); and

comparing said values, respectively, with a leu+ile flag threshold and a leu+ile/ala flag threshold;

identifying whether or not there is an elevation of any of said values above any of said flag thresholds; and

interpreting said sample as being normal for maple syrup urine disease (MSUD), provided there is no said elevation of any of said values.

56. The method of claim 55, wherein:

the sample was derived from a dry blood spot on filter paper;

the sample was derivatized; and

in the acquiring step, said data was acquired using a tandem mass spectrometer.

57. The method of claim 56, wherein said leu+ile flag threshold, said val flag threshold, said leu+ile/phe flag threshold, and said leu+ile/ala flag threshold are, respectively, at least about 229 μ

M, 215 μ

M, 4.36, and 1.19.

58. The method of claim 56, wherein said leu+ile flag threshold, said val flag threshold, said leu+ile/phe flag threshold, and said leu+ile/ala flag threshold are, respectively, at least about 291 μ

M, 288 μ

M, 5.53, and 1.54.

59. The method of claim 56, wherein said leu+ile flag threshold, said val flag threshold, said leu+ile/phe flag threshold, and said leu+ile/ala flag threshold are, respectively, about 325 μ

M, 300 μ

M, 8.0, and 2.25.

60. A method comprising steps of:

acquiring mass spectral data from a sample that has been derived from the blood of a patient, wherein said data includes values comprised of a leucine+isoleucine concentration (leu+ile), a molar ratio of leucine+isoleucine and phenylalanine (leu+ile/phe), and a molar ratio of leucine+isoleucine and alanine (leu+ile/ala); and

comparing said values, respectively, with a leu+ile flag threshold, a leu+ile/phe flag threshold, and a leu+ile/ala flag threshold;

identifying whether or not there is an elevation of any of said values above any of said flag thresholds; and

interpreting said sample as being normal for maple syrup urine disease (MSUD), provided there is no said elevation of any of said values.

61. A method comprising steps of:

acquiring mass spectral data from a sample that has been derived from the blood of a patient, wherein said data includes values comprised of an octanoyl carnitine concentration (C8) and one or more of a molar ratio of octanoyl carnitine and palmitoyl carnitine (C8/C16), a hexanoyl carnitine concentration (C6), a decanoyl carnitine concentration (C10;

1), and a decanoyl carnitine concentration (C10);

comparing said values, respectively, with one or more of a C8 flag threshold and one or more of a C8/C16 flag threshold, a C6 flag threshold, a C10;

1 flag threshold, and a C10 flag threshold;

identifying whether or not there is an elevation of any of said values above any of said flag thresholds; and

interpreting said sample as being normal for medium-chain acyl-coenzyme A dehydrogenase (MCAD) deficiency, provided there is no said elevation of any of said values.

62. The method of claim 61, wherein:

in the acquiring step, said data includes values comprised of an octanoyl carnitine concentration (C8) and a molar ratio of octanoyl carnitine and palmitoyl carnitine (C8/C16); and

in the comparing step, said C8and C8/C16 are compared with a C8 flag threshold and a C8/C16 flag threshold, respectively.

63. The method of claim 61, wherein:

in the acquiring step, said data includes values comprised of an octanoyl carnitine concentration (C8) and a hexanoyl carnitine concentration (C6); and

in the comparing step, said C8and C6are compared with a C8 flag threshold and a C6 flag threshold, respectively.

64. The method of claim 61 wherein:

in the acquiring step, said data includes values comprised of an octanoyl carnitine concentration (C8), a decanoyl carnitine concentration (C10), and a decenoyl carnitine concentration (C10;

1); and

in the comparing step, said C8, C10, and C10;

1 are compared with a C8 flag threshold, a C10 flag threshold, and a C10;

1 flag threshold, respectively.

65. The method of claim 61, wherein:

in the acquiring step, said data includes values comprised of an octanoyl carnitine concentration (C8), a hexanoyl carnitine concentration (C6), and a decenoyl carnitine concentration (C10;

1); and

in the comparing step, said C8, C6, and C10;

1 are compared with a C8 flag threshold, a C6 flag threshold, and a C10;

1 flag threshold, respectively.

66. The method of claim 61 further comprising:

flagging the sample for re-analysis provided any of the following occur;

i. said C8is greater than or equal to about 0.4 μ

M;

ii. said C8 is greater than about 0.3 μ

M and said C8/C16 is greater than about 0.15;

iii. said C8is greater than about 0.3 μ

M and said C6is greater than about 0.2 μ

M;

or

iv. said C10;

1 is greater than about 0.2 μ

M and said C10 is greater than about 0.3 μ

M.

67. The method of claim 66 further comprising:

performing a re-analysis of said sample or a new sample from the same patient to obtain mean values.

68. The method of claim 67 further comprising:

initiating a follow-up protocol based on follow-up criteria utilizing said mean values.

72. The method of claim 61, wherein said C8 flag threshold, said C8/C16 flag threshold, said C6 flag threshold, said C10:

1 flag threshold, and said C10 flag threshold are, respectively, at least about 0.19 μ

M, 0.08, 0.16 μ

M, 0.18 μ

M, and 0.31 μ

M.

73. The method of claim 61 wherein said C8 flag threshold, said C8/C16 flag threshold, said C6 flag threshold, said C10:

1 flag threshold, and said C10 flag threshold are, respectively, at least about 0.26 μ

M, 0.12, 0.22 μ

M, 0.24 μ

M, and 0.42 μ

M.

74. The method of claim 61 wherein said C8 flag threshold, said C8/C16 flag threshold, said C6 flag threshold, said C10:

1 flag threshold, and said C10 flag threshold are, respectively, about 0.35 μ

M, 0.28, 0.16 μ

M, 0.32 μ

M, and 0.42 μ

M.

75. The method of claim 61 wherein the sample was derived from a dry blood spot on filter paper.

76. The method of claim 75 wherein the sample was derivatized.

77. The method of claim 61, wherein:

in the acquiring step, said data was acquired using a tandem mass spectrometer.

78. The method of claim 61, wherein:

the sample was derived from a dry blood spot on filter paper;

the sample was derivatized; and

in the acquiring step, said data was acquired using a tandem mass spectrometer.

79. The method of claim 78, wherein said C8 flag threshold, said C8/C16 flag threshold, said C6 flag threshold, said C10:

1 flag threshold, and said C10 flag threshold are, respectively, at least about 0.19 μ

M, 0.08, 0.16 μ

M, 0.18 μ

M, and 0.31 μ

M.

80. The method of claim 78, wherein said C8 flag threshold, said C8/C16 flag threshold, said C6 flag threshold, said C10:

1 flag threshold, and said C10 flag threshold are, respectively, at least about 0.26 μ

M, 0.12, 0.22 μ

M, 0.24 μ

M, and 0.42 μ

M.

81. The method of claim 78, wherein said C8 flag threshold, said C8/C16 flag threshold, said C6 flag threshold, said C10:

1 flag threshold, and said C10 flag threshold are, respectively, about 0.35 μ

M, 0.28, 0.16 μ

M, 0.32 μ

M, and 0.42 μ

M.

82. A method comprising steps of:

acquiring mass spectral data from a sample that has been derived from the blood of a patient, wherein said data includes values comprised of one or more of a saturated myristoyl carnitine concentration (C14), an unsaturated myristoyl carnitine concentration (C14;

1), and a molar ratio of myristoyl carnitine and palmitoyl carnitine (C14;

1/C16);

comparing said values, respectively, with one or more of a C14 flag threshold, a C14;

1 flag threshold, and a C14;

1/C16 flag threshold;

identifying whether or not there is an elevation of any of said values above any of said flag thresholds; and

interpreting said sample as being normal for VLCAD, provided there is no said elevation of any of said values.

83. The method of claim 82, wherein:

in the acquiring step, said data includes a saturated myristoyl carnitine concentration (C14); and

in the comparing step, said C14 is compared with a C14 flag threshold.

84. The method of claim 82, wherein:

in the acquiring step, said data includes an unsaturated myristoyl carnitine concentration (C14;

1); and

in the comparing step, said C14;

1 is compared with a C14;

1 flag threshold.

85. The method of claim 82, wherein:

in the acquiring step, said data includes a molar ratio of myristoyl carnitine and palmitoyl carnitine (C14;

1/C16); and

in the comparing step, said C14;

1/C16 is compared with a C14;

1/C16 flag threshold.

86. The method of claim 82, wherein:

in the acquiring step, said data includes values comprised of a saturated myristoyl carnitine concentration (C14) and a molar ratio of myristoyl carnitine and palmitoyl carnitine (C14;

1/C16); and

in the comparing step, said C14 and C14;

1/C16 are compared with a C14 flag threshold and a C14;

1/C16 flag threshold, respectively.

87. The method of claim 82, wherein:

in the acquiring step, said data includes values comprised of an unsaturated myristoyl carnitine concentration (C14;

1) and a molar ratio of myristoyl carnitine and palmitoyl carnitine (C14;

1/C16); and

in the comparing step, said C14;

1 and C14;

1/C16 are compared with a C14;

1 flag threshold and a C14;

1/C16 flag threshold, respectively.

88. The method of claim 82, wherein:

in the acquiring step, said data includes values comprised of a saturated myristoyl carnitine concentration (C14), an unsaturated myristoyl carnitine concentration (C14;

1) and a molar ratio of myristoyl carnitine and palmitoyl carnitine (C14;

1/C16); and

in the comparing step, said C14, C14;

1, and C14;

1/C16 are compared with a C14 flag threshold, a C14;

1 flag threshold, and a C14;

1/C16 flag threshold, respectively.

89. The method of claim 82 further comprising:

flagging the sample for re-analysis provided any of the following occur;

i. said C14 is greater than said C14 flag threshold;

ii. said C14;

1 is greater than said C14;

1 flag threshold;

or

iii. said C14 is greater than about 0.75 μ

M or said C14;

1 is greater than about 0.65 μ

M and said C14;

1/C16 is greater than about 0.24.

90. The method of claim 89 further comprising:

performing a re-analysis of said sample or a new sample from the same patient to obtain mean values.

91. The method of claim 90 further comprising:

initiating a follow-up protocol based on follow-up criteria utilizing said mean values.

92. The method of claim 82 further comprising:

flagging the sample for re-analysis provided any of the following occur;

i. said C14 is greater than about 2.0 μ

M

ii. said C14;

1 is greater than about 1.5 μ

M;

or

ii. said C14 is greater than about 1.5 μ

M and said C14;

1 is greater than about 1.0 μ

M and said C14;

1/C16 is greater than about 0.3.

93. The method of claim 92 further comprising:

performing a re-analysis of said sample or a new sample from the same patient to obtain mean values.

94. The method of claim 93 further comprising:

initiating a follow-up protocol based on follow-up criteria utilizing said mean values.

95. The method of claim 82, wherein said C14 flag threshold, said C14:

1 flag threshold, and said C14;

1/C16 flag threshold are, respectively, at least about 0.45 μ

M, 0.28 μ

M, and 0.11.

96. The method of claim 82, wherein said C14 flag threshold, said C14:

1 flag threshold, and said C14;

1/C16 flag threshold are, respectively, at least about 0.56 μ

M, 0.38 μ

M, and 0.17.

97. The method of claim 82, wherein said C14 flag threshold, said C14:

1 flag threshold, and said C14;

1/C16 flag threshold are, respectively, about 0.85 μ

M, 0.70 μ

M, and 0.24.

98. The method of claim 82, wherein the sample was derived from a dry blood spot on filter paper.

99. The method of claim 95 wherein the sample was derivatized.

100. The method of claim 82, wherein:

in the acquiring step, said data was acquired using a tandem mass spectrometer.

101. The method of claim 82, wherein:

the sample was derived from a dry blood spot on filter paper;

the sample was derivatized; and

in the acquiring step, said data was acquired using a tandem mass spectrometer.

102. The method of claim 101, wherein said C14 flag threshold, said C14:

1 flag threshold, and said C14;

1/C16 flag threshold are, respectively, at least about 0.45 μ

M, 0.28 μ

M, and 0.11.

103. The method of claim 101, wherein said C14 flag threshold, said C14:

1 flag threshold, and said C14;

1/C16 flag threshold are, respectively, at least about 0.56 μ

M, 0.38 μ

M, and 0.17.

104. The method of claim 101, wherein said C14 flag threshold, said C14:

1 flag threshold, and said C14;

1/C16 flag threshold are, respectively, about 0.85 μ

M, 0.70 μ

M, and 0.24.

105. A method comprising steps of:

acquiring mass spectral data from a sample that has been derived from the blood of a patient, wherein said data includes values comprised of one or more of a hydroxyisovalerylcarnitine concentration (C5OH) and a tiglylcarnitine concentration (C5;

1);

comparing said values, respectively, with one or more of a C5OH flag threshold and a C5;

1 flag threshold;

identifying whether or not there is an elevation of any of said values above any of said flag thresholds; and

interpreting said sample as being normal for crotonyl co-A carboxylase deficiency, provided there is no said elevation of any of said values.

106. The method of claim 105, wherein:

in the acquiring step, said data includes a hydroxyisovalerylcarnitine concentration (C5OH); and

in the comparing step, said C5OH is compared with a C5OH flag threshold.

107. The method of claim 105, wherein:

in the acquiring step, said data includes a tiglylcarnitine concentration (C5;

1); and

in the comparing step, said C5;

1 is compared with a C5;

1 flag threshold.

108. The method of claim 105 further comprising:

flagging the sample for re-analysis provided any of the following occur;

i. said C5OH is greater than or equal to said C5OH flag threshold;

or

ii. said C5;

1 is greater than or equal to said C5;

1 flag threshold.

109. The method of claim 108 further comprising:

performing a re-analysis of said sample or a new sample from the same patient to obtain mean values.

110. The method of claim 109 further compising:

initiating a follow-up protocol based on follow-up criteria utilizing said mean values.

111. The method of claim 105 further comprising:

flagging the sample for re-analysis provided any of the following occur;

i. said C5OH is greater than about 3.0 μ

M;

or

ii. said C5;

1 is greater than about 1.0 μ

M.

112. The method of claim 111 further comprising:

performing a re-analysis of said sample or a new sample from the same patient to obtain mean values.

113. The method of claim 112 further comprising:

initiating a follow-up protocol based on follow-up criteria utilizing said mean values.

114. The method of claim 105, wherein said C5OH flag threshold and said C5:

1 flag threshold are, respectively, at least about 0.42 μ

M and 0.09 μ

M.

115. The method of claim 105, wherein said C5OH flag threshold and said C5:

1 flag threshold are, respectively, at least about 0.57 μ

M and 0.13 μ

M.

116. The method of claim 105, wherein said C5OH flag threshold and said C5:

1 flag threshold are, respectively, about 0.85 μ

M and 0.35 μ

M.

117. The method of claim 105, wherein the sample was derived from a dry blood spot on filter paper.

118. The method of claim 117 wherein the sample was derivatized.

119. The method of claim 105 wherein:

in the acquiring step, said data was acquired using a tandem mass spectrometer.

120. The method of claim 105, wherein:

the sample was derived from a dry blood spot on filter paper;

the sample was derivatized; and

in the acquiring step, said data was acquired using a tandem mass spectrometer.

121. The method of claim 120, wherein said C5OH flag threshold and said C5:

1 flag threshold are, respectively, at least about 0.42 μ

M and 0.09 μ

M.

122. The method of claim 120, wherein said C5OH flag threshold and said C5:

1 flag threshold are, respectively, at least about 0.57 μ

M and 0.13 μ

M.

123. The method of claim 120, wherein said C5OH flag threshold and said C5:

1 flag threshold are, respectively, about 0.85 μ

M and 0.35 μ

M.