Method of Determining Hypoxia

• US 20080213744A1
• Filed: 04/11/2008
• Published: 09/04/2008
• Est. Priority Date: 10/13/2006
• Status: Active Grant

• Current Assignee: Calmark Sweden AB
• Original Assignee: Calmark Sweden AB
• Inventors: Ornas, Sofia Hirot af, Karlsson, Mathias
• Granted Patent: US 8,945,824 B2
• Time in Patent Office: Days
• US Class Current: 435/2
• CPC Class Codes: A61B 10/0045 Devices for taking samples of body liquids devices for taking blood samples A61B5/15
• CPC Class Codes: A61B 10/0048 for taking amniotic fluid samples
• CPC Class Codes: A61B 2503/02 Foetus
• CPC Class Codes: A61B 5/1405 Devices for taking blood samples devices for taking samples of other body liquids A61B10/0045
• CPC Class Codes: A61B 5/14542 for measuring blood gases A61B5/14551 takes precedence
• CPC Class Codes: A61B 5/14546 for measuring analytes not otherwise provided for, e.g. ions, cytochromes
• CPC Class Codes: A61B 5/150015 Source of blood
• CPC Class Codes: A61B 5/157 Devices characterised by integrated means for measuring characteristics of blood
• CPC Class Codes: A61B 5/412 Detecting or monitoring sepsis
• CPC Class Codes: A61B 5/4362 Assessing foetal parameters
• CPC Class Codes: A61F 7/00 Heating or cooling appliances for medical or therapeutic treatment of the human body heating or cooling means in connection with bedsteads or mattresses A47C21/00 ; massage, therapeutic baths A61H; hyperthermia using electric or magnetic fields or ultrasound A61N; electric heating elements H05B
• CPC Class Codes: C12Q 1/32 involving dehydrogenase
• CPC Class Codes: C12Q 1/52 involving transaminase
• CPC Class Codes: G01N 2333/904 acting on CHOH groups as donors, e.g. glucose oxidase, lactate dehydrogenase (1.1)
• CPC Class Codes: G01N 2333/91188 transferring nitrogenous groups (2.6)
• CPC Class Codes: G01N 2800/2871 Cerebrovascular disorders, e.g. stroke, cerebral infarct, cerebral haemorrhage, transient ischemic event
• CPC Class Codes: G01N 2800/7038 Hypoxia
• CPC Class Codes: G01N 33/492 Determining multiple analytes
• CPC Class Codes: G01N 33/54386 Analytical elements
• CPC Class Codes: G01N 33/573 for enzymes or isoenzymes

Abstract

A method of determining hypoxia in fetal scalp blood sampled during labor comprises the determination of total lactate dehydrogenase (LDH) in plasma obtained from the sample. The method can comprise the additional determination of K, Mg, Ca, AST, ALT, lactate in the plasma and/or blood. Increased values of one or more of LDH, Mg, Ca, AST, ALT, lactate are indicative of hypoxia in the fetus. Also disclosed is the use of a plasma separation apparatus in the method.

Claims

1. A method of assessing hypoxia at a tissue site of a mammal, comprising:

a. collecting a blood sample from the tissue site, said blood sample comprising plasma and blood cells;

b. separating the plasma from the blood cells;

c. determining the amount of LDH in the plasma; and

d. assessing the presence of hypoxia at the tissue site from the amount of LDH in the plasma.

2. The method of claim 1, further comprising determining the amount of at least one additional prognostic marker in the plasma selected from the group consisting essentially of K, Mg, Ca, AST, ALT and lactate.

3. The method of claim 1, wherein the volume of the blood sample comprises from 5 μ

L to 150 μ

L.

4. The method of claim 1, wherein the volume of the blood sample comprises from 5 μ

L to 25 μ

L

5. The method of claim 1, wherein the volume of the blood sample comprises from about 5 to 15 μ

L.

6. The method of claim 1, wherein the plasma is separated from the blood cells by means of a semi-permeable membrane such that the plasma passes through the membrane and the blood cells are retained on the membrane.

7. The method of claim 1, wherein the blood sample is collected from a mammal'"'"'s bowel.

8. The method of claim 7, wherein the blood sample is collected prior to a medical procedure and further comprising:

a. collecting a second blood sample after a medical procedure, said blood sample comprising plasma and blood cells;

b. separating the plasma from the blood cells of the second blood sample; and

c. determining the total amount of LDH in the plasma of the second blood sample.

9. The method of claim 8, further comprising determining the amount of at least one additional prognostic marker in the plasma of both blood samples selected from the group consisting essentially of K, Mg, Ca, AST, ALT and lactate.

10. The method of claim 9, further comprising comparing the amount of the prognostic markers in the blood samples.

11. The method of claim 8, wherein the medical procedure comprises anastomosis of the gastrointestinal tract.

12. The method of claim 1, wherein the blood sample is collected from one or more of a specific organ, cerebrospinal fluid, urine, or intraperitoneal fluid of a mammal during a surgical procedure.

13. The method of claim 12, wherein the organ comprises a mammal'"'"'s aorta.

14. The method of claim 12, further comprising determining the amount of at least one additional prognostic marker in the plasma selected from the group consisting essentially of K, Mg, Ca, AST, ALT and lactate.

15. The method of claim 14, wherein the at least one prognostic marker includes lactate.

16. The method of claim 1, wherein the blood sample is collected from an organ to be transplanted into a mammal of need for said organ.

20. The method of claim 2, further comprising determining the amount of at least one additional prognostic marker in the plasma selected from the group consisting essentially of K, Mg, Ca, AST, ALT and lactate.

21. The method of claim 20, wherein the determination of the prognostic markers allows an individual to assess the health of a mammal'"'"'s liver.

22. The method of claim 1, wherein the determination of hypoxia allows the prediction of brain injury after prenatal asphyxia.

23. The method of claim 22, further comprising determining the amount of at least one additional prognostic marker in the plasma selected from the group consisting essentially of AST, ALT and lactate.

24. The method of claim 22, further comprising determining the amount of ALT in the plasma.

25. The method of claim 22, further comprises providing hypothermia treatment to a mammal in need thereof, wherein the determination of LDH in the plasma indicate delayed cell death in a mammal'"'"'s brain.

26. The method of claim 1, wherein determining the total amount of LDH in the plasma allows the assessment of blood circulation to a mammal'"'"'s limbs before, during, and after a medical or surgical treatment.

17. The method of claim 17, wherein the blood sample is collected prior to transplantation surgery.

18. The method of claim 17, wherein the organ comprises a liver.

19. The method of claim 17, further comprising determining the amount of at least one additional prognostic marker in the plasma selected from the group consisting essentially of K, Mg, Ca, AST, ALT and lactate.

27. A method of determining acute hypoxia in fetal scalp blood, comprising collecting a blood sample during labor and treating the blood sample to look for at least one indication to determine hypoxia characterized in that it comprises the determination of total lactate dehydrogenase (LDH) in plasma obtained from the sample.

28. The method of claim 27, wherein the sample is from 5 μ

L to 60 μ

L.

29. The method of claim 27, wherein the sample is from 5 μ

L to 25 μ

L.

30. The method of claim 27, wherein the sample is about 5 to 15 μ

L.

31. The method of claim 27, comprising separating plasma from blood cells by means of a semi-permeable membrane and/or filter.

32. The method of claim 31, wherein the membrane has an area of from 5 mm to 1000 mm², preferably less than 100 mm².

33. The method of claim 31, comprising the steps of loading one face of the membrane with a sample, applying negative pressure to the other face of the membrane, and collecting plasma in a compartment facing said other face.

34. The method of claim 33, comprising the step of transferring the plasma from the compartment to an apparatus and/or device for determination of LDH.

35. The method of claim 33, comprising the step of determining LDH in the plasma disposed in the compartment.

36. The method of claim 31, wherein the membrane is disposed on or integrated with a porous matrix into which plasma migrates attracted by capillary forces.

37. The method of claim 36, wherein the membrane is removed from the matrix after a given time to determine LDH in the matrix.

38. The method of claim 27, comprising the steps of separating plasma from blood cells in the sample transferring the supernatant plasma to an apparatus for determination of LDH.

39. The method of claim 27, further comprising the comparison of LDH so determined with LDH determined in plasma obtained from fetal scalp blood sampled during partum from a fetus in a non-hypoxic state.

40. The method of claim 39, wherein a statistically significant increased LDH level is indicative of hypoxia.

41. The method of claim 39, wherein a statistically significant LDH level increased by a factor of three or more is indicative of a risk of hypoxic-ischemic encephalopathy.

42. The method of claim 27, additionally comprising the determination of one or several members of the group consisting of K, Mg, Ca, AST, ALT, and lactate in said scalp blood and/or said plasma.

43. The method of claim 42, wherein a statistically significant increased level of any of K, Mg, Ca, AST, ALT, and lactate in combination with a statistically significant increased level of LDH increased is indicative of hypoxia.

44. The method according to claim 27 characterized in that sample is collected on a disposable device including means for separation of plasma and a compartment arranged to perform detection of LDH.

45. Use of an apparatus for separation of plasma from blood cells in the method of claim 27, the apparatus comprising a housing, a compartment in the housing, a first opening in the housing covered by a semi-permeable membrane, a second opening in the housing communicating the compartment with a means of negative pressure or capillary force via a conduit, and if negative pressure a valve means for controlling said communication.

46. The use of claim 45, wherein the valve means is disposed in the conduit, wherein preferably the compartment additionally comprises a second conduit arranged to communicate with a bottom section thereof and provided with a second valve means for controlled emptying of plasma accumulated at the bottom of the compartment.

47. The use of claim 45, wherein the compartment houses or is in communication with a means for determining LDH in the plasma accumulated in the housing.

48. Testing system for performing the method according to claim 27, said testing system comprising a disposable device having a blood sample collecting portion, a plasma separation device and a compartment arranged to indirectly or directly determine the total lactate dehydrogenase (LDH) in the plasma.

49. Testing system according to claim 48, characterized in that said disposable device has a flat-shaped body, wherein said receiving portion forms a compartment and interface for capillary sample collector.

50. Testing system according to claim 49, characterized in that there is a number of detection compartments arranged on the card, wherein preferably each one of said compartments is connected to an upstream compartment with a reagent.

51. Testing system according to claim 48, characterized in that there is arranged a sample splitter in between the said inter-face and said plasma separation device.

52. Testing system according to claim 49, characterized in that said compartment is in the form of an optical detection cell and that the body is formed to fit into an instrument arranged with at least one device for optical spectrometry.

53. Testing system according to claim 48, characterized in that said body is arranged with a unique code and preferably that said instrument is arranged with a reader.

54. Testing system according to claim 48, characterized in that said disposable device includes a compartment arranged with dry chemical means for visual detection.