Apparatus and method for separating and concentrating fluids containing multiple components

• US 9,897,589 B2
• Filed: 10/09/2014
• Issued: 02/20/2018
• Est. Priority Date: 05/24/2002
• Status: Active Grant

• Current Assignee: Biomet Manufacturing, LLC (Zimmer Biomet Holdings, Inc.)
• Original Assignee: Biomet Biologics LLC (Zimmer Biomet Holdings, Inc.)
• Inventors: Woodell-May, Jennifer E.
• Primary Examiner(s): Mellon, David C
• Application Number:
  US14/510,828
• Publication Number:
  US 20150023939A1
• Time in Patent Office: 1,230 Days
• CPC Class Codes: A61K 2300/00 Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00
• CPC Class Codes: A61K 35/14 Blood; Artificial blood perfluorocarbons A61K31/02; umbilical cord blood A61K35/51; haemoglobin A61K38/42
• CPC Class Codes: A61K 35/17 Lymphocytes; B-cells; T-cells; Natural killer cells; Interferon-activated or cytokine-activated lymphocytes when activated by a specific antigen A61K39/00
• CPC Class Codes: A61K 35/19 Platelets; Megacaryocytes
• CPC Class Codes: A61K 35/28 Bone marrow; Haematopoietic stem cells; Mesenchymal stem cells of any origin, e.g. adipose-derived stem cells
• CPC Class Codes: B01D 21/262 by using a centrifuge
• CPC Class Codes: B01D 21/307 Passive control mechanisms without external energy, e.g. using a float
• CPC Class Codes: B01L 1/52 Transportable laboratories; Field kits
• CPC Class Codes: B01L 2200/026 Fluid interfacing between devices or objects, e.g. connectors, inlet details
• CPC Class Codes: B01L 2300/0681 Filter
• CPC Class Codes: B01L 2400/0409 centrifugal forces
• CPC Class Codes: B01L 2400/0478 pistons
• CPC Class Codes: B01L 2400/0605 check valves
• CPC Class Codes: B01L 3/502 with fluid transport, e.g. in multi-compartment structures centrifugal-type cuvettes G01N21/07; analysis by separation into components G01N30/00; automatic analysers G01N35/00
• CPC Class Codes: B01L 3/5021 Test tubes specially adapted for centrifugation purposes centrifuges B04B5/04
• CPC Class Codes: B01L 3/50215 using a float to separate phases
• CPC Class Codes: B01L 9/54 Supports specially adapted for pipettes and burettes automated pipetting stations G01N35/10
• CPC Class Codes: G01N 33/491 by separating the blood components G01N15/05 takes precedence

Abstract

An apparatus that allows for separating and collecting a fraction of a sample. The apparatus, when used with a centrifuge, allows for the creation of at least three fractions in the apparatus. It also provides for a new method of extracting the buffy coat phase from a whole blood sample. A buoy system that may include a first buoy portion and a second buoy member operably interconnected may be used to form at least three fractions from a sample during a substantially single centrifugation process. Therefore, the separation of various fractions may be substantially quick and efficient.

Claims

1. A method of isolating a fraction comprising nucleated cells, comprising:

obtaining a volume of whole blood from a patient through venipuncture;

obtaining a volume of bone marrow aspirate from the patient;

loading the volume of whole blood and the volume of bone marrow aspirate into a container configured to be placed in a centrifuge, the container comprising a buoy having a selected density operable to physically separate a plurality of fractions of whole blood based on the density of the fractions, the fractions including the fraction comprising nucleated cells;

centrifuging the container, so as to create the fraction comprising nucleated cells; and

collecting the fraction comprising the nucleated cells.

2. The method according to claim 1, wherein the fraction comprising nucleated cells comprises cells selected from the group consisting of platelets, white blood cells, stem cells, and mixtures thereof.

3. The method according to claim 1, wherein the centrifuging creates at least three distinct fractions comprising a red blood cell fraction, a buffy coat fraction, and a plasma fraction.

4. The method according to claim 1, wherein the buoy is configured to have a density less than the density of red blood cells and a density greater than the density of the fraction comprising nucleated cells.

5. The method according to claim 1, wherein the container further comprises an extraction port, and the collecting of the fraction comprising nucleated cells comprises affixing a syringe to the extraction port and drawing the fraction comprising nucleated cells into the syringe.

6. The method according to claim 1, further comprising administering the fraction comprising nucleated cells to the patient.

7. A method of isolating a fraction comprising nucleated cells from a multi-component fluid comprising nucleated cells, comprising:

obtaining a volume of whole blood from a patient through venipuncture;

obtaining a volume of bone marrow aspirate from the patient;

loading the volume of whole blood and the volume of bone marrow aspirate into a container of a separator configured to be placed in a centrifuge, the volume of whole blood and the volume of bone marrow combining to form a multi-component fluid;

centrifuging the separator to separate the multi-component fluid into at east three fractions, the fractions including the fraction comprising nucleated cells; and

collecting the fraction comprising nucleated cells.

8. The method according to claim 7, wherein the three fractions are a fraction comprising red blood cells, a fraction comprising buffy coat, and a fraction comprising plasma.

9. The method according to claim 7, wherein the fraction comprising nucleated cells comprises cells selected from the group consisting of platelets, white blood cells, stem cells, and mixtures thereof.

10. The method according to claim 8, wherein the separator comprises a buoy and a piston.

11. The method according to claim 10, wherein the buoy is configured to have a density less than the density of red blood cells.

12. The method according to claim 11, wherein the buoy is configured to provide for isolation of the buffy coat and undifferentiated bone marrow cells.

13. The method according to claim 11, wherein the buoy is configured to have a density greater than the density of the fraction comprising nucleated cells.

14. The method according to claim 7, further comprising administering the fraction comprising nucleated cells to a patient.

15. A method of treating a patient with a biological material comprising undifferentiated nucleated cells, comprising:

obtaining a volume of whole blood from the patient through venipuncture;

obtaining a volume of bone marrow aspirate from the patient;

loading the volume of whole blood and the volume of bone marrow aspirate into a container of a separator configured to be placed in a centrifuge, the volume of whole blood and the volume of bone marrow aspirate combining to form a multi-component fluid, the separator being operable to separate the multi-component fluid into at least three fractions, the fractions including a fraction comprising undifferentiated nucleated cells, wherein the separator comprises a buoy with a density less than the density of a fraction of red blood cells;

centrifuging the separator;

collecting the biological material comprising the fraction comprising undifferentiated nucleated cells, wherein the fraction comprising undifferentiated nucleated cells further comprises a blood component selected from the group consisting of platelets, white blood cells, and mixtures thereof; and

administering the fraction comprising undifferentiated nucleated cells to the patient.

16. The method according to claim 15, wherein centrifuging the container separates the mixture of bone marrow aspirate and whole blood into three fractions based on density, wherein said fractions, in decreasing order of density, comprise:

i. a fraction rich in blood cells;

ii. a buffy coat fraction;

iii. a plasma fraction; and

the collecting comprises isolating the buffy coat fraction alone or in combination with all or part of the plasma fraction to form the biological material.

17. The method according to claim 15, wherein the separator further comprises a piston.

18. The method according to claim 15, wherein the separator further comprises an extraction port;

and the collecting of the fraction comprising undifferentiated nucleated cells comprises affixing a syringe to the extraction port and drawing the fraction comprising undifferentiated nucleated cells into the syringe.

20. The method according to claim 18, wherein the biological sample is loaded into a tube of a separation device comprising an extraction port and a buoy moveable within the tube, and centrifuging comprises centrifuging the separation device.

19. A method for obtaining a biological material comprising a bone marrow fraction, comprising:

a. centrifuging a biological sample comprising bone marrow aspirate obtained from a patient and whole blood obtained from the patient through venipuncture to provide a separation of components of the sample based on density, said separation providing the following fractions in decreasing order of density;

i. a fraction rich in blood cells;

ii. a buffy coat fraction;

iii. a plasma fraction; and

b. isolating the huffy coat fraction alone or in combination with all or part of the plasma fraction to form the biological material.