Devices and methods for infrared (IR) based quantitation of biomolecules

• US 9,018,584 B2
• Filed: 03/08/2012
• Issued: 04/28/2015
• Est. Priority Date: 04/14/2011
• Status: Active Grant

• Current Assignee: EMD Millipore Corporation (Merck & Co., Inc.)
• Original Assignee: EMD Millipore Corporation (Merck & Co., Inc.)
• Inventors: Chernokalskaya, Elena, Joshi, Vivek, Clark, Phillip, Utzat, Christopher, Amara, Ryan, Rider, Timothy Scott
• Primary Examiner(s): Porta, David
• Assistant Examiner(s): BOOSALIS, FANI POLYZOS
• Application Number:
  US13/415,400
• Publication Number:
  US 20130062523A1
• Time in Patent Office: 1,146 Days
• Field of Search: 250/339.07, 738/649.1
• US Class Current: 250/339.07
• CPC Class Codes: G01J 5/0255 Sample holders for pyrometry; Cleaning of sample using a gas purge G01J5/051
• CPC Class Codes: G01N 2021/3572 Preparation of samples, e.g. salt matrices
• CPC Class Codes: G01N 21/25 Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
• CPC Class Codes: G01N 21/3563 for analysing solids; Preparation of samples therefor
• CPC Class Codes: G01N 21/3577 for analysing liquids, e.g. polluted water
• CPC Class Codes: G01N 21/359 using near infrared light

Abstract

The present invention provides methods for quantitating one or more biomolecules in a sample using IR based techniques, sample holder devices for use in such methods as well as methods for manufacturing such sample holder devices.

Claims

1. A method for quantitation of one or more biomolecules in a sample, the method comprising the steps of:

(a) providing a sample holder comprising a porous membrane which comprises a hydrophilic region surrounded by a hydrophobic region for sample containment;

(b) contacting the hydrophilic region of the membrane with a sample volume, wherein the hydrophilic region has a diameter;

(c) drying the sample volume on the membrane;

(d) exposing the sample volume on the membrane to an infrared beam having a diameter equal to or larger than the diameter of the hydrophilic region, and comprising a wavelength in the spectral range of 4000-400 cm<sup>−

1</sup> or any portion of the spectral range of 4000-400 cm<sup>−

1</sup>, thereby to obtain an infrared absorption spectrum;

wherein one or more absorption peak areas in the infrared absorption spectrum correlates with the quantity of the one or more biomolecules in the sample.

2. The method of claim 1, wherein the one or more biomolecules is selected from the group consisting of nucleic acids, proteins, lipids, polysaccharides and lipopolysaccharides.

3. The method of claim 2, wherein the lipopolysaccharide is an endotoxin.

4. The method of claim 1, wherein the method does not require a user to generate a calibration curve each time a sample is analyzed for quantitation of the one or more biomolecules.

5. The method of claim 1, wherein sample volume ranges from 0.1-20 μ

l.

6. The method of claim 5, wherein sample volume is 1 μ

l or less.

7. The method of claim 1, wherein the porous membrane is contained within a device.

8. The method of claim 7, wherein the device is a sample holder card.

9. The method of claim 8, wherein the sample holder card comprises a porous membrane which comprises an area within which the sample is contained on the membrane.

10. The method of claim 9, wherein the area for sample containment on the membrane comprises a hydrophilic region surrounded by a hydrophobic region.

11. The method of claim 10, wherein the hydrophobic region is created by plasma treatment of a hydrophilic porous membrane.

12. The method of claim 10, wherein the sample is contained within the hydrophilic region.

13. The method of claim 10, wherein the hydrophobic region is created by heat treatment of a hydrophilic porous membrane.

14. The method of claim 1, wherein the sample comprises a biological fluid.

15. The method of claim 14, wherein the biological fluid is selected from the group consisting of blood, plasma, serum and urine.

16. The method of claim 1, wherein the sample comprises cell or tissue lysate.

17. The method of claim 1, wherein the sample is a crude sample.

18. The method of claim 1, wherein the porous membrane is an ultrafiltration membrane.

19. The method of claim 1, wherein the porous membrane is a microporous membrane.

20. The method of claim 1, wherein the porous membrane comprises a polymeric material selected from the group consisting of PVDF (Polyvinylidene fluoride), polytetrofluoroethylene, hydrophilic polytetrofluoroethylene, polyethylene and polypropylene.

21. A sample holder card for use in the method of claim 1, wherein the sample holder card comprises a porous membrane which comprises a hydrophilic region surrounded by a hydrophobic region, wherein the sample is contained within the boundaries of the hydrophilic region.

22. The sample holder card of claim 1, wherein the diameter of the hydrophilic region ranges from 2.0 mm through 10 mm.

23. The sample holder card of claim 1, wherein the diameter of the hydrophilic region ranges from 3.0 mm through 6 mm.

24. A method for quantitation of one or more biomolecules in a sample, the method comprising the steps of:

(a) providing a sample holder card comprising a porous membrane which comprises a hydrophilic region surrounded by a hydrophobic region for sample containment;

(b) contacting the hydrophilic region of the membrane with a sample volume;

(c) drying the sample volume on the membrane;

(d) detecting the presence of water on the membrane using infrared absorbance and repeating step (c), if necessary, until no water is detected; and

(e) exposing the sample volume on the membrane to an infrared beam comprising a wavelength in the spectral range of 4000-400 cm<sup>−

1</sup> or any portion of the spectral range, thereby to obtain an infrared, absorption spectrum;

wherein one or more absorption peak areas in the infrared absorption spectrum correlates with the quantity of the one or more biomolecules in the sample.