Devices and methods for infrared (IR) based quantitation of biomolecules
View Patent ↗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.
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 −1 or any portion of the spectral range of 4000-400 cm −1 , 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 μl.
6. The method of claim 5 , wherein sample volume is 1 μl 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 −1 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.