Plasmon resonance measuring method and apparatus

• US 20050213101A1
• Filed: 03/25/2005
• Published: 09/29/2005
• Est. Priority Date: 03/25/2004
• Status: Active Grant

• Current Assignee: PerkinElmer LAS, Inc. (Perkinelmer Incorporated)
• Original Assignee: PerkinElmer LAS, Inc. (Perkinelmer Incorporated)
• Inventors: Schermer, Mack
• Granted Patent: US 7,271,885 B2
• Time in Patent Office: Days
• US Class Current: 356/445
• CPC Class Codes: B01L 2300/0654 Lenses; Optical fibres
• CPC Class Codes: B01L 2300/0829 Multi-well plates; Microtitration plates
• CPC Class Codes: B01L 2300/0851 Bottom walls
• CPC Class Codes: B01L 3/5085 for multiple samples, e.g. microtitration plates
• CPC Class Codes: G01N 21/253 for batch operation, i.e. multisample apparatus analytical automats G01N35/00
• CPC Class Codes: G01N 21/553 and using surface plasmons fluorescence excitation G01N21/648; enhanced Raman G01N21/658

Abstract

Detection apparatus for performing surface plasmon resonance (SPR) measurements on a sample includes a support having side walls and a bottom wall defining a plurality of adjacent wells. The bottom wall has integrated therein SPR detectors underlying the wells for detecting the refractive indexes of samples deposited in the wells. Each SPR detector includes a prism having an exterior entrance surface, an exterior exit facet and an interior sensing surface located at the bottom of the overlying well, and a thin metal layer coated on the sensing surface. The prism may be molded from a transparent polymer into the bottom of a standard-format microplate structure. That structure may be used in conjunction with an SPR measuring or reading instrument to perform label-less assay measurements on various samples in an efficient and reliable manner. A method of acquiring the assay measurements is also disclosed.

Claims

1. Apparatus for performing surface plasmon resonance (SPR) measurements on a sample, said apparatus comprising a support having side walls and a bottom wall defining a plurality of wells, said bottom wall having integrated therein SPR detectors underlying the wells for detecting the refractive indexes of samples deposited in the wells, each SPR detector including a prism having an exterior entrance surface, an exterior exit surface and an interior sensing surface located at the bottom of the overlying well, and a thin metal layer coated on the sensing surface.

2. The apparatus defined in claim 1 wherein the plurality of wells is arranged in a rectangular array of columns and rows surrounded by a skirt in a standard microplate format.

3. The apparatus defined in claim 1 or 2 wherein the prisms are molded into the bottom wall.

4. The apparatus defined in claim 3 wherein the prisms are of a transparent polymer.

5. The apparatus defined in claim 4 wherein the polymer is one of the group consisting of polycarbonate, polystyrene, polysulfone and polymethylmethacrylate.

6. The apparatus defined in claim 4 wherein the side walls and the bottom wall are molded as a unitary part.

7. The apparatus defined in claim 1 or 2 wherein the side walls and the bottom wall are formed separately and secured together by a bonding agent.

8. The apparatus defined in claim 7 wherein the side walls and the bottom wall are of different materials.

9. The apparatus defined in claim 8 wherein the bottom wall is of glass.

10. The apparatus defined in claim 2 wherein each prism is a segment of a longer prism element which extends under an entire row or column of wells in the microplate.

11. The apparatus defined in claim 1 or 2 wherein the bottom wall includes a separation between the prisms underlying adjacent wells.

12. The apparatus defined in claim 1 wherein the metal layer is of gold.

13. The apparatus defined in claim 1 wherein each prism cross-section is an inverted irregular trapezoid whose base corresponds to said sensing surface and whose opposite sides correspond to said entrance and exit surface, each prism also having a lowermost surface opposite the sensing surface which declines toward the exit surface.

14. The apparatus defined in claim 13 wherein said sides descend from the base at equal angles and the lowermost surface declines at an angle of about 10°

relative to the base.

15. The apparatus defined in claim 1 or 2 and including a first member of a specific binding pair resident in at least one of said wells in contact with the metal layer therein to facilitate an assay.

16. The apparatus defined in claim 1 or 2 and including cells or cell fragments resident in at least one of said wells and immobilized against the metal layer therein to facilitate an assay.

17. Apparatus for performing surface plasmon resonance (SPR) measurements on a sample, said apparatus comprising a support having side walls and a bottom wall defining at least one well, said bottom wall having integrated therein a prism underlying said at least one well, each prism in cross-section having the shape of an inverted irregular trapezoid and whose base constitutes the bottom of the overlying well, whose opposite sides descend from the base toward one another and each prism also having a lowermost surface opposite the base which declines toward one of said sides.

18. The apparatus defined in claim 17 wherein said sides descend from the base at equal angles and the lowermost surface declines at an angle of about 10°

relative to the base.

19. The apparatus defined in claim 17 and further including a thin metal layer coated on the base of each prism.

20. The apparatus defined in claim 19 wherein said support is of plastic and said metal layer is of gold.

21. Surface plasmon resonance (SPR) measuring apparatus comprising

a fixture;

a support removably received in the fixture, said support having side walls and a bottom wall defining a multiplicity of wells for receiving samples, said bottom wall also defining prisms underlying said wells, each prism having an exterior entrance surface and an exterior exit surface, said entrance and exit surface descending from an interior metallized sensing surface located at the bottom of the associated well;

a light source for directing interrogating light rays into a selected one of said prisms through the entrance surface thereof, said interrogating rays being reflected at the corresponding sensing surface and leaving the selected prism from the exit surface thereof as measuring rays;

an optical detector paired with the light source for collecting said measuring rays and producing a corresponding output signal;

a positioning mechanism for moving the fixture and the source/detector pair relatively, and

a controller for controlling the positioning mechanism so that said multiplicity of wells can be interrogated in a selected order.

22. The apparatus defined in claim 21 and further including an output device responsive to the output signal for indicating the refractive index or index change of the sample in each interrogated well.

23. The apparatus defined in claim 21 wherein the light source provides interrogating rays which converge to a point near the entrance surface so that the interrogating rays are diverging as they pass into the selected prism.

24. The apparatus defined in claim 21 wherein the light source produces polarized light and includes a wavelength filter so that the interrogating rays are monochromatic.

25. The apparatus defined in claim 21 wherein the multiplicity of wells is arranged in a rectangular array of columns and rows surrounded by a skirt in a standard microplate format.

26. The apparatus defined in claim 25 including one or more additional source/detector pairs mechanically coupled in parallel with the first-mentioned source/detector pair enabling simultaneous SPR measurements to be taken of samples in a plurality of wells in a selected column or row of said array.

27. A method of acquiring assay measurements comprising the steps of

providing a support having side walls and a bottom wall defining a well, said bottom wall also defining a prism under the well which has an exterior entrance surface, an exterior exit surface and an interior metallized sensing surface at the bottom of the well with a selected SPR characteristic;

depositing a sample in the well so that the sample contacts the sensing surface thereby changing said SPR characteristic, and

measuring the change in said SPR characteristic.

28. The method defined in claim 27 wherein a first member of a specific binding pair is immobilized on the metallized sensing surface.

29. The method defined in claim 27 wherein cells or cell fragments are immobilized on the metallized sensing surface.

30. The method defined in claim 27 wherein the measuring step is performed by taking a first measurement at or just prior to the depositing of the sample in the well and then taking a second measurement after the assay has been incubated to a desired state of completion.

31. The method defined in claim 27 wherein a plurality of measurements are taken from the time the sample is deposited to the completion of incubation thereof thereby providing an indication of assay kinetics.