Claims
- 1. A method for transiently permeabilizing a target cell, comprising the steps of
(a) illuminating a population of substantially stationary cells contained in a frame; (b) obtaining a static representation of at least one property of light directed simultaneously from the frame; (c) locating a target cell in the population of cells, wherein the target cell is located with reference to the static representation; and (d) irradiating the target cell with a pulse of radiation; whereby the target cell is transiently permeabilized.
- 2. A method for transiently permeabilizing a target cell, comprising the steps of
(a) illuminating a population of cells contained in a frame, wherein the cells are illuminated through a lens having a numerical aperture of at most 0.5; (b) detecting at least one property of light directed from the frame and through the lens; (c) locating a target cell in the population of cells, wherein the target cell is located with reference to the detected property of light; and (d) irradiating the target cell with a pulse of radiation; whereby the target cell is transiently permeabilized.
- 3. A method for transiently permeabilizing a target cell, comprising the steps of
(a) illuminating a population of cells contained in a frame; (b) detecting at least one property of light directed from the frame; (c) locating a target cell in the population of cells, wherein the target cell is located with reference to the detected property of light; and (d) irradiating the target cell with a pulse of radiation, wherein the pulse of radiation has a diameter of at least 5 microns at the point of contact with the target cell; whereby the target cell is transiently permeabilized.
- 4. A method for transiently permeabilizing a target cell, comprising the steps of
(a) illuminating a population of cells contained in a frame; (b) detecting at least one property of light directed from the frame; (c) locating a target cell in the population of cells, wherein the target cell is located with reference to the detected property of light; and (d) irradiating the target cell with a pulse of radiation, wherein the pulse of radiation delivers at most 1 μJ/μm2; whereby the target cell is transiently permeabilized.
- 5. The method of claim 2, 3 or 4, wherein the population of cells is substantially stationary.
- 6. The method of claim 2, 3 or 4, wherein the property of light detected in step (b) is obtained as a static representation of light transmitted simultaneously from the frame, whereby the target cell located in step (c) is located with reference to the static representation.
- 7. The method of claim 2, 3 or 4, wherein step (b) further comprises obtaining a static representation of the at least one property of light transmitted simultaneously from the frame, whereby step (c) further comprises locating the target cell with reference to the static representation.
- 8. The method of claim 1, 2, 3 or 4, wherein at least one property of light is fluorescence and the target cell is located with reference to the fluorescence.
- 9. The method of claim 1, 3 or 4, wherein the population of cells is illuminated through a lens having numerical aperture of at most 0.5 and the target cell is located with reference to a property of light directed from the frame and through the lens.
- 10. The method of claim 2, wherein the lens has a numerical aperture of at most 0.4.
- 11. The method of claim 2, wherein the lens has a numerical aperture of at most 0.3.
- 12. The method of claim 2, wherein the lens has flat field correction.
- 13. The method of claim 2, wherein the lens has a working distance of at least 5 mm.
- 14. The method of claim 2, wherein the lens has a working distance of at least 10 mm.
- 15. The method of claim 1, 2 or 4, wherein the pulse of radiation has a diameter of at least 5 microns at the point of contact with the target cell.
- 16. The method of claim 3, wherein the pulse of radiation has a diameter of at least 7 microns at the point of contact with the target cell.
- 17. The method of claim 3, wherein the pulse of radiation has a diameter of at least 10 microns at the point of contact with the target cell.
- 18. The method of claim 3, wherein the pulse of radiation has a diameter of at least 20 microns at the point of contact with the target cell.
- 19. The method of claim 1, 2 or 3, wherein the pulse of radiation delivers at most 2 μJ/μm2.
- 20. The method of claim 4, wherein the pulse of radiation delivers at most 0.1 μJ/μm2.
- 21. The method of claim 4, wherein the pulse of radiation delivers at most 0.01 μJ/μm2.
- 22. The method of claim 1, further comprising the step of (e) adjusting the direction of the pulse of radiation to irradiate a second target cell in the population, whereby the second target cell is transiently permeabilized.
- 23. The method of claim 1, wherein the frame has an area of at least 50 mm2.
- 24. The method of claim 1, wherein the frame has an area of at least 85 mm2.
- 25. The method of claim 1, wherein the frame has an area of at least 115 mm2.
- 26. The method of claim 1, wherein the population of cells is illuminated with a laser.
- 27. The method of claim 1, wherein the population of cells is illuminated with a lamp.
- 28. The method of claim 1, wherein the population of cells is illuminated with light selected from the group consisting of visible, ultraviolet, and infrared wavelengths.
- 29. The method of claim 1, wherein the property of light is selected from the group consisting of visible, ultraviolet, or infrared wavelengths.
- 30. The method of claim 1, wherein the property of light is transmittance and the target cell is located with reference to the transmittance.
- 31. The method of claim 1, wherein the property of light is polarization and the target cell is located with reference to the polarization.
- 32. The method of claim 1, wherein the property of light is reflectance and the target cell is located with reference to the reflectance.
- 33. The method of claim 1, wherein the property of light is phase contrast illumination and the target cell is located with reference to the phase contrast illumination.
- 34. The method of claim 1, wherein the property of light is intensity and the target cell is located with reference to the intensity.
- 35. The method of claim 1, wherein greater than 50% of the irradiated target cells are viable after the method is performed.
- 36. The method of claim 1, wherein greater than 80% of the irradiated target cells are viable after the method is performed.
- 37. The method of claim 1, wherein greater than 90% of the irradiated target cells are viable after the method is performed.
- 38. The method of claim 1, wherein the target cell is a procaryotic cell.
- 39. The method of claim 1, wherein the target cell is a eucaryotic cell.
- 40. The method of claim 1, wherein the target cell is selected from the group consisting of an animal cell, plant cell, yeast cell, human cell and non-human primate cell.
- 41. The method of claim 1, wherein the population of cells contains cells associated with an exogenous label.
- 42. The method of claim 41, wherein the label is a fluorophore.
- 43. The method of claim 1, wherein the target cell is associated with an exogenous label.
- 44. The method of claim 1, wherein the target cell is in the presence of an exogenous molecule, whereby the exogenous molecule enters the transiently permeabilized cell.
- 45. The method of claim 44, wherein the exogenous molecule is selected from the group consisting of a nucleic acid, polypeptide, carbohydrate, lipid, and small molecule.
- 46. The method of claim 45, wherein the small molecule is a dye capable of absorbing visible, ultraviolet or infrared light.
- 47. The method of claim 44, wherein the molecular weight of the exogenous molecule is greater than 0.1 kiloDalton.
- 48. The method of claim 44, wherein the molecular weight of the exogenous molecule is greater than 0.3 kiloDalton.
- 49. The method of claim 44, wherein the molecular weight of the exogenous molecule is greater than 1 kiloDalton.
- 50. The method of claim 44, wherein the molecular weight of the exogenous molecule is greater than 3 kiloDaltons.
- 51. The method of claim 44, wherein the molecular weight of the exogenous molecule is greater than 10 kiloDaltons.
- 52. The method of claim 44, wherein the molecular weight of the exogenous molecule is greater than 30 kiloDaltons.
- 53. The method of claim 44, wherein the molecular weight of the exogenous molecule is greater than 70 kiloDaltons.
- 54. The method of claim 1, wherein step (b) further comprises obtaining a second static representation of at least one property of light directed simultaneously from the frame.
- 55. The method of claim 54, wherein the target cell is located with reference to said first and second static representation.
- 56. The method of claim 1, wherein steps (c) and (d) are repeated so that more than one target cell is located and irradiated.
- 57. The method of claim 1, further comprising the steps of
(e) illuminating a population of cells contained in a second frame (f) obtaining a static representation of at least one property of light directed from the second frame and through the lens, and repeating steps (c) through (d).
- 58. The method of claim 57, wherein the population of cells remains in a substantially stationary location relative to the lens.
- 59. The method of claim 57, wherein at least 10,000 cells are irradiated per minute.
- 60. The method of claim 57, wherein at least 20,000 cells are irradiated per minute.
- 61. The method of claim 57, wherein at least 50,000 cells are irradiated per minute.
- 62. The method of claim 57, wherein at least 100,000 cells are irradiated per minute.
- 63. The method of claim 57, further comprising the step of
(g) moving the population of cells relative to the lens and repeating steps (a) through (f).
- 64. The method of claim 57, wherein steps (a) through (f) are automated.
- 65. The method of claim 1, further comprising the steps of
(e) moving the population of cells relative to the lens and repeating steps (a) through (d).
- 66. The method of claim 1, wherein steps (a) through (d) are automated.
- 67. The method of claim 1, wherein the static representation comprises an image.
- 68. The method of claim 1, wherein the static representation comprises a set of data stored in computer memory.
- 69. The method of claim 1, further comprising a camera having a magnification between 2× and 40×.
- 70. The method of claim 1, further comprising a camera having a magnification between 2.5× and 25×.
Parent Case Info
[0001] This application is a continuation in part of U.S. patent application Ser. No. 09/728,281, filed Nov. 30, 2000; which is a continuation in part of application Ser. No. 09/451,659, filed Nov. 30, 1999; which is a continuation in part of application No. 09/049,677, filed Mar. 27, 1998, now U.S. Pat. No. 6,143,535; which is a continuation in part of application No. 08/824,968, filed Mar. 27, 1997, now U.S. Pat. No. 5,874,266, each of which is incorporated by reference herein.
Continuation in Parts (4)
|
Number |
Date |
Country |
Parent |
09728281 |
Nov 2000 |
US |
Child |
09961691 |
Sep 2001 |
US |
Parent |
09451659 |
Nov 1999 |
US |
Child |
09728281 |
Nov 2000 |
US |
Parent |
09049677 |
Mar 1998 |
US |
Child |
09451659 |
Nov 1999 |
US |
Parent |
08824968 |
Mar 1997 |
US |
Child |
09049677 |
Mar 1998 |
US |