Determining the distribution of a substance by scanning with a measuring front

Patent metadata

Field	Value
Publication number	US-9024279-B2
Application number	US-201213657071-A
Country	US
Kind code	B2
Filing date	Oct 22, 2012
Priority date	Apr 22, 2010
Publication date	May 5, 2015
Grant date	May 5, 2015

Abstract

For determining the distribution of a substance, a measuring front is formed of a first and a second optical signal. Intensities of the first and second optical signals, over a depth of the measuring front which is smaller than the diffraction limit at the wavelengths of the first and second optical signals, increase so steeply that a portion of the substance in a measurement state in which a measurement signal is available from the substance increases from essentially zero due to transferring the substance by means of the first optical signal into the measurement state, and decreases to essentially zero again due to transferring the substance by means of the second optical signal back out of the measurement state. The measuring front is moved over a measurement region. The measurement signal is recorded for different positions of the measuring front in the measurement region and assigned to these positions.

First claim

I claim:

1. A method of determining the distribution of a substance in a measurement region, the substance being transferable
by means of a first optical signal (a) out of a first state in which no measurement signal is available from the substance into a measurement state in which a measurement signal is available from the substance, and
by means of a second optical signal (b) into one of the first state and a second state in which no measurement signal is available from the substance, the method comprising:
forming a measuring front of the first and second optical signals in the measurement region, wherein intensities of the first and second optical signals, over a depth of the measuring front in the direction of the combined intensity profiles of the first and second optical signals, the depth being smaller than the diffraction limit at the wavelengths of the first and second optical signals, increase so steeply that a portion of the substance in the measurement state over the depth of the measuring front increases from essentially zero due to transferring the substance by means of the first optical signal (a) out of the first state into the measurement state, and decreases to essentially zero again due to transferring the substance by means of the second optical signal (b) into the one of the first and second states;
moving the measuring front over the measurement region in a direction opposite to the increase of the intensities of the first and second optical signals over the depth of the measuring front;
at least recording the measurement signal emitted out of the measuring front for different positions of the measuring front in the measurement region; and
assigning the recorded measurement signal to the corresponding position of the measuring front in the measurement region,
wherein the first and second optical signals have different wavelengths, wherein the intensity distributions of both the first and second optical signals over the measuring front are structured by the same optical elements, and wherein the first and second optical signals whose intensities increase over the measuring front essentially have no intensity in front of the measuring front being sufficient for driving any changes of state of the substance.

2. The method of claim 1 , wherein a distribution of the measurement signal is recorded with spatial resolution along the measuring front and assigned to the corresponding position of the measuring front in the measurement region.

3. The method of claim 1 , wherein the measuring front is straight, and wherein the intensities of the first and second optical signals increasing over the depth of the measuring front are constant along the measuring front.

4. The method of claim 3 , wherein the measuring front is moved in two linearly independent directions over the measurement region.

5. The method of claim 1 , wherein the measuring front spans over at least hundred times an diffraction limit at the wavelengths of the first and second optical signals.

6. The method of claim 1 , wherein the measuring front extends over a full width of the measurement region.

7. The method of claim 1 , wherein the measurement signal is recorded with temporal resolution for each position of the measuring front in the measurement region.

8. The method of claim 2 , wherein the distribution of the measurement signal is recorded with a line detector aligned along the measuring front.

9. The method of claim 1 , wherein a distribution of the measurement signal is recorded with a detector array covering the entire measurement region.

10. The method of claim 1 , wherein the intensities of the first and second optical signals increase so steeply over the measuring front that the depth of the measuring front over which the portion of the substance in the measurement state increases from essentially zero, and decreases to essentially zero again is smaller than half of the diffraction limit at the wavelengths of the first and second optical signals.

11. The method of claim 1 , wherein the substance emits fluorescence light in the measurement state and no fluorescence light in the one of the first and second states.

12. The method of claim 1 , wherein the substance is excitable for emission of fluorescence light by means of excitation light in the measurement state and not excitable for the emission of the fluorescence light by means of the excitation light in the one of the first and second states, the fluorescence light being the measurement signal.

13. The method of claim 1 , wherein a resetting optical signal is applied to the substance in the measurement region over which the measuring front has been moved prior to moving the measuring front again over the measurement region.

14. The method of claim 13 , wherein the resetting optical signal is selected from the blue to ultraviolet wavelength range.

15. The method of claim 1 , wherein the measurement region is a part of a data carrier and wherein the distribution of the substance in the measurement region is transcribed into data stored on the data carrier.

16. The method of claim 15 , wherein the substance is arranged in tracks arranged at a distance of at least the diffraction limit at the wavelength of the measurement signal and wherein the measuring front is aligned perpendicular to the tracks.

17. A scanning light microscope for determining the distribution of a substance in a measurement region, comprising
a light source configured to provide first and second optical signals of different wavelength, wherein the first optical signal transfers the substance out of a first state in which no measurement signal is available from the substance into a measurement state in which a measurement signal is available from the substance, and the second optical signal transfers the substance into one of the first state and a second state in which no measurement signal is available from the substance
optics configured to form a measuring front of the first and second optical signals in the measurement region, wherein intensities of the first and second optical signals, over a depth of the measuring front in the direction of the combined intensity profiles of the first and second optical signals, the depth being smaller than the diffraction limit at the wavelengths of the first and second optical signals, increase so steeply that a portion of the substance in the measurement state over the depth of the measuring front, increases from essentially zero due to transferring the substance by means of the first optical signal (a) out of the first state into the measurement state, and decreases to essentially zero again due to transferring the substance by means of the second optical signal (b) into the one of the first and second states
a scanner configured to move the measuring front with regard to the measurement region, and
a detector configured to at least record a distribution of the measurement signal emitted out of the measuring front with spatial resolution along the measuring front,
wherein the optics comprise same optical elements for structuring intensity distributions of the first and second optical signals over the measuring front, wherein the first and second optical signals whose intensities increase over the measuring front essentially have no intensity in front of the measuring front being sufficient for driving any changes of state of the substance.

18. The scanning light microscope of claim 17 , wherein the detector includes a line detector.

19. A method of locally initiating a conversion of a sub

Assignees

• Max Planck Gesellschaft

Inventors

• Hell Stefan W

Classifications

• G01N21/6428

  Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" (in vivo A61B5/00; immunoassay G01N33/53) · CPC title

• G01N21/6458 Primary

  Fluorescence microscopy (fluorescence microscopes per se G02B21/0076 and G02B21/16) · CPC title