Accurately calculating acoustic time-of-flight

Patent metadata

Field	Value
Publication number	US-11009486-B2
Application number	US-201715624694-A
Country	US
Kind code	B2
Filing date	Jun 15, 2017
Priority date	Dec 17, 2014
Publication date	May 18, 2021
Grant date	May 18, 2021

Abstract

The subject disclosure presents systems and computer-implemented methods for determining an acoustic time-of-flight (TOF) of sound waves through a sample material with greater accuracy and in a more repeatable fashion, by invoking one or more of an envelope generation for an error function, fitting a non-linear curve to an ultrasound frequency sweep, or performing a clustered piece-wise linear regression on individual linear parts of the ultrasonic frequency sweep. The systems and methods are useful for, among other things, monitoring diffusion of fluids through porous materials, such as tissue samples.

First claim

We claim:

1. A method of obtaining a tissue sample that is sufficiently diffused with a fixative, said method comprising: (a) immersing an unfixed tissue sample into a volume of the fixative, wherein the fixative comprises a temperature raging from between about 0° C. to about 15° C.; (b) tracking diffusion of the fixative through the unfixed tissue sample by: (b1) transmitting an acoustical signal from a transmitter through the unfixed tissue sample when the tissue sample is immersed in the fixative; (b2) detecting the transmitted acoustical signal after the transmitted acoustical signal has passed through the unfixed tissue sample by a receiver; (b3) calculating time of flight (ToF) of the acoustical signal on a computing system comprising a processor; and (b4) repeating (b1)-(b3) until the calculated ToF reaches a predefined threshold value, wherein the predefined threshold value indicates that the unfixed tissue sample is sufficiently diffused with the fixative, wherein the unfixed tissue sample and the fixative are maintained at a temperature ranging from between about 0° C. to about 15° C. while performing the steps (b1)-(b4).

2. The method of claim 1 , wherein the fixative solution is a cross-linking fixative solution.

3. The method of claim 2 , wherein the cross-linking fixative is at a temperature from about 0° C. to about 15° C.

4. The method of claim 1 , further comprising upon the predefined threshold value is reached, allowing the temperature of the tissue sample and the fixative to rise to the ambient temperature or heating the temperature of the tissue sample and the fixative to a temperature of more than about 20° C.

5. The method of claim 1 , wherein step (b1) is performed by transmitting a set of acoustic signals respectively having a plurality (N) of different frequencies from an ultrasonic transmitter to an ultrasonic receiver, wherein the ultrasonic transmitter and ultrasonic receiver are positioned so that the frequency passes through the unfixed tissue sample before encountering the receiver.

6. The method of claim 1 , wherein the TOF calculation in step (b3) having a temporal resolution of a nanosecond.

7. The method of claim 1 , wherein the transmitter and the receiver are transducers of an acoustic monitoring system, the acoustic monitoring system being operatively coupled to a computer system comprising a processor and computer-interpretable instructions on a computer-readable, non-transitory medium, the instructions causing the processor to process the measured phase shifts provided by the acoustic monitoring system and information on the plurality (N) of frequencies applied on the biological sample for performing step (b3).

8. The method of claim 7 , wherein the transmitter comprises an ultrasonic transmitter; and wherein the receiver comprises an ultrasonic receiver.

9. The method of claim 1 , further comprising: after determining that the calculated TOF has reached a predefined threshold value, fixing the tissue sample by actively or passively raising the temperature of the tissue sample; contacting the fixed tissue sample with a specific binding entity capable of binding to the labile biomarker; and detecting binding of the specific binding entity.

10. The method of claim 1 , wherein the ToF is calculated using an envelope method of ToF calculation.

11. The method of claim 1 , wherein the ToF is calculated using a curve fitting method of ToF calculation.

12. The method of claim 1 , wherein the ToF is calculated using a linear regression method of ToF calculation.

13. The method of claim 1 , wherein the fixative comprises neutral buffered formalin.

14. The method of claim 1 , wherein the acoustic signal is an ultrasonic signal.

15. A method of fixing a tissue sample, said method comprising (a) obtaining a tissue sample sufficiently diffused with a cross-linking fixative solution by: (a1) immersing an unfixed tissue sample freshly obtained from a subject into a volume of the cross-linking fixative solution, said cross-linking fixative solution; (a2) tracking diffusion of the cross-linking fixative solution through the tissue sample by: (a2a) transmitting an acoustical signal from a transmitter through the tissue sample when the tissue sample is immersed in the cross-linking fixative solution; (a2b) detecting the transmitted acoustical signal after the transmitted acoustical signal has passed through the tissue sample; (a2c) calculating time of flight (ToF) of the acoustical signal on a computing system comprising a processor and (a2d) repeating (a2a)-(a2c) until the calculated ToF reaches a predefined threshold value, wherein the predefined threshold value indicates that the tissue sample is sufficiently diffused with the cross-linking fixative solution; (b) raising the temperature of the tissue sample after the tissue sample is sufficiently diffused with the cross-linking fixative solution to a temperature in the range of room temperature to 50° C., and holding the tissue sample in said range of temperatures for a period of time sufficient to allow fixation of the tissue sample and provide a fixed tissue sample.

16. The method of claim 15 , further comprising detecting a labile biomarker in the fixed tissue sample by contacting the fixed tissue sample with a specific binding entity capable of binding to the labile biomarker and detecting binding of the specific binding entity.

17. The method of claim 16 , wherein the labile biomarker is selected from the group consisting of a phosphorylated protein, an mRNA, and a miRNA.

18. The method of claim 15 , wherein the ToF is calculated using an envelope method of ToF calculation.

19. The method of claim 15 , wherein the ToF is calculated using a curve fitting method of ToF calculation.

20. The method of claim 15 , wherein the ToF is calculated using a linear regression method of ToF calculation.

21. The method of claim 15 , wherein the fixative comprises neutral buffered formalin.

22. A system for monitoring diffusion of a liquid into a tissue sample comprising: (a) a container for immersing the tissue sample in a volume of the liquid; (b) a transmitter arranged in or around the container in a position that permits transmission of the acoustical signal through the tissue sample when the tissue sample is immersed in the volume of liquid in the container; (c) a receiver arranged in or around the container at a fixed distance from the transmitter and positioned to receive the acoustical signal after the acoustical signal has been transmitted through the tissue sample; (d) a computing device communicatively coupled to the transmitter and receiver, the computing device configured to: (d1) calculate a time of flight (ToF) of the acoustical signal between the transmitter and receiver, by (d1a) transmitting a frequency sweep of the acoustical signal between a transmitter and a receiver, wherein the transmitter and receiver are positioned so that the frequency passes through the material before encountering the receiver; (d1b) detecting the frequency sweep of the acoustic signal with the receiver; (d1c) calculating time of flight of the acoustical signal by obtaining an error function of a frequency sweep of the acoustic signal; and, (d1d) generating an envelope of the error function, wherein the time-of-flight is based on a minimum of the error function; and, (d2) generate a signal when the ToF reaches a predetermined threshold.

23. The system of claim 22 , wherein the liquid is a fixative.

24. The

Assignees

• Ventana Med Syst Inc

Inventors

• Bauer Daniel
• Otter Michael
• Stevens Benjamin

Classifications

• G01N2291/02475

  Tissue characterisation · CPC title

• G01N29/024 Primary

  by measuring propagation velocity or propagation time of acoustic waves · CPC title

• G01N1/30

  Staining; Impregnating {; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis} · CPC title

• G01N29/4418

  with a model, e.g. best-fit, regression analysis · CPC title

• G01N29/07

  by measuring propagation velocity or propagation time of acoustic waves · CPC title