# US 11,441,979 B2 · Thawing methods and apparatus

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- Number: US 11,441,979 B2
- Document ID: US11441979B2
- Status: Granted
- Kind: B2
- Application number: 15/532,815
- Filed: 2015-12-04
- Published: 2019-01-03
- Granted: 2022-09-13
- Earliest claimed: 2014-12-04
- Assignees: Asymptote Ltd.
- Applicants: Asymptote Ltd.
- Inventors: Stuart Milne; Stephen Lamb; Christopher Creasey; Alex Nancekievill; Rupert Rutledge
- Web page: https://patentgrep.com/patent/US11441979B2

## CPC

- G01N 1/44: Sample treatment involving radiation, e.g. heat
- A01N 1/142: Apparatus
- A01N 1/162: Temperature processes, e.g. following predefined temperature changes over time
- B01L 3/5082: {Test tubes per se}
- B01L 7/54: {using spatial temperature gradients}
- B01L 7/02: Water baths; Sand baths; Air baths
- B01L 2300/1827: using resistive heater
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)
- A01N: PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF (preservation of food or foodstuff A23); BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES (preparations for medical, dental or toiletry purposes which kill or prevent the growth or proliferation of unwanted organisms A61K); PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- B01L: CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE

## Abstract

The invention provides a method of thawing a sample comprised in a container, the method comprising the steps of: a) calculating an agitation program as a function of either or both of the sample volume and the type of the container, and the ice fraction in the sample, and optionally the thermal conductivity of the sample container; b) agitating said sample according to the program to agitate at least one region of the sample; and c) changing the agitation program applied to at least one region of the sample in response to changes in the sample volume and/or sample ice fraction. The invention further provides a method of reducing shearing to cells during a method of agitation, and methods for thawing a sample wherein a sample container is differentially heated. An apparatus for use in the methods is also provided, as is an apparatus for thawing and/or cooling a sample which comprises a resilient vessel wall.

## Claims

### Claim 1

A method of thawing a sample wherein said sample is within a container comprising an outer wall and an inner wall, said method comprising
a) heating at least one region of the outer wall of said container to a temperature equal to or above that at which cellular damage occurs and wherein the inner wall of the container is at a temperature lower than that at which cellular damage occurs, or
b) heating the container such that an outer wall region of said container corresponding to an inner wall region which contacts the sample, is heated to a temperature higher than another outer wall region corresponding to another inner wall region which does not contact the sample; and
c) i) calculating an agitation program as a function of 1) either or both of a sample volume and a type of container, 2) a sample ice fraction in the sample, and 3) the thermal conductivity of the sample container;
ii) agitating said sample according to the agitation program to agitate at least one region of the sample;
iii) changing the agitation program applied to the at least one region of the sample in response to changes in the sample volume and the sample ice fraction; and
iv) further agitating said sample according to the changed agitation program to agitate the at least one region of the sample.

### Claim 2 (depends on 1)

The method according to claim 1, wherein in step b) the outer wall region of the container corresponding to the inner wall region which contacts the sample is heated to a temperature at which cellular damage would occur, and wherein the outer wall region of the container corresponding to the inner wall region which does not contact the sample is at a temperature less than that at which cellular damage would occur.

### Claim 3 (depends on 1)

The method according to claim 1, wherein step b) further comprises a step of monitoring the temperature of the inner wall region contacting the sample and the inner wall region not contacting the sample and adjusting the heat or energy applied to one or both of the corresponding outer wall regions of the sample container in response to changes in the temperature of the two regions and/or a step of monitoring the sample temperature and/or the ice fraction in the sample and adjusting the heat or energy applied to the outer wall region, the corresponding inner wall region to which contacts the sample material.

### Claim 4 (depends on 1)

The method according to claim 1, wherein in b) the inner wall of the container is at a temperature less than that at which cellular damage would occur.

### Claim 5 (depends on 1)

The method according to claim 1, wherein in step a) or b) said temperature at which cellular damage would occur is a temperature above 37° C. and/or said temperature less than that at which cellular damage would occur is a temperature at or less than 37° C.

### Claim 6 (depends on 1)

The method as claimed in claim 1, wherein in step a) or b) said sample is agitated.

### Claim 7 (depends on 1)

The method according to claim 1, wherein in step c) steps ii) and iii) are repeated at least once, or wherein steps ii) and iii) are carried out continuously.

### Claim 8 (depends on 1)

The method according to claim 1, wherein in step c) said method comprises a step of monitoring sample volume and/or sample ice fraction carried out within or sequentially to step ii) and/or a step of determining the container type and/or the volume of the sample material and optionally comprises determining the thermal conductivity of the sample container, before step i).

### Claim 9 (depends on 1)

The method according to claim 1, wherein in step c) (1) agitation of the sample comprises agitation of a single region of the sample, of multiple regions of the sample, or the whole sample, (2) the change in agitation applied to the at least one region of the sample in step iii) comprises changing the frequency, amplitude or both frequency and amplitude of the agitation, (3) the sample is heated and/or (4) a heating and agitation program as a function of the volume of the sample, and the ice fraction is calculated, wherein the sample is heated according to the heating program to heat at least one region of the sample and wherein the heating program is changed to at least one region of the sample in response to changes in the sample volume and/or ice fraction in the sample.

### Claim 10 (depends on 1)

The method of claim 1, wherein in step c) the method reduces shearing during the agitation of the sample comprised in the container.

### Claim 11 (depends on 1)

The method as claimed in claim 1, wherein said sample is a biological sample comprising a biopharmaceutical, a cellular material, a biological tissue, a biological organ or a part thereof, nucleic acids, proteins, polypeptides and/or amino acids.