Methods of determining exposure to UV light

Patent metadata

Field	Value
Publication number	US-10955291-B2
Application number	US-201314385249-A
Country	US
Kind code	B2
Filing date	Mar 7, 2013
Priority date	Mar 15, 2012
Publication date	Mar 23, 2021
Grant date	Mar 23, 2021

Abstract

Methods of determining the dose of UVC light delivered to a sample comprising a low optical transmission complex fluid are provided. Also provided are methods of inactivation of an organism, such as a spore, a bacteria or a virus, in a sample comprising dose of UVC light delivered to a sample comprising a low optical transmission complex fluid.

First claim

What is claimed is:

1. A method of determining the dose of UVC light delivered to a sample comprising a low optical transmission complex fluid comprising:
(a) measuring the fluence rate delivered by a NIST traceable UVC source;
(b) generating a standard curve by:
(i) contacting a bleachable fluorescence emitter with a low optical transmission complex fluid comprising cell culture media to form a control mixture, wherein the bleachable fluorescence emitter is not contacted with a high optical transmission solution or water that does not contain carbohydrates and/or amino acids as solutes;
(ii) exposing the control mixture to UVC light for an initial residence time;
(iii) obtaining an aliquot from the control mixture;
(iv) measuring the fluorescence emitted by the aliquot of (iii);
(v) repeating (ii)-(iv) one or more times, wherein the control mixture is exposed to UVC light for a residence time that is longer than the initial residence time;
(vi) correlating the fluorescence emitted with the residence time;

(c) contacting a bleachable fluorescence emitter with a test fluid comprising a low optical transmission complex fluid comprising cell culture media to form a test mixture;
(d) exposing the test mixture to UVC light for a selected residence time;
(e) measuring the fluorescence emitted by the test mixture; and
(f) determining the dose delivered to the test mixture using the standard curve of (b).

2. The method of claim 1 , wherein the UVC light has a wavelength in the range of about 200 nm to about 280 nm.

3. The method of claim 2 , wherein the UVC light has a wavelength of 254 nm.

4. The method of claim 1 , wherein the bleachable fluorescence emitter comprises a UV-sensitive fluorescent microsphere.

5. The method of claim 1 , wherein the dose is provided as one of a dose distribution, a mean dose, a P10 dose, a P50 dose and a P90 dose.

6. The method of claim 1 , wherein the fluence rate delivered by the UVC source is measured using a NIST traceable UVC detector.

7. The method of claim 1 , wherein correlating the fluorescence emitted with the residence time in step (vi) comprises measuring the fluorescence distributions of aliquots from the control mixture taken at different residence times and determining photo-bleaching kinetic parameters in a photo-bleaching equation based on the fluorescence distributions.

8. The method of claim 7 , wherein the UVC light dose delivered to the test mixture is determined from the fluorescence distribution of the test mixture using the photo-bleaching equation and determined photo-bleaching kinetic parameters.

9. The method of claim 1 , wherein the fluorescence emitted from the aliquots of the control mixture and the test mixture is measured with a digital flow cytometer.

10. A method of determining the dose of UVC light delivered to a sample comprising a low optical transmission complex fluid comprising:
(a) measuring the fluence rate delivered by a NIST traceable UVC source;
(b) generating a standard curve by:
(i) contacting a bleachable fluorescence emitter with a low optical transmission complex fluid to form a control mixture comprising cell culture media, wherein the bleachable fluorescence emitter is not contacted with a high optical transmission solution or water that does not contain carbohydrates and/or amino acids as solutes;
(ii) exposing the control mixture to UVC light from the UVC source for an initial residence time;
(iii) obtaining an aliquot from the control mixture;
(iv) measuring the fluorescence emitted by the aliquot of (iii);
(v) repeating (ii)-(iv) one or more times, wherein the control mixture is exposed to UVC light for a residence time that is longer than the initial residence time;
(vi) calculating the UVC dose for each aliquot based on the measured fluence rate from the UVC source and residence time;
(vii) correlating the fluorescence emitted with the UVC dose by measuring the fluorescence distributions of aliquots from the control mixture taken at different residence times and determining photo-bleaching kinetic parameters in a photo-bleaching equation based on the fluorescence distributions;

(c) contacting a bleachable fluorescence emitter with a test fluid comprising a low optical transmission complex fluid comprising cell culture media to form a test mixture;
(d) exposing the test mixture to UVC light for a selected residence time;
(e) measuring the fluorescence emitted by the test mixture; and
(f) determining the dose delivered to the test mixture using the standard curve of (b).

11. The method of claim 10 , wherein the UVC light dose delivered to the test mixture is determined from the fluorescence distribution of the test mixture using the photo-bleaching equation and the photo-bleaching kinetic parameters determined in step (vii).

12. The method of claim 11 , wherein the low optical transmission complex fluid comprises a protein A-purified monoclonal antibody pool.

13. A method of determining the dose of UVC light delivered to a sample comprising a low optical transmission complex fluid comprising:
(a) measuring the fluence rate delivered by a NIST traceable UVC source;
(b) generating a standard curve by:
(i) contacting a bleachable fluorescence emitter with a low optical transmission complex fluid to form a control mixture, wherein the wherein the bleachable fluorescence emitter is not contacted with a high optical transmission solution or water that does not contain carbohydrates and/or amino acids as solutes;
(ii) exposing the control mixture to UVC light from the UVC source for an initial residence time;
(iii) obtaining an aliquot from the control mixture;
(iv) measuring the fluorescence emitted by the aliquot of (iii);
(v) repeating (ii)-(iv) one or more times, wherein the control mixture is exposed to UVC light for a residence time that is longer than the initial residence time;
(vi) calculating the UVC dose for each aliquot based on the measured fluence rate from the UVC source and residence time;
(vii) correlating the fluorescence emitted with the UVC dose by measuring the fluorescence distributions of aliquots from the control mixture taken at different residence times and determining photo-bleaching kinetic parameters in a photo-bleaching equation based on the fluorescence distributions;

(c) contacting a bleachable fluorescence emitter with a test fluid comprising a low optical transmission complex fluid to form a test mixture;
(d) exposing the test mixture to UVC light for a selected residence time;
(e) measuring the fluorescence emitted by the test mixture; and
(f) determining the dose delivered to the test mixture using the standard curve of (b), wherein the UVC light dose delivered to the test mixture is determined from the fluorescence distribution of the test mixture using the photo-bleaching equation and the photo-bleaching kinetic parameters determined in step (vii).

14. The method of claim 13 , wherein the low optical transmission complex fluid is selected from a fluid comprising cell culture media, a fluid comprising serum, a fluid comprising a mixture comprising a vitamin, a sugar and a pigment, and a fluid comprising a solution containing amino acids, peptides or proteins.

Assignees

• Amgen Inc

Inventors

• Hart Roger
• Lewis Amanda
• Welborn Brent

Classifications

• A61L2/10

  Ultraviolet [UV] radiation · CPC title

• A61L2/081

  Gamma radiation · CPC title

• A61L2103/05

  Living organisms or biological materials · CPC title

• A23B2/53 Primary

  with ultraviolet light · CPC title

• G01N21/6486

  Measuring fluorescence of biological material, e.g. DNA, RNA, cells (G01N21/6428 takes precedence) · CPC title