METHOD FOR DETECTING MICRO-COLONIES GROWING ON A MEMBRANE OR AN AGAROSE MEDIUM OF A SAMPLE AND A STERILITY TESTING APPARATUS

• US 20170044588A1
• Filed: 03/30/2015
• Published: 02/16/2017
• Est. Priority Date: 04/22/2014
• Status: Active Application

• Current Assignee: Merck Patent GmbH (Merck & Co., Inc.)
• Original Assignee: Merck Patent GmbH (Merck & Co., Inc.)
• Inventors: Felden, Luc, BOUTHILLON, Marine, Olivier, Stephane, Woehl, Pierre, Guedon, Pierre, Allard, Florian
• Application Number:
  US15/305,703
• Publication Number:
  US 20170044588A1
• Time in Patent Office: Days
• CPC Class Codes: C12Q 1/04 Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor C12Q1/6897 takes precedence
• CPC Class Codes: C12Q 1/22 Testing for sterility conditions
• CPC Class Codes: G01N 2021/1765 Method using an image detector and processing of image signal
• CPC Class Codes: G01N 2021/4704 Angular selective
• CPC Class Codes: G01N 2021/4709 Backscatter
• CPC Class Codes: G01N 2021/4711 Multiangle measurement
• CPC Class Codes: G01N 21/47 Scattering, i.e. diffuse reflection G01N21/25, G01N21/41 take precedence G01N21/55 takes precedence
• CPC Class Codes: G01N 21/55 Specular reflectivity
• CPC Class Codes: G01N 2201/103 Scanning by mechanical motion of stage
• CPC Class Codes: G06T 2207/10012 Stereo images
• CPC Class Codes: G06T 2207/10056 Microscopic image
• CPC Class Codes: G06T 2207/30024 Cell structures in vitro; Tissue sections in vitro
• CPC Class Codes: G06T 7/0012 Biomedical image inspection
• CPC Class Codes: G06V 20/693 Acquisition

Abstract

The present invention is directed to a method and an apparatus for detecting micro-colonies growing on a membrane or an agarose medium of a sample in a closed device. According to the invention the sample is irradiated with a light incident at an angle (β) with respect to the normal to the membrane or the surface of the agarose medium from outside the device. An incident area of the light on the membrane or the surface of the agarose medium is imaged by means of a light receiving element using an imaging angle (α) different from angle (β) with respect to the normal to the membrane or the surface of the agarose medium from outside the device. The light reflected, scattered and/or diffused from the membrane or the surface of the agarose medium and/or the micro-colonies on the membrane and/or the micro-colonies on the agarose medium is detected.

Claims

1. Method for detecting micro-colonies growing on a membrane (1) or an agarose medium of a sample (5) in a closed device (3), the method comprising the steps:

irradiating the sample (5) with a light incident at an angle (β

) with respect to the normal to the membrane (1) or the surface of the agarose medium from outside the device (3);

imaging an incident area (7) of the light on the membrane (1) or the surface of the agarose medium by means of a light receiving element (9) using an imaging angle (α

) different from angle (β

) with respect to the normal to the membrane (1) or the surface of the agarose medium from outside the device (3);

detecting the light reflected, scattered and/or diffused from the membrane or the surface of the agarose medium and/or the micro-colonies on the membrane and/or the micro-colonies on the agarose medium.

2. Method according to claim 1, using an electric powered light source, with a pattern illuminating the sample, said pattern being a point, a line, a plurality of lines, a square pattern, an arc or circle pattern, a cross pattern, a multi-square/round pattern, a grid pattern or a two-dimensional area, a circle spot, a rectangular spot, or a square spot and wherein the light receiving element (9) contains an array of pixel sensors.

3. The method of claim 1, comprising the step of determining variations in the height of the incident area (7) of the membrane (1) or the surface of the agarose medium based on the position of the image of the incident area on the light receiving element (9) using a triangulation method.

4. Method according to claim 1, wherein the detection of micro-colonies is performed by determining variation in the light by using the number of pixels receiving a signal level above a threshold value around a light intensity peak for each line or column of an array of pixels of the pixel sensor.

5. Method according to claim 1, wherein the detection of micro-colonies is performed by determining variation in the peak intensity value of the light by using the highest signal level for each line or column of the array of pixels of the pixel sensor.

6. Method according to claim 1, wherein the detection of micro-colonies is performed by determining variation in the intensity value of the light by using the signal level of the array of pixel sensor.

7. Method according to claim 1 further comprising:

scanning the membrane (1) or the surface of the agarose medium by a linear and/or rotational movement of the sample relative to the light receiving element (9).

8. Method according to claim 7 wherein:

the relative movement of the sample (5) relative to the light receiving element (9) is formed by a succession of a linear movement, displacing the incident area (7) from the center of the sample (5) towards the border thereof, followed by a rotational movement of the sample (5) around a center axis thereof, and displacing the incident area (7) from the border of the sample (5) towards the center thereof, or

displacing the incident area (7) from the center of the sample (5) towards the border thereof, displacing the incident area (7) from the border of the sample (5) towards the center thereof, followed by a rotational movement of the sample (5) around a center axis thereof.

9. Method according to claim 1, comprising:

performing a relative motion of the sample in respect to the light source (13) and the light receiving element (9) so as to illuminate and to image the entire membrane (1) or the entire surface of the agarose medium while.

10. Method according to claim 7, wherein an angle (δ

) between the direction of the linear movement displacing the incident area (7) from the center to the border and an imaging direction of the light receiving element (9) is set to exceed an angle (γ

) between the direction of the linear movement displacing the incident area (7) from the center to the border and the irradiation direction of the incident light.

11. Method according to claim 10, wherein the angle (γ

) between the direction of the linear movement displacing the incident area (7) from the center to the border and the irradiation direction of the incident light is set to exceed the angle (δ

) between the direction of the linear movement displacing the incident area (7) from the center to the border and an imaging direction of the light receiving element (9).

12. An apparatus for detecting micro-colonies growing on a membrane (1) or an agarose medium of a sample (5) in a closed device (3), the apparatus comprising:

a light source for irradiating the sample (5) with a light incident at an angle (β

) with respect to the normal to the membrane (1) or the surface of the agarose medium from outside the device (3);

a light receiving means (9) for imaging an incident area (7) of the light on the membrane (1) or the surface of the agarose medium by means of the light receiving element (9) using an imaging angle (α

) different from angle (β

) with respect to the normal to the membrane (1) or the surface of the agarose medium from outside the device (3); and

means for detecting the light reflected, scattered and/or diffused from the membrane or the surface of the agarose medium and/or the micro-colonies on the membrane and/or the micro-colonies on the agarose medium.

13. The apparatus according to claim 12, wherein the light source (13) is an electric powered light source configured to irradiate a pattern illuminating the sample, said pattern being a point, a line, a plurality of lines, a square pattern, an arc or circle pattern, a cross pattern, a multi-square/round pattern, a grid pattern or a two-dimensional area, a circle spot, a rectangular spot, or a square spot and wherein the light receiving element (9) contains an array of pixel sensors.

14. The apparatus according to claim 12, further comprising:

means for scanning the membrane (1) or the surface of the agarose medium by a linear and/or rotational movement of the sample relative to the light receiving element (9).