Method and device for detecting bacteria and determining the concentration thereof in a liquid sample

• US 9,644,230 B2
• Filed: 10/28/2014
• Issued: 05/09/2017
• Est. Priority Date: 10/29/2013
• Status: Active Grant

• Current Assignee: IDEXX Laboratories Incorporated
• Original Assignee: IDEXX Laboratories Incorporated
• Inventors: Hammond, Jeremy, Russell, James
• Primary Examiner(s): Shen, Bin
• Application Number:
  US14/525,628
• Publication Number:
  US 20150118708A1
• Time in Patent Office: 924 Days
• CPC Class Codes: B01L 2200/0652 Sorting or classification of particles or molecules
• CPC Class Codes: B01L 2300/0681 Filter
• CPC Class Codes: B01L 2300/0851 Bottom walls
• CPC Class Codes: B01L 2300/0864 comprising only one inlet and multiple receiving wells, e.g. for separation, splitting
• CPC Class Codes: B01L 2300/0877 Flow chambers
• CPC Class Codes: B01L 2400/086 using baffles or other fixed flow obstructions
• CPC Class Codes: B01L 3/502 with fluid transport, e.g. in multi-compartment structures centrifugal-type cuvettes G01N21/07; analysis by separation into components G01N30/00; automatic analysers G01N35/00
• CPC Class Codes: B01L 3/502761 specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads, for physically stretching molecules investigating characteristics of particles G01N15/00
• CPC Class Codes: C12Q 1/06 Quantitative determination
• CPC Class Codes: G01N 15/01 specially adapted for biological cells, e.g. blood cells investigating sedimentation of particle suspensions in blood G01N15/05
• CPC Class Codes: G01N 15/14 Optical investigation techniques, e.g. flow cytometry
• CPC Class Codes: G01N 15/149 specially adapted for sorting particles, e.g. by their size or optical properties
• CPC Class Codes: G01N 2015/1006 for cytology
• CPC Class Codes: G01N 2015/1447 Spatial selection
• CPC Class Codes: G01N 2015/1452 Adjustment of focus; Alignment
• CPC Class Codes: G01N 2015/1486 Counting the particles
• CPC Class Codes: G01N 2333/195 from bacteria
• CPC Class Codes: G01N 33/493 urine

Abstract

A method for detecting bacteria and determining the concentration thereof in a liquid sample includes the steps of taking an optical section through a container holding a volume of the liquid sample at a predetermined field of view and at a predetermined focal plane depth or angle and after a period of time has elapsed to allow non-bacteria in the sample to settle to the bottom of the container. Since bacteria auto arranges in the liquid sample, forming a lattice-like grid pattern, an optical section through the volume of auto-arranged bacteria may be used to measure the quantity of bacteria residing in that section. A container for holding the liquid sample has particular structure which aids in separating the non-bacteria from the bacteria.

Claims

1. A method for detecting bacteria and determining the concentration thereof in a liquid sample, which comprises the steps of:

taking at least one optical section through a volume of the liquid sample with a fluid imaging system having a camera at a predetermined field of view and at a predetermined focal plane depth or angle in the volume and after a predetermined time has elapsed to allow bacteria in the liquid sample to auto arrange in a uniform distribution;

counting the number of bacteria present within the at least one optical section with the fluid imaging system;

calculating the number of optical sections into which the volume of the liquid sample may be divided with the fluid imaging system thereby determining a total number of possible optical sections;

multiplying the number of bacteria present in the at least one optical section by the total number of possible optical sections with the fluid imaging system thereby determining at least an approximation of the total number of bacteria within the volume of the liquid sample; and

determining the concentration of bacteria within the liquid sample based on the at least approximation of the total number of bacteria within the volume of the liquid sample by the fluid imaging system.

2. A method for detecting bacteria and determining the concentration thereof in a liquid sample as defined by claim 1, wherein the predetermined time allowed for bacteria in the liquid sample to auto arrange is between about three minutes and about ninety minutes.

3. A method for detecting bacteria and determining the concentration thereof in a liquid sample as defined by claim 1, wherein the predetermined time allowed for bacteria in the liquid sample to auto arrange is between about three minutes and about 10 minutes.

4. A method for detecting bacteria and determining the concentration thereof in a liquid sample as defined by claim 1, wherein the at least one optical section taken through the volume of the liquid sample has a focal plane angle of about zero (0) degrees to about ninety (90) degrees relative to a vertical plane through the volume.

5. A method for detecting bacteria and determining the concentration thereof in a liquid sample as defined by claim 1, wherein the at least one optical section taken through the volume of the liquid sample has a focal plane angle of about zero degrees relative to a vertical plane through the volume.

6. A method for detecting bacteria and determining the concentration thereof in a liquid sample as defined by claim 1, wherein the at least one optical section taken through the volume of the liquid sample has a focal plane angle of about ninety degrees relative to a vertical plane through the volume.

7. A method for detecting bacteria and determining the concentration thereof in a liquid sample as defined by claim 6, wherein the optical section has a focal plane depth of at least one of about 100 microns, about 200 microns, about 400 microns, about 600 microns, about 800 microns, about 1,000 microns and about 1,200 microns above the bottom of the volume of the liquid sample.

8. A method for detecting bacteria and determining the concentration thereof in a liquid sample, which comprises the steps of:

taking a plurality of optical sections through a volume of the liquid sample with a fluid imaging system having a camera at a predetermined field of view and at one or more predetermined focal plane depths or angles in the volume and after a predetermined time has elapsed to allow bacteria in the liquid sample to auto arrange in a uniform distribution;

counting the number of bacteria present within each optical section of the plurality of optical sections with the fluid imaging system;

calculating an average of the number of bacteria present by dividing the total number of bacteria present in the plurality of optical sections by the number of optical sections taken through the volume of the liquid sample with the fluid imaging system thereby determining an average number of bacteria present within the optical sections of the plurality of optical sections;

calculating the number of optical sections into which the volume of liquid sample may be divided with the fluid imaging system thereby determining a total number of possible optical sections;

multiplying the average number of bacteria present in the optical sections of the plurality of optical sections by the total number of possible optical sections with the fluid imaging system thereby determining at least an approximation of the total number of bacteria within the volume of the liquid sample; and

determining the concentration of bacteria within the liquid sample based on the at least approximation of the total number of bacteria within the volume of the liquid sample by the fluid imaging system.

9. A method for detecting bacteria and determining the concentration thereof in a liquid sample, which comprises the steps of:

taking at least one optical section through a volume of the liquid sample with a fluid imaging system having a camera at a predetermined field of view and at a predetermined focal plane depth or angle in the volume and after a predetermined time has elapsed to allow bacteria in the liquid sample to auto arrange in a uniform distribution;

determining the average spacing between bacteria present within the at least one optical section with the fluid imaging system thereby determining the average bacteria spacing;

calculating the three dimensional area occupied by the volume of the liquid sample with the fluid imaging system thereby determining a three dimensional volumetric area;

dividing the three dimensional volumetric area by the average spacing between bacteria with the fluid imaging system thereby determining at least an approximation of the total number of bacteria within the volume of the liquid sample; and

determining the concentration of bacteria within the liquid sample based on the at least approximation of the total number of bacteria within the volume of the liquid sample by the fluid imaging system.