Fluorescent biological particle detection system

• US 5,895,922 A
• Filed: 05/23/1997
• Issued: 04/20/1999
• Est. Priority Date: 03/19/1996
• Status: Expired due to Term

• Current Assignee: TSI Incorporated
• Original Assignee: Her Majesty The Queen In Right of Canada As Represented By The Minister of National Defence
• Inventors: Ho, Jim Yew-Wah
• Primary Examiner(s): Hannaher, Constantine
• Application Number:
  US08/863,023
• Time in Patent Office: 697 Days
• Field of Search: 250/461.2
• US Class Current: 250/492.1
• CPC Class Codes: C12M 41/46 of cellular or enzymatic activity or functionality, e.g. cell viability
• CPC Class Codes: G01N 15/1459 the analysis being performed on a sample stream
• CPC Class Codes: G01N 2015/019 Biological contaminants; Fouling
• CPC Class Codes: G01N 21/64 Fluorescence; Phosphorescence

Abstract

A process and apparatus are provided for detection of viable and potentially hazardous biological particles which may be dispersed in a particulate-containing airstream. The process comprises directing each of the contained particles along a linear path through air, in a sequential manner, and sampling them for determination of their size, whether they are biological and viable, and whether they are present in concentrations greater than background levels. The particle size identifies the particles as respirable or not and the particles are characterized as biological and viable by subjecting each particle in turn, to 340 nm, ultraviolet laser light and looking for the emission of fluorescence which is typically emitted from bacteria or bacterial spore. Fluorescence detected in the 400-540 nm range signals the presence of nicotinamide adenine dinucleotide hydrogen, which is indicative of biological activity or viability. A more compact, and power-saving apparatus results with the preferential use of a solid state, ultraviolet laser, which is actuated only when the particle is passing the laser and only if it is deemed to be a biologically viable candidate.

Claims

1. Apparatus for identifying the existence of viable biological particles within an airstream containing a mixture of biologically viable and biologically inert particles, comprising:

means for directing the particles individually and sequentially along a substantially linear path through air;

means for calculating the position of each particle as a function of time;

a source for emitting an ultraviolet laser beam when the calculated position of the particle is such that it is passing the laser beam source, said beam being directed to contact each particle moving along its linear path in the air and having a wavelength operative to excite biomolecules contained therein to produce fluorescence;

means for measuring the intensity of the fluorescence emitted from each particle and producing a signal indicative thereof, said intensity being measured during the period of time when the position of the particle is calculated to be substantially coincident with the ultraviolet laser beam; and

means for comparing each particle'"'"'s fluorescence intensity signal against pre-determined criteria and establishing whether that particle is a biologically viable particle or an inert particle.

2. Apparatus as recited in claim 1 wherein the source of the ultraviolet laser beam

emits an ultraviolet laser beam in one or more discontinuous pulses,

each laser beam pulse being directed along a light path which intersects the path of each particle moving along its linear path in the air, and

said laser beam pulse being emitted during the period of time when the position of the particle is calculated to be substantially coincident with the light path of the ultraviolet laser beam.

3. Apparatus as recited in claim 2 wherein the means for directing the particles individually and sequentially along a substantially linear path through air comprises:

an air discharge nozzle capable of accelerating the flow of the particle-containing airstream through the nozzle so that the particles exit the nozzle individually and sequentially along a substantially linear path, the air flow acting to accelerate a particle passing through the nozzle due to the aerodynamic drag acting thereon, said particle resisting the acceleration by its inertia, whereby the velocity of the particle exiting the nozzle is less than that of the airstream.

4. Apparatus as recited in claim 3 wherein the means for calculating the position of the particle comprises:

means for measuring the velocity of the airstream and producing a signal indicative thereof;

means for emitting spaced apart laser beams, both of which intersect the particle'"'"'s linear path, the first beam intersecting the path adjacent the nozzle, the second of beam being spaced a known distance downstream;

means for measuring the time taken for the particle to move between the first and second laser beams and producing a signal indicative thereof; and

means for processing the known distance between the first and second laser beams and the time signal so as to establish the velocity of the particle and producing a signal indicative thereof; and

means for processing the difference between the particle velocity signal and the airstream velocity signal so as to establish the size of the particle and producing a signal indicative thereof;

means for calculating the position of the particle as a function of time knowing the particle size, the airstream velocity, and the particle'"'"'s velocity.

5. Apparatus as recited in claim 4 wherein the ultraviolet laser beam has a wavelength between 320 and 360 nm and the means for measuring the intensity of the fluorescence is operative to measure it in the wavelength range of 400-540 nm.

6. Apparatus as recited in claim 5 wherein the ultraviolet laser beam has a wavelength operative to excite nicotinamide adenine dinucleotide hydrogen and to produce fluorescence therefrom.

7. Apparatus as recited in claim 6 further comprising:

means for storing the fluorescence intensity signal and the corresponding particle size signal for each particle;

means for counting the number of particles and producing signals indicative thereof;

means for processing the stored fluorescence intensity, particle size and particle count signals and establishing the concentration of biologically viable particles as a function of particle size.

8. Apparatus as recited in claim 7 wherein the means for emitting two spaced apart laser beams comprises:

the source emitting the ultraviolet laser beam; and

means for splitting the ultraviolet laser beam into two beams.

9. Apparatus for identifying the existence of viable biological particles within an airstream containing a mixture of biologically viable and biologically inert particles, comprising:

an air discharge nozzle capable of accelerating the flow of the particle-containing airstream through the nozzle so that the particles exit the nozzle individually and sequentially along a substantial linear path, the air flow acting to accelerate the particular particle passing through the nozzle due to the aerodynamic drag acting thereon, said particle resisting the acceleration by its inertia, whereby the velocity of the particle exiting the nozzle is less than that of the airstream;

means for measuring the velocity of the airstream and producing a signal indicative thereof;

means for emitting spaced apart laser beams, both of which intersect the particle'"'"'s linear path, the first beam intersecting the path adjacent the nozzle, the second of beam being spaced a known distance downstream;

means for measuring the time taken for the particle to move between the first and second laser beams and producing a signal indicative thereof; and

means for processing the known distance between the first and second laser beams and the time signal so as to establish the velocity of the particle and producing a signal indicative thereof;

means for processing the difference between the particle velocity signal and the airstream velocity signal so as to establish the size of the particle;

means for processing the signals for the particle'"'"'s size and comparing it against pre-determined criteria and establishing whether the particle is a candidate as being a biologically viable particle;

means for calculating the position of the candidate particle as a function of time and producing a signal indicative thereof;

a source for repeatedly emitting an ultraviolet laser beam in discontinuous pulses, each laser beam pulse being directed along a light path which intersects the path of each particle moving along its linear path in the air, said beam having a wavelength operative to excite biomolecules contained therein to produce fluorescence;

means for selectively triggering a ultraviolet laser beam pulse when the calculated position of the candidate particle corresponds with the intersection of the particle'"'"'s linear path and the ultraviolet laser light path;

means for measuring the intensity of the fluorescence emitted from each candidate particle and producing a signal indicative thereof; and

means for comparing each candidate particle'"'"'s fluorescence intensity signal against pre-determined criteria and establishing whether that particle is a biologically viable or an inert particle.

10. Apparatus as recited in claim 9 wherein the ultraviolet laser beam has a wavelength between 320 and 360 nm and the means for detecting fluorescence is operative to measure it in the wavelength range of 400-540 nm.

11. Apparatus as recited in claim 10 wherein the ultraviolet laser beam has a wavelength operative to excite nicotinamide adenine dinucleotide hydrogen and to produce fluorescence therefrom.

12. Apparatus as recited in claim 11 further comprising:

means for storing the fluorescence intensity signal and the corresponding particle size signal for each candidate particle;

means for counting the number of particles and producing signals indicative thereof;

means for processing the stored fluorescence intensity, particle size and particle count signals and establishing the concentration of biologically viable particles as a function of particle size.

13. Apparatus as recited in claim 12 wherein the means for emitting two spaced apart laser beams comprises:

the source emitting the ultraviolet laser beam; and

means for splitting the ultraviolet laser beam into two beams.

14. A process for the detecting the existence of viable biological particles within a stream of air containing a mixture of biologically viable and biologically inert particles, comprising the steps of:

(a) continuously processing the stream of air so as to direct the particles along a substantially linear path through air in a sequential manner;

(b) calculating the position of each particle as a function of time;

(c) providing an ultraviolet laser source for producing a beam of ultraviolet light directed along a light path;

(d) actuating the laser source and emitting an ultraviolet laser beam when the calculated position of the particle is substantially coincident with the light path of the laser beam so that the ultraviolet laser beam contacts the particle, the laser beam having a wavelength suitable to excite biomolecules contained therein and produce fluorescence;

(e) detecting fluorescence from the particle and establishing its intensity;

(f) comparing the intensity of the particle'"'"'s fluorescence against pre-determined criteria so as to establish whether that particle is biologically viable or is an inert particle; and

(g) performing the steps (b)-(f) for the next particle in sequence.

15. A process for the detecting the existence of viable biological particles within a stream of air containing a mixture of biologically viable and biologically inert particles, comprising the steps of:

(a) continuously processing the stream of air so as to direct the particles along a substantially linear path through air in a sequential manner;

(b) measuring biological characteristics associated with each particle;

(c) comparing the particle'"'"'s biological characteristics against pre-determined criteria and establishing whether that particle is a candidate as being a biologically viable particle;

(d) selectively contacting the candidate particle with an ultraviolet laser beam having a wavelength suitable to excite biomolecules contained therein and produce fluorescence;

(e) detecting fluorescence from the candidate particle and establishing its intensity;

(f) comparing the intensity of the candidate particle'"'"'s fluorescence against pre-determined criteria so as to establish whether that particle is biologically viable or is an inert particle; and

(g) performing the steps (b)-(f) for the next particle in sequence.

16. The process as set forth in claim 15 further comprising:

measuring the size of the particle as a biological characteristic; and

establishing the particle as a candidate particle if its size is within the range of particles which are respirable.

17. The process as set forth in claim 16 wherein the candidate particle is selectively contacted with an ultraviolet laser beam by

pulsing the beam when a candidate particle is substantially coincident with the path of the ultraviolet laser beam.

18. The process as recited in claim 17 wherein the stream of air is continuously processed by:

rapidly accelerating the stream of air containing particles to a known velocity wherein each particle accelerates to reach a characteristic velocity, slower than the velocity of the stream of air itself, so that, as a function of the particle'"'"'s inertia and aerodynamic properties, each particle becomes separated from the others and becomes distributed substantially sequentially along a linear path through the air;

directing each particle to intersect two laser beams of known spacing;

measuring the particle'"'"'s transit time between the two laser beams;

establishing measures of the particle'"'"'s velocity from the particle'"'"'s transit time and the laser beam spacing; and

comparing the relative velocities of the particle and stream of air so as to establish the size of the particle;

establishing the particle as a candidate particle if its size is within the range of particles which are potentially biologically viable; and

then

establishing measures of the particle'"'"'s position as a function of time if it is a candidate particle.