Imaging system and method for enhancing microscopic images of unstained cells

1. An imaging system ( 100 ) for enhancing microscopic images of unstained cells, the imaging system ( 100 ) comprising:

a light source ( 102 ) for producing light ( 120 a );

a sample holder ( 109 ) for containing cells to be imaged;

a condenser ( 104 ) for focussing the light ( 120 b ) at a focal plane within the sample holder ( 109 ) on the cells to be imaged;

a translation mechanism for moving the focal plane of the light ( 120 b ) relative to the sample holder ( 109 ); and

a detector system ( 112 ) configured to acquire a plurality of images at respective focal planes within the sample holder ( 109 ) and to process the plurality of images to provide an resultant enhanced image (U r ), wherein processing of the plurality of images comprises:

applying a pixel-by-pixel minimisation operation to each of the plurality of images to identify a dark image (U 1 dark );

applying a non-linear top-hat transform to the dark image (U 1 dark ) to obtain an intermediate image (U 2 dark ); and

processing the intermediate image (U 2 dark ) to produce the resultant enhanced image (U r ).

2. The imaging system ( 100 ) of claim 1 , wherein the detector system ( 112 ) is further configured to identify a best focus within the sample holder ( 109 ) and to acquire the plurality of images at respective focal planes within the sample holder ( 109 ) near to the best focus.

3. The imaging system ( 100 ) of claim 1 , further comprising an aperture stop ( 106 ) provided between the light source ( 102 ) and the detector system ( 112 ).

4. The imaging system ( 100 ) of claim 3 , wherein the size of the aperture stop ( 106 ) is variable.

5. The imaging system ( 100 ) of claim 1 , wherein:

the dark image is defined by:

U 1 dark =min k {U ( Z k )},

where U(Z k ) is the k th image in the Z-plane;

the intermediate image is defined by:

U 2 dark =NTH ( inv[U 1 dark ];ε|.),

where NTH is a non-linear ε-pixel top-hat operation applied to the dark image, inv[ . . . ] is image inversion operation; and

the resultant enhanced image is obtained by applying the following transform:

U

r

=

U

T

+

(

U

2

dark

-

U

T

)

+



U

2

dark

-

U

T



2

,

where U T defines a predetermined threshold image.

6. The imaging system ( 100 ) of claim 1 , wherein processing of the plurality of images further comprises applying thresholding to the resultant enhanced image (U r ).

7. An imaging system ( 100 ) for enhancing microscopic images of unstained cells, the imaging system ( 100 ) comprising:

a light source ( 102 ) for producing light ( 120 a );

a sample holder ( 109 ) for containing cells to be imaged;

a condenser ( 104 ) for focussing the light ( 120 b ) at a focal plane within the sample holder ( 109 ) on the cells to be imaged;

a translation mechanism for moving the focal plane of the light ( 120 b ) relative to the sample holder ( 109 ); and

a detector system ( 112 ) configured to acquire a plurality of images at respective focal planes within the sample holder ( 109 ) and to process the plurality of images to provide an resultant enhanced image (U r ), wherein processing of the plurality of images comprises:

applying a pixel-by-pixel maximisation operation to each of the plurality of images to identify a bright image (U 1 bright );

applying a non-linear top-hat transform to the bright image (U 1 bright ) to obtain an intermediate image (U 2 bright ); and

processing the intermediate image (U 2 bright ) to produce the resultant enhanced image (U r ).

8. The imaging system ( 100 ) of claim 7 , wherein:

the bright image is defined by:

U 1 bright =max k {U ( Z k )},

where U(Z k ) is the k th image in the Z-plane;

the intermediate image is defined by:

U 2 bright =NTH ( U 1 bright ;ε|.),

where NTH is a non-linear ε-pixel top-hat operation applied to the bright image; and

the resultant enhanced image is obtained by applying the following transform:

U

r

=

U

T

+

(

U

2

bright

-

U

T

)

+



U

2

bright

-

U

T



2

,

where U T defines a predetermined threshold image.

9. The imaging system ( 100 ) of claim 7 , wherein processing of the plurality of images further comprises applying thresholding to the enhanced processed image (U r ).

10. The imaging system ( 100 ) of claim 7 , wherein the detector system ( 112 ) is further configured to identify a best focus within the sample holder ( 109 ) and to acquire the plurality of images at respective focal planes within the sample holder ( 109 ) near to the best focus.

11. The imaging system ( 100 ) of claim 7 , further comprising an aperture stop ( 106 ) provided between the light source ( 102 ) and the detector system ( 112 ).

12. The imaging system ( 100 ) of claim 11 , wherein the size of the aperture stop ( 106 ) is variable.

13. A method ( 200 ) for enhancing microscopic images of unstained cells, the method ( 200 ) comprising:

acquiring ( 202 ) a plurality of images at respective focal planes within a sample holder ( 109 ); and

processing ( 210 ) the plurality of images to provide an resultant enhanced image (U r ), wherein the step ( 210 ) of processing of the plurality of images comprises:

applying a pixel-by-pixel minimisation operation to each of the plurality of images to identify a dark image (U 1 dark );

applying a non-linear top-hat transform to the dark image (U 1 dark ) to obtain an intermediate image (U 2 dark ); and

processing the intermediate image (U 2 dark ) to produce the resultant enhanced image (U r ).

14. The method ( 200 ) of claim 13 , further comprising the step of setting an aperture stop prior to acquiring the plurality of images.

15. The method ( 200 ) of claim 13 , wherein:

the dark image is defined by:

U 1 =min k {U ( Z k )},

where U(Z k ) is the k th image in the Z-plane;

the intermediate image is defined by:

U 2 dark =NTH ( inv[U 1 dark ];ε|.),

where NTH is a non-linear ε-pixel top-hat operation applied to the dark image; inv[ . . . ] is image inversion operation and

the resultant enhanced image is obtained by applying the following transform:

U

r

=

U

T

+

(

U

2

dark

-

U

T

)

+



U

2

dark

-

U

T



2

,

where U T defines a predetermined threshold image.

16. The method ( 200 ) of claim 13 , wherein the step ( 210 ) of processing the plurality of images further comprises applying thresholding to the enhanced processed image (U r ).

17. A method ( 200 ) for enhancing microscopic images of unstained cells, the method ( 200 ) comprising:

acquiring ( 202 ) a plurality of images at respective focal planes within a sample holder ( 109 ); and

processing ( 210 ) the plurality of images to provide an resultant enhanced image (U r ), wherein the step ( 210 ) of processing the plurality of images comprises:

applying a pixel-by-pixel maximisation operation to each of the plurality of images to identify a bright image (U 1 bright );

applying a non-linear top-hat transform to the bright image (U 1 bright ) to obtain an intermediate image (U 2 bright ); and

processing the intermediate image (U 2 bright ) to produce the resultant enhanced image (U r ).

18. The method ( 200 ) of claim 17 , wherein:

the bright image is defined by:

U 1 bright =max k {U ( Z k )},

where U(Z k ) is the k th image in the Z-plane;

the intermediate image is defined by:

U 2 bright =NTH ( U 1 bright ;ε|.),

where NTH is a non-linear ε-pixel top-hat operation applied to the bright image; and

the resultant enhanced image is obtained by applying the following transform:

U

r

=

U

T

+

(

U

2

bright

-

U

T

)

+



U

2

bright

-

U

T



2

,

where U T defines a predetermined threshold image.

19. The method ( 200 ) of claim 17 , further comprising the step of setting an aperture stop prior to acquiring the plurality of images.

20. The method ( 200 ) of claim 17 , wherein the step ( 210 ) of processing the plurality of images further comprises applying thresholding to the enhanced processed image (U r ).