Plate assembly for rapid, automatic, high penetration depth examination of samples with a light-sheet microscope, as well as a method for using such plate assembly

1 . A plate assembly ( 1 ) for rapid, automatic, high penetration depth examination of samples ( 12 ) with a light-sheet microscope having a scanning unit,

wherein the plate assembly ( 1 ) comprises:

a first sample holder ( 10 ) configured to receive a plurality of samples ( 12 ) separated from each other, the first sample holder ( 10 ) being a transparent foil sheet and comprising a plurality of cuvettes ( 11 ) individually configured to receive a single one of the plurality of samples ( 12 ),

a sample fixing plate ( 20 ) configured to position the plurality of samples ( 12 ) placed in the cuvettes ( 11 ), wherein the sample fixing plate ( 20 ) comprises a plurality of inserts ( 21 ) configured to be individually insertable into one of the cuvettes ( 11 ) of the first sample holder ( 10 ) in such a way that a chamber ( 22 ) is formed between the first sample holder ( 10 ) and the inserted sample fixing plate ( 20 ) in individual ones of the cuvettes ( 11 ), the chamber ( 22 ) being configured to ensure integrity of a structure of a single sample ( 12 ) in a respective individual one of the cuvettes ( 11 ) of the first sample holder ( 10 ), wherein

at least one slot ( 23 ) is formed next to each insert ( 21 ) to allow transmission of light emitted by a light-sheet microscope,

the sample fixing plate ( 20 ) further comprising at least two opposite sample fixing plate magnets ( 24 ),

a sample holder frame ( 30 ) configured to receive the sample holder ( 10 ) and the sample fixing plate ( 20 ) fitted to each other, the sample holder frame ( 30 ) having a planar flat sheet ( 31 ), which comprises at least one receiving element ( 32 ) configured to receive the first sample holder ( 10 ) and the sample fixing plate ( 20 ) inserted therein, wherein

a bottom of the receiving element ( 32 ) is made of transparent material,

the sample holder frame ( 30 ) comprising a planar flat edge ( 34 ) extending around a circumference of the planar flat sheet ( 31 ), wherein dimensions of the planar flat edge ( 34 ) are configured to allow a scanning unit of the light-sheet microscope to operate continuously,

wherein the sample holder frame ( 30 ) further comprises at least two opposite holder frame magnets ( 35 ) located in the planar flat edge ( 34 ), wherein individual ones of the at least two opposite holder frame magnets ( 35 ) in the edge ( 34 ) have a position substantially matching a position of one of the at least two opposite sample fixing plate magnets ( 24 ) arranged in the sample fixing plate ( 20 ) to connect to each other and thereby hold the sample fixing plate ( 20 ) stable in an inserted position.

2 . The plate assembly ( 1 ) according to claim 1 , wherein the sample holder frame ( 30 ) further comprises a circumferentially extending, protruding side wall ( 36 ), wherein at least two opposite parts of the side wall ( 36 ) comprises at least one L-shaped groove ( 37 ), and the sample fixing plate ( 20 ) comprises fixing tabs arranged opposite each other and/or the plate assembly ( 1 ) further comprises a grid clamping element ( 50 ), either of which is suitable for inserting into the at least one L-shaped groove ( 37 ) and can be fixed stably to press down the sample fixing plate ( 20 ) in the inserted position, respectively.

3 . The plate assembly ( 1 ) according to claim 1 , wherein each individual chamber ( 22 , 42 ) formed in the cuvettes ( 11 ) of the first sample holder ( 10 ) is configured to receive a sample ( 12 ) and a mounting medium, cell culture medium and/or hydrogel together.

4 . The plate assembly ( 1 ) according to claim 1 , wherein the sample holder frame ( 30 ) comprises more than one receiving element ( 32 ) with dimensions that correspond to dimensions of a standard slide, wherein individual ones of the more than one receiving element ( 32 ) are arranged next to each other and separated from each other by a partition wall ( 33 ).

5 . The plate assembly ( 1 ) according to claim 1 , wherein a thickness of the bottom of the receiving elements ( 32 ) is about 150 μm and is made of a material selected from a group consisting of: glass, transparent plastic, transparent resin, or transparent FEP (Fluorinated Ethylene Propylene) foil.

6 . The plate assembly ( 1 ) according to claim 1 , wherein the sample holder frame ( 30 ), except for the bottom of the receiving element ( 32 ), is made of plastic, resin, and/or metal.

7 . The plate assembly ( 1 ) according to claim 1 , wherein the sample ( 12 ) is a 3-dimensional cell culture or tissue sample.

8 . A method of using the plate assembly ( 1 ) according to claim 1 , comprising the steps of:

providing (S 10 ) the first sample holder ( 10 ) and filling the plurality of cuvettes ( 11 ) with a sample ( 12 ) stored in mounting medium, cell culture medium, and/or hydrogel,

providing (S 20 ) the sample fixing plate ( 20 ) and inserting individual ones of the plurality of inserts ( 21 ) into individual ones of the plurality of cuvettes ( 11 ) of the first sample holder ( 10 ) containing samples ( 12 ), thereby forming the chamber ( 22 ) having a volume that ensures integrity of the structure of the single sample ( 12 ),

providing (S 30 ) the sample holder frame ( 30 ) with the at least one receiving element ( 32 ) and placing the sample fixing plate ( 20 ) inserted into the first sample holder ( 10 ) in the at least one receiving element ( 32 ), thereby assembling the plate assembly ( 1 ),

connecting (S 40 ) one or more of the at least two opposite sample fixing plate magnets ( 24 ) of the sample fixing plate ( 20 ) and one or more of the at least two holder frame magnets ( 35 ) of the sample holder frame ( 30 ) to stably fix the plate assembly ( 1 ),

inverting (S 50 ) the plate assembly ( 1 ) in such a way that the cuvettes ( 11 ) of the first sample holder element ( 10 ) filled with samples ( 12 ) face upwards, and

illuminating (S 60 ) the plate assembly ( 1 ) from below with a light-sheet microscope for examining the samples ( 12 ).

9 . The method according to claim 8 , further comprising first filling individual ones of the plurality of cuvettes ( 11 ) of the first sample holder ( 10 ) with a mounting medium, cell culture medium, and/or hydrogel, and then filling with the sample ( 12 ).

10 . The method according to claim 8 , further comprising filling individual ones of the plurality of cuvettes ( 11 ) of the first sample holder ( 10 ) with the sample ( 12 ) stored in a mounting medium, cell culture medium, and/or hydrogel, manually or with a pipetting robot.

11 . The method according to claim 8 , wherein the sample ( 12 ) is a 3-dimensional cell culture or tissue sample.

12 . The method according to claim 8 , further comprising simultaneously using several of the at least one receiving elements ( 32 ) of the sample holder frame ( 30 ) for continuous examination of the samples ( 12 ), without recalibration of the light-sheet microscope.

13 . The method according to claim 8 , wherein each individual chamber ( 22 , 42 ) formed in the cuvettes ( 11 ) of the first sample holder ( 10 ) is configured to receive a sample ( 12 ) and a mounting medium, cell culture medium and/or hydrogel together, further comprising the step of:

before inverting the plate assembly ( 1 ), sliding and stably securing fixing tabs of the sample fixing plate ( 20 ) and/or grid clamping element ( 50 ) into L-shaped grooves ( 37 ) formed on side walls ( 36 ) of the sample holder frame ( 30 ) (S 41 or S 510 ).

14 . The method according to claim 8 , comprising the step of producing any of the first sample holder ( 10 ) from a FEP (fluorinated ethylene propylene) foil by vacuum forming.

15 . The plate assembly ( 1 ) according to claim 1 , further comprising a second sample holder ( 40 ), which is a transparent foil sheet and comprises a plurality of second cuvettes ( 41 ), individual ones of the plurality of second cuvettes ( 41 ) being insertable into a respective one of the cuvettes ( 11 ) of the first sample holder ( 10 ) in such a way that a second chamber ( 42 ) is formed between the first sample holder ( 10 ) and the second sample holder ( 40 ) in an individual one of the cuvettes ( 11 ), the chamber second ( 42 ) being configured to ensure integrity of the structure of the single sample ( 12 ) in the respective individual one of the cuvettes ( 11 ) of the first sample holder ( 10 ), and wherein the individual ones of the plurality of second cuvettes ( 41 ) of the second sample holder ( 40 ) is configured to receive individual inserts ( 21 ) of the sample fixing plate ( 20 ).

16 . The plate assembly ( 1 ) according to claim 15 , wherein the first sample holder ( 10 ) and the second sample holder ( 40 ) are a transparent foil sheet selected from at least one of the group consisting of: ECTFE foil (ethylene-chlorotrifluoroethylene), THV foil (tetrafluoroethylene-hexafluoropropylene-vinylidene fluoride), PFA foil (perfluoroalkoxy), PFA-PG foil, PFA-GK foil, PTFE (polytetrafluoroethylene, Teflon) foil, especially FEP foil, wherein a thickness of the foil sheet is between 25-100 μm, and wherein the first sample holder ( 10 ) and second sample holder ( 40 ) are formed by vacuum forming.

17 . A method of using of the plate assembly ( 1 ) according to claim 15 , comprising the steps of:

providing (S 100 ) the first sample holder ( 10 ) and filling the plurality of cuvettes ( 11 ) with a sample ( 12 ) stored in mounting medium, cell culture medium, and/or hydrogel,

providing (S 200 ) the second sample holder ( 40 ) and inserting the plurality of second cuvettes ( 41 ) into respective ones of the cuvettes ( 11 ) of the first sample holder ( 10 ) containing samples ( 12 ), thereby the second chamber ( 42 ) is formed, which has a volume that ensures the integrity of the structure of the single sample ( 12 ),

providing (S 300 ) the sample fixing plate ( 20 ) and inserting individual ones of the plurality of inserts ( 21 ) into individual ones of the plurality of second cuvettes ( 41 ) of the second sample holder ( 40 ),

providing (S 400 ) the sample holder frame ( 30 ) having the at least one receiving element ( 32 ) and placing the first sample holder ( 10 ), the second sample holder ( 40 ), and the sample fixing plate ( 20 ) in the at least one receiving element ( 32 ), which are aligned one after the other, thereby assembling the plate assembly ( 1 ),

connecting (S 500 ) one or more of the at least two opposite sample fixing plate magnets ( 24 ) of the sample fixing plate ( 20 ) and one or more of the at least two holder frame magnets ( 35 ) of the sample holder frame ( 30 ) to stably fix the plate assembly ( 1 ),

inverting (S 600 ) the plate assembly ( 1 ) in such a way that the cuvettes ( 11 ) of the first sample holder element ( 10 ) filled with samples ( 12 ) face upwards, and

illuminating (S 700 ) the plate assembly ( 1 ) from below with a light-sheet microscope for examining the samples ( 12 ).

18 . A method of using the plate assembly ( 1 ) according to claim 1 , comprising the steps of:

providing (S 7 ) the first sample holder ( 10 ) and sterilizing it,

filling (S 8 ) individual ones of the plurality of cuvettes ( 11 ) of the sterilized first sample holder ( 10 ) with sample-forming material ( 9 ),

placing (S 9 ) the first sample holder ( 10 ) in an incubator, wherein the individual ones of the plurality of cuvettes ( 11 ) of which are filled with sample-forming material ( 9 ), until obtaining a sample ( 12 ), and

sequentially performing steps comprising:

providing (S 10 ) the first sample holder ( 10 ) and filling the plurality of cuvettes ( 11 ) with a sample ( 12 ) stored in mounting medium, cell culture medium, and/or hydrogel,

providing (S 20 ) the sample fixing plate ( 20 ) and inserting individual ones of the plurality of inserts ( 21 ) into individual ones of the plurality of cuvettes ( 11 ) of the first sample holder ( 10 ) containing samples ( 12 ), thereby forming the chamber ( 22 ) having a volume that ensures integrity of the structure of the single sample ( 12 ),

providing (S 30 ) the sample holder frame ( 30 ) with the at least one receiving element ( 32 ) and placing the sample fixing plate ( 20 ) inserted into the first sample holder ( 10 ) in the at least one receiving element ( 32 ), thereby assembling the plate assembly ( 1 ),

connecting (S 40 ) one or more of the at least two opposite sample fixing plate magnets ( 24 ) of the sample fixing plate ( 20 ) and one or more of the at least two holder frame magnets ( 35 ) of the sample holder frame ( 30 ) to stably fix the plate assembly ( 1 ),

inverting (S 50 ) the plate assembly ( 1 ) in such a way that the cuvettes ( 11 ) of the first sample holder element ( 10 ) filled with samples ( 12 ) face upwards, and

illuminating (S 60 ) the plate assembly ( 1 ) from below with a light-sheet microscope for examining the samples ( 12 ).

19 . The method according to claim 18 , wherein the sample-forming material ( 9 ) comprises single cells in a cell culture medium or hydrogel, or one or more small cell aggregates and wherein the sample ( 12 ) is a 3-dimensional cell culture of living cells.

20 . The method according to claim 18 , wherein steps S 7 -S 9 are performed in a sterile environment.