Analysis of liquid-based biological substances by microscopy, and systems comprising such carriers and microscopes

Abstract

A carrier object comprises a glass slide (11) and a cover glass (12) fixed by adhesive lines (13) perimetrically defining individual compartments (14) for receiving samples of biological matter; characterized in that the upper surface (17) of the glass slide is bare and uncovered on at least one front edge region (41) and on at least one back edge region (42), each edge region having: an extension in the front-back direction at least equal to 2.5 mm; and an extension in the left-right direction at least equal to the spacing between the respective axes of the two individual compartments farthest apart from each other. The system comprises the carrier object (40) and a microscope for clamping the carrier object between a stage and a platen having pressing fingers distal ends of which contact only the edge regions (41, 42).

Description

Technical Field

[0001] This invention relates to the microscopic analysis of liquid-based biological substances, such as animal semen. More specifically, the invention relates to a carrier for such substances, designed for mounting in a microscope configured to perform this analysis, and also to a system comprising such a carrier and such a microscope. Background Technology

[0002] Specifically, through European Patent EP0809815 corresponding to US Patent US6551554 and US Design Patent USD566849S, a carrier for analyzing liquid-based biological substances such as animal semen by microscope is known. The carrier includes a glass slide, a coverslip, and an adhesive thread. The adhesive thread is positioned between the glass slide and the coverslip to secure them to each other, while defining a separate compartment for receiving the biological sample on its periphery. The glass slide is a parallelepiped with two principal surfaces, namely an upper surface and a lower surface, and side surfaces extending from one of the principal surfaces to the other, namely a left surface, a right surface, a front surface, and a rear surface. The distance between the left and right surfaces is greater than the distance between the front and rear surfaces, and the distance between the front and rear surfaces is itself greater than the distance between the lower and upper surfaces. The carrier has a left-right orientation, transverse to the left and right surfaces, and parallel to the front, rear, and two principal surfaces; a front-back orientation... The coverslip is a parallelepiped with two main surfaces, namely the upper and lower surfaces, and side surfaces extending from one of the two main surfaces to the other, namely the left, right, front, and rear surfaces. The lower surface of the coverslip faces the upper surface of the slide, while the front, rear, left, and right surfaces of the coverslip face the front, rear, left, and right surfaces of the slide and are separated from them by a distance. Each individual compartment is defined by the upper surface of the slide and the lower surface of the coverslip in the downward direction, and is defined by an adhesive line on the periphery. Each individual compartment has an opening to a groove on only one side and an opening to a vent on the other side. Each groove and each vent are defined by an adhesive line. Each groove faces one of the front and rear surfaces of the slide, and each vent faces the other of the front and rear surfaces of the slide.

[0003] To analyze biological material, a drop of biological material is placed in each groove of the microscope mount. Each droplet is then carried by capillary action to a separate compartment connected by the grooves. The mount is then mounted in a microscope, with the individual compartments placed one after another under a lens for observation of the material.

[0004] A system comprising such a carrier and such a microscope is also known. The microscope includes a lens, a stage on which the carrier rests, a lower surface of a slide abuts the stage, and a separate compartment aligned with the lens. The microscope also includes a frame on which the lens is mounted, and the stage is mounted on the frame to allow translational movement along the left-right direction of the carrier to sequentially align other separate compartments with the lens. The microscope also includes a pressure plate and a clamping member configured to actuate the pressure plate and the stage toward each other. The pressure plate is hinged to the stage to allow it to be displaced from or folded onto the carrier. The system is further configured to focus the lens via autofocus. Summary of the Invention

[0005] In a first aspect, the present invention proposes a carrier similar to the carrier described above, but the carrier of the present invention has improved performance, particularly regarding the positioning of the individual compartments relative to the lenses of the microscope.

[0006] Therefore, the present invention proposes a sample carrier for microscopic analysis of liquid-based biological materials, such as animal semen. The sample carrier includes a glass slide, a coverslip, and an adhesive thread. The adhesive thread is disposed between the glass slide and the coverslip to fix the glass slide and the coverslip to each other, while defining a separate compartment for receiving the biological material sample on the periphery. The glass slide is a parallelepiped with two main surfaces, namely an upper surface and a lower surface, and side surfaces extending from one of the two main surfaces to the other, namely a left surface, a right surface, a front surface, and a back surface. The distance between the left surface and the right surface is greater than the distance between the front surface and the back surface. The distance between surfaces is greater between the front and back surfaces than between the bottom and top surfaces; the object being carried has the following characteristics: in the left-right direction, it is transverse to the left and right surfaces, and parallel to the front, back, and two main surfaces; in the front-back direction, it is transverse to the front and back surfaces, and parallel to the left, right, and two main surfaces; in the up-down direction, it is transverse to the two main surfaces and parallel to the side surfaces; the coverslip is a parallelepiped, having two main surfaces, namely the top and bottom surfaces, and side surfaces extending from one of the two main surfaces to the other, namely the left, right, and front surfaces. The back surface; the lower surface of the coverslip faces the upper surface of the slide, while the front, back, left, and right surfaces of the coverslip face the front, back, left, and right surfaces of the slide, respectively, and are spaced apart from them; each individual compartment is defined in the up-down direction by the upper surface of the slide and the lower surface of the coverslip, and is peripherally defined by an adhesive line; each individual compartment has an opening to a groove on only one side and an opening to a vent on the other side; each groove and each vent is defined by an adhesive line; each groove faces one of the front and back surfaces of the slide, and each vent faces the front of the slide. The other of the upper surface and the rear surface; characterized in that the upper surface of the slide is exposed and uncovered in at least one front edge region and at least one rear edge region, the front edge region extending along the front surface of the slide and the rear edge region extending along the rear surface of the slide, each of the front edge region and the rear edge region having: an extension dimension in the front-rear direction of at least 2.5 mm; and an extension dimension in the left-right direction, the extension dimension in the left-right direction being at least equal to the distance between the respective axes of the two most distant individual compartments and not greater than the distance between the left and right surfaces of the slide.

[0007] By means of the edge region, the positioning of the object relative to the microscope according to the invention can be achieved by pressing the upper surface of the slide instead of the lower surface of the slide as is conventional.

[0008] This allows for particularly precise positioning of the sample to be analyzed in the top-bottom direction, as this avoids uncertainties in the thickness of the slide due to manufacturing tolerances.

[0009] It should be noted that the term "naked" in relation to the upper surface of a slide means that the upper surface does not directly bear any locally added coverings, such as adhesive lines or ink lines.

[0010] It should also be noted that the expression “uncovered” in relation to the upper surface of the slide means that the upper surface is not opposite any element (such as a coverslip) that could constitute an obstacle for another element to move in the down-up direction toward the object to make contact with the upper surface.

[0011] The edge region allows direct access to the upper surface of the slide carrying the object, enabling mechanical determination of its position, and consequently, the positions of other elements, whose positions relative to the upper surface are fixed and / or known. It should be particularly noted that mechanically determining the position of the upper surface is equivalent to mechanically determining the position of the individual compartments defined by the upper surface.

[0012] The invention is also based on the observation that the optical quality of the surface of the slide (which is necessary to minimize interference with the light passing through the slide to illuminate the sample present in the individual compartment) relates to the mechanical regularity of the surface that facilitates sliding; and that pressure can be applied to the surface while maintaining the ability to slide on it.

[0013] Therefore, the front and rear edge regions can serve as sliding tracks for at least one pressing element, such as the pressing finger explained later, with the upper surface abutting against the pressing element through its edge regions in order to mechanically determine its position.

[0014] It should also be noted that the position of the edge areas, i.e., along the front and rear surfaces, means that these areas are as far apart as possible from each other, which is beneficial to the accuracy of the mechanically determined positioning by the pressing element and the stability during the sliding process.

[0015] Regarding the lower limit of the edge area's extension in the front-back direction, i.e., 2.5mm, the applicant determined that, due to the manufacturing tolerances of various components of the system, a value of at least 2.5mm ensures that the pressing finger remains in contact only with the front and back edge areas, regardless of the stage's position in the left-right direction.

[0016] The particularly simple, convenient, and economical advantages of the carrier according to the present invention are as follows:

[0017] - For at least one of the front edge region and the rear edge region, the extension dimension in the left-right direction is equal to the distance between the left and right surfaces of the slide;

[0018] - Each groove has a front-to-back extension dimension smaller than the front-to-back extension dimension of each of the front and rear edge regions.

[0019] - The distance between the left and right surfaces of the slide is between 74.8 mm and 75.2 mm; the distance between the front and back surfaces of the slide is between 24.8 mm and 25.2 mm; the distance between the left and right surfaces of the coverslip is between 31.8 mm and 32.2 mm; and the distance between the front and back surfaces of the coverslip is between 16.8 mm and 16.2 mm; and / or

[0020] - All grooves face one of the front and back surfaces of the slide, and all vents face the other of the front and back surfaces of the slide.

[0021] In a second aspect, the present invention proposes a system for analyzing liquid-based biological substances, the system comprising a carrier as described above, and a microscope comprising: a lens; a stage configured to receive the carrier in a first observation position, wherein the lower surface of a slide rests against the stage, and one of the individual compartments is aligned with the lens; a frame on which the lens is mounted at least one fixed predetermined position, and the stage is mounted on the frame to be translatable in a left-right direction along the carrier to sequentially align other individual compartments with the lens; and a pressure plate, at least partially The plate extends relative to the object being carried; and a clamping member is configured to actuate the pressure plate and the platform toward each other, such that the object is clamped between the pressure plate and the platform; the pressure plate is fixedly mounted on the frame and includes a base and pressing fingers, the pressing fingers projecting from the base toward the platform and having their distal ends contacting only edge regions, wherein at least two pressing fingers contact one of the front edge region and the rear edge region, and at least one pressing finger contacts the other of the front edge region and the rear edge region; the platform has positioning stops for fixing the object relative to the platform in a left-right direction.

[0022] Because the pressure plate is fixedly installed on the frame and the upper surface of the glass slide is held against the pressing finger by the stage under the action of the clamping member, the pressing finger determines the position of the upper surface relative to the frame in the downward and upward direction.

[0023] It should be noted that, in this way, the position of the upper surface relative to the frame does not depend on the thickness of the slide (i.e., the distance between the lower and upper surfaces).

[0024] As the stage moves laterally, the pressing finger slides on the edge region. Since the lateral extension of the edge region is at least equal to the distance between the corresponding axes of the two furthest individual compartments, the pressing finger remains in contact with the edge region regardless of which individual compartment is aligned with the lens.

[0025] Therefore, the position of the upper surface of the slide in the downward direction is determined in the same way by the pressing finger, regardless of which individual compartment is aligned with the lens.

[0026] Once the lens has been focused for a given individual compartment, this specifically allows that focus to be maintained for each of the other individual compartments.

[0027] This further allows the focus to be maintained on another object instead of the first object that has been focused on, because the position of the upper surface of the other object along the down-up direction will be determined by the pressing finger in the same way as on the first object.

[0028] It should be noted that this focusing can thus correspond to the factory settings made during microscope manufacturing.

[0029] It should also be noted that in existing systems, the mechanical reference plane is the stage on which the lower surface of the slide rests, making the position of the upper surface of the slide dependent on the thickness of the slide. Therefore, these systems do not provide the aforementioned advantages.

[0030] The particularly simple, convenient, and economical advantages of the system according to the invention are as follows:

[0031] - The pressure plate includes four pressing fingers, two of which contact the front edge area and the other two of which contact the rear edge area;

[0032] - One of the stage and the pressure plate is magnetic, and the clamping member includes a magnet fixed to the other of the stage and the pressure plate;

[0033] - The base has an opening through which the lens points to a separate compartment aligned with the lens at a first observation position, and at least two of the pressing fingers that contact the same edge area are arranged on both sides of the opening.

[0034] - The frame includes a mounting wall, and the base includes two arms, each arm projecting laterally from the mounting wall. The two arms extend facing each other and define the opening. Each arm is provided with two pressing fingers, one pressing finger contacting the front edge region and the other pressing finger contacting the rear edge region.

[0035] -The clamping member includes a magnet housed in the arm;

[0036] - One of the stage and the pressure plate has a ramp, and the other of the stage and the pressure plate has a protrusion. The protrusion is configured to engage with the ramp during the translational movement of the stage in the left and right directions in order to overcome the force applied by the clamping members to separate the stage from the pressure plate to a position in which the load can be accessed to remove the load from the stage.

[0037] - The stage is mechanically connected to the frame via a sliding pivot connector, which includes a cylindrical guide oriented longitudinally in the left-right direction and a cylinder mounted on the guide for sliding, with a gap between the cylinder and the guide; and / or

[0038] - The lens has a fixed focal length.

[0039] The present invention also relates to the use of the above-described system, wherein the liquid-based biological material is animal semen, and the analysis includes a sperm counting step. Attached Figure Description

[0040] The invention will now continue to be described with reference to the accompanying drawings and a detailed description of exemplary embodiments given below by way of illustrative and non-limiting examples. In the drawings:

[0041] - Figure 1 It is a view of a carrier for analyzing liquid-based biological materials by microscope, based on existing technology.

[0042] - Figure 2 It is similar to the carrier of the present invention. Figure 1 And the view is in the same proportion.

[0043] - Figure 3 The system according to the present invention (in) Figure 9-12 A highly schematic cross-sectional view (shown in more detail below) of the system, which includes a microscope and Figure 2 The illustrated object is in an observation configuration, wherein the object is held between the stage of the microscope and the pressing finger of the pressure plate, and one of the individual compartments of the object is aligned with the lens; the profile is along the front-rear direction of the object and passes through the middle of the individual compartment.

[0044] - Figure 4 yes Figure 3 The diagram shows a cross-sectional view of the system, but here it is along the left-right direction of the load and passes through the middle of the individual compartment.

[0045] - Figure 5 Similar to the stage moving left and right to align another separate compartment with the lens. Figure 4 The view.

[0046] - Figure 6 It is similar to Figure 2 The view shows the axes of the individual compartments, and when the stage is located... Figure 4 The position of the middle and the stage are located Figure 5 The position of the pressure plate on the surface of the glass slide.

[0047] - Figure 7 It is similar to Figure 6 The view shows when the stage moves from... Figure 4 The position in the middle is moved to Figure 5 The path of each pressing finger on the pressure plate when it is in position.

[0048] - Figure 8 It is similar to Figure 3 However, this is a view of the system after the platform has been removed from the pressure plate, changing it to a loading / unloading configuration in which the system user can access the loaded object.

[0049] - Figure 9 yes Figures 2 to 8 The diagram shows a detailed exploded perspective view of the system, but the lenses are not shown.

[0050] - Figure 10 It is similar to Figure 9 The view is for systems in an assembled state and in a clean configuration, where the stage is far from the pressure plate.

[0051] - Figure 11 Is it like this? Figure 9 and Figure 10 The system shown is a cross-sectional view taken along the front-to-back direction of the load and passing through one of its individual compartments. The system is located in... Figure 8 The loading / unloading configuration is schematically represented in the diagram.

[0052] - Figure 12 yes Figure 9 and Figure 11 A side view of a portion of the system shown, in which the system is located Figures 3 to 5 The observation configuration is schematically represented in the diagram.

[0053] - Figure 13 It is a variation of the carrier according to the invention, similar to Figure 7 The view. Detailed Implementation

[0054] In respectively Figure 1 and Figure 2 Both the carrier 10 and the carrier 40 shown include a glass slide 11, a cover glass 12, and an adhesive line 13 for carrying the object. The adhesive line 13 is disposed between the glass slide 11 and the cover glass 12 to secure them to each other, while defining a separate compartment 14 on the periphery.

[0055] First, it should be noted that, for the sake of simplicity, the same numerical designations are used for the corresponding components of carrier 10 and carrier 40 in this report.

[0056] Therefore, in the following description of the carrier 10, if a component of the reference carrier 10 is in Figure 1 If it is not visible, for the purpose of understanding, one of the diagrams showing the visible corresponding elements of the carrying object 40 can be referred to.

[0057] Each of the individual compartments 14 of the carrier 10 is configured to receive diluted or undiluted liquid-based biological material, in this case, a sample of animal semen.

[0058] The object 10 here includes multiple individual compartments 14.

[0059] Each individual compartment 14 has an opening to a recess 15 on only one side and an opening to a vent 16 on the other side, with each recess 15 and each vent 16 defined by an adhesive line 13.

[0060] To fill the individual compartment 14 with a biological sample, a droplet of material (not shown) is deposited in the corresponding groove 15. The droplet is then entrained into the individual compartment 14 by capillary action, while the air initially present in the individual compartment 14 escapes through the vent 16.

[0061] Both the slide 11 and the cover glass 12 are made of float glass covered with a surfactant film, which promotes the entrainment of droplets into a separate compartment 14 by capillary action.

[0062] Both the slide 11 and the coverslip 12 are transparent.

[0063] The slide 11 is a parallelepiped with two main surfaces: an upper surface 17 and a lower surface 18. Figure 3 and Figure 4 ); and side surfaces extending from one main surface to another, namely left surface 19, right surface 20, front surface 21 and rear surface 22.

[0064] The distance between the left surface 19 and the right surface 20 is greater than the distance between the front surface 21 and the rear surface 22. The distance between the front surface 21 and the rear surface 22 is itself greater than the distance between the lower surface 18 and the upper surface 19.

[0065] The distance between the left surface 19 and the right surface 20 is between 74.8 mm and 75.2 mm.

[0066] The distance between the front surface 21 and the rear surface 22 is between 24.8 mm and 25.2 mm.

[0067] The distance between the lower surface 18 and the upper surface 19, which is the thickness of the glass slide 11 that supports the object, is approximately 1 mm.

[0068] The carrier 10 has the following characteristics: in the left-right direction, it is transverse to the left surface 19 and the right surface 20, and parallel to the front surface 21, the rear surface 22, and the two main surfaces 17 and 18; in the front-back direction, it is transverse to the front surface 21 and the rear surface 22, and parallel to the left surface 19, the right surface 20, and the two main surfaces 17 and 18; and in the up-down direction, it is transverse to the two main surfaces 17 and 18 and parallel to the side surfaces 19, 20, 21, and 22.

[0069] The coverslip 12 is a parallelepiped and has: two main surfaces, namely the upper surface 23 and the lower surface 24; and side surfaces extending from one of the two main surfaces to the other, namely the left surface 25, the right surface 26, the front surface 27 and the back surface 28.

[0070] The distance between the left surface 25 and the right surface 26 is greater than the distance between the front surface 27 and the rear surface 28. The distance between the front surface 27 and the rear surface 28 is itself greater than the distance between the lower surface 24 and the upper surface 23.

[0071] The distance between the left surface 25 and the right surface 26 is between 31.8 mm and 32.2 mm.

[0072] The distance between the front surface 27 and the rear surface 28 is between 20.8 mm and 21.2 mm.

[0073] The distance between the lower surface 24 and the upper surface 23, which is the thickness of the cover glass 12, is 0.7 mm + / - 0.05 mm.

[0074] The lower surface 24 of the coverslip 12 faces the upper surface 17 of the slide 11, while the front surface 27, rear surface 28, left surface 25 and right surface 26 of the coverslip 12 face the front surface 21, rear surface 22, left surface 19 and right surface 20 of the slide 11 respectively and are at a distance from them.

[0075] Here, the cover glass 12 is centered relative to the slide 11 in the left-right direction.

[0076] Each individual compartment 14 is defined in the upper direction by the upper surface 17 of the slide 11 and the lower surface 24 of the cover glass 12, respectively, and is defined on the periphery by an adhesive line 13.

[0077] The thickness of adhesive line 13 is 20 μm here.

[0078] It should be noted that, in Figures 3 to 5 and Figure 8 In the middle, for readability reasons, the thickness of adhesive line 13 is exaggerated.

[0079] Here, the capacity of each individual compartment 14 is 3 μl.

[0080] The individual compartments 14 follow each other along the left and right directions, and are placed side by side here.

[0081] As described above, each individual compartment 14 has an opening to a recess 15 on one side and an opening to a vent 16 on the other side. The recess 15 faces the rear surface 22 of the slide 11, and the vent 16 faces the front surface 21 of the slide 11.

[0082] Here, all the grooves 15 face the rear surface 22, and all the vents 16 face the front surface 21.

[0083] Each groove 15 is defined here by a portion 29 of adhesive line 13, which is shaped as a U and extends to an end 30, which is located strictly outside the space between the slide 11 and the cover glass 12, and is flush with the rear surface 22 of the slide 11.

[0084] Each vent 16 is defined here by straight sections 31 of adhesive lines 13 extending face to face, each straight section being strictly outside the space between the slide 11 and the coverslip 12 and extending to the corresponding end 32, which is flush with the front surface 21 of the slide 11.

[0085] Figure 2 The carrier 40 shown is similar to the carrier 10, except that the coverslip 12 and adhesive line 13 of the carrier 40 are smaller in size along the front-to-back direction, and the upper surface 17 of the slide 11 of the carrier 40 is exposed and uncovered on the front edge region 41 and the rear edge region 42.

[0086] More precisely, the distance between the front surface 27 and the back surface 28 of the coverslip 12 is between 16.8 mm and 16.2 mm.

[0087] It should be noted that the length of the individual compartment 14 of the object 40, i.e. the distance between the recess 15 and the vent 16—which here corresponds to the distance between the front surface 27 and the rear surface 28 of the cover glass 12—is also less than the length of the individual compartment 14 of the object 10.

[0088] The front edge region 41 extends along the front surface 21 of the slide 11, while the rear edge region 42 extends along the rear surface 22 of the slide 11.

[0089] Here, the front surface 21 and the rear surface 22 of the slide 11 are directly located on the edges of the front edge region 41 and the rear edge region 42, respectively.

[0090] The front edge region 41 has an extension dimension of 2.5 mm in the front-to-back direction and an extension dimension in the left-to-right direction equal to the distance between the left surface 19 and the right surface 20 of the slide 11.

[0091] The extension limit of the front edge region 41 in the front-to-back direction is Figure 2 The dotted line 43 indicates that it is flush with each end 32 of the vent 16.

[0092] This is because, beyond the rearward limit 43, the upper surface 17 is covered by the adhesive line 13 and / or located opposite the lower surface 24 of the cover glass 12, and is therefore not bare and uncovered.

[0093] The rear edge region 42 has an extension dimension of 2.6 mm in the front-to-back direction and an extension dimension in the left-to-right direction equal to the distance between the left surface 19 and the right surface 20 of the slide 11.

[0094] The extension limit of the rear edge region 42 in the front-to-back direction is Figure 2 The dotted line 44 indicates that the groove is flush with the end 30 of the groove 15.

[0095] This is because, beyond the forward limit 44, the upper surface 17 is covered by the adhesive line 13 and / or located opposite the lower surface 24 of the cover glass 12, and is therefore not bare and uncovered.

[0096] Typically, each of the front edge region 41 and the rear edge region 42 has an extension dimension in the front-rear direction of no more than 3 mm, or in variations, the extension dimension in the front-rear direction is equal to a value greater than 3 mm, such as 3.5 mm, 4 mm, 4.5 mm, 5 mm, or even greater than 5 mm.

[0097] Furthermore, it should be noted that the longitudinal extension dimension of each groove 15, i.e. the distance between the rear surface 28 of the cover glass 12 and the limit 44 (which is flush with the end 30 of the groove 15) measured at the upper surface 17, is equal to 2.4 mm, which is significantly smaller than the longitudinal extension dimension of the front edge region 41 (2.5 mm) and the longitudinal extension dimension of the rear edge region 42 (2.6 mm).

[0098] Figures 3 to 5 and Figure 8 A system 100 for analyzing liquid-based biological material is shown, the system including a carrier 40 and a microscope 50.

[0099] exist Figures 3 to 5In the observation configuration, the microscope 50 is coupled with the carrier 40 to observe a biological material sample contained in a separate compartment 14.

[0100] The microscope 50 includes a lens 51 and a stage 52, the stage 52 being configured to receive a slide 40 at at least one observation position, in which the lower surface 18 of a slide 11 rests against the stage 52, and one of the individual compartments 14 is aligned with the lens 51.

[0101] exist Figure 3 and Figure 4 In the middle, the object 40 is in the first observation position, and in this first observation position, the compartment 14 aligned with the lens 51 is... Figure 2 The compartment seen from the far left.

[0102] As explained in more detail below, the microscope 50 is configured to arrange the object 40 at multiple observation positions, at each observation position, the stage 52 receives the object 40, and a corresponding separate compartment 14 is aligned with the lens 51.

[0103] For example, in Figure 5 In the middle, the object 40 is in the second observation position, in which the compartment 14 aligned with the lens 51 is... Figure 2 The compartment seen on the far right.

[0104] The microscope 50 also includes a frame ( Figures 3 to 5 and Figure 8 Not shown in the image, but Figures 9 to 11 The mounting wall 60 is visible in the image. The lens 51 is fixedly mounted on the frame, and the stage 52 is mounted on the frame so that it can be translated in the left and right directions along the object 40 so that other individual compartments 14 can be aligned with the lens 51 in turn.

[0105] The mechanical connection between the stage 52 and the frame will be achieved by means of Figures 9 to 12 It will be described in detail below.

[0106] The stage 52 is further configured to receive the object 40 at a predetermined position.

[0107] To position and fix the object 40 in the predetermined position, the stage 52 includes positioning stops, each of which is configured to engage with a corresponding side surface of the slide 11 opposite to the stop, so as to fix the object 40 in a direction transverse to the corresponding side surface (left-right or front-back, as appropriate).

[0108] The positioning stops here include two front positioning stops 33 opposite to the front side surface 21, a left positioning stop 34 opposite to the left side surface 19, a right positioning stop 35 opposite to the right side surface 20, and two rear positioning stops 36 opposite to the rear side surface 22, so that the object 40 is fixed relative to the stage 52 in both directions along the front-rear axis and in both directions along the left-right axis.

[0109] The positioning stop is formed by a pin protruding from the upper surface 37 of the stage 52, and the lower surface 18 of the slide 11 rests against the upper surface.

[0110] The stage 52 also includes a cutout 45 that extends relative to each of the individual compartments 14 and is configured to allow light from an illumination element (not shown) disposed under the stage 52 to reach each individual compartment 14.

[0111] The microscope 50 also includes a pressure plate 53 and a clamping member, the pressure plate 53 extending at least partially opposite the object 40, the clamping member being configured to actuate the pressure plate 53 and the stage 52 toward each other such that the object 40 is clamped between the pressure plate 53 and the stage 52.

[0112] The clamping member here includes a magnet 54 fixed to the pressure plate 53, and the stage 52 is magnetic, the magnet 54 being configured to actuate the stage 53 toward the pressure plate 53.

[0113] The pressure plate 53 is fixedly mounted on the frame and includes a base 55 and pressing fingers 56, each pressing finger protruding from the base 55 toward the stage 52 to the distal end 57.

[0114] The distal ends 57 of the pressing fingers 56 each contact only the edge regions 41 and 42 of the upper surface 17 of the slide 11.

[0115] Therefore, the slide 11 is held between the pressing finger 56 and the stage 52, and the slide 11 is in contact with the pressure plate 53 only through the pressing finger 56.

[0116] Here, the pressure plate 53 includes four pressing fingers 56, two of which are in contact with the front edge region 41 and the other two fingers 56 are in contact with the rear edge region 42.

[0117] The four pressing fingers 56 are arranged here so that each is located at the corresponding vertex of the imaginary rectangle.

[0118] The base 55 also has an opening 58 through which the lens 51 points to a separate compartment 14 aligned with the lens 51, where the lens 51 is partially received in the opening 58.

[0119] Pressing fingers 56 that contact the front edge region 41 are located on both sides of the opening 58. Pressing fingers 56 that contact the rear edge region 42 are located on both sides of the opening 58.

[0120] Lens 51 has a fixed focal length here.

[0121] System 100 is configured such that the focal point 59 of lens 51 is located approximately in the middle of a separate compartment 14 aligned with lens 51.

[0122] As the stage 52 moves horizontally along the object 40, the pressing fingers 56 slide on their respective edge regions 41 and 42, while each remains in contact only with the edge region.

[0123] exist Figure 6 The axis 38 of each individual compartment 14 has been marked.

[0124] When a separate compartment 14 is aligned with lens 51 to observe the sample received in the compartment 14, the axis 38 of the compartment 14 is more precisely aligned with the optical axis of lens 51.

[0125] Therefore, when the stage is Figure 4 The location of the middle and its Figure 5 Between the positions in the middle, the platform 52 has moved a distance equal to the distance between the corresponding axes 38 of the leftmost and rightmost individual compartments 14, that is, the two individual compartments 14 that are furthest apart from each other on the object 40.

[0126] The distance between the axes 38 of two adjacent compartments is 7.8 mm, so the distance between the axes 38 of the two most distant individual compartments 14 is 23.4 mm.

[0127] exist Figure 6 In the middle, when the stage 52 is in Figure 4 The corresponding position of the pressing finger 56 when the stage 52 is in the middle is further marked by four black rectangles 46. Figure 5 The corresponding positions of these pressing fingers 56 when they are in the middle are marked by four shaded rectangles 47.

[0128] Each black rectangle 46 or shaded rectangle 47 corresponds to the portion of the upper surface 17 that contacts the distal end 57 of the pressing finger 56, which here has an extension dimension of approximately 1 mm in the front-back direction and an extension dimension of approximately 3 mm in the left-right direction.

[0129] exist Figure 7 In the middle, when the platform 52 is in its Figure 4 The position above moves to its position in Figure 5The path of each pressing finger 56 at the position above is further marked by the shaded rectangle 48.

[0130] Therefore, each shaded rectangle 48 corresponds to all corresponding rectangles 46 and 47 along the entire path of the corresponding pressing finger 56. Thus, each rectangle 48 here has an extension dimension of approximately 1 mm in the front-to-back direction and an extension dimension of approximately 3 mm plus the length of the pressing finger path 56 in the left-to-right direction, the pressing finger path length being equal to the distance between the respective axes 38 of the leftmost and rightmost individual compartments 14, i.e., the two individual compartments 14 furthest apart from each other on the carrying object 40.

[0131] It should be noted that, in order for each pressing finger 56 to remain in full contact with the corresponding edge region 41 or 42 it travels through, regardless of which individual compartment 14 is aligned with the lens 51, the left-right extension dimension of that edge region must be at least equal to the left-right extension dimension of the shadow rectangle 48.

[0132] It should also be noted that the left-right extension dimension of the shaded rectangle 48 is actually greater than the distance between the corresponding axes 38 of the two most distant individual compartments 14 on the load 40 (because the left-right extension dimension of the pressing finger must be taken into account).

[0133] It should also be noted that, in Figure 7 In this configuration, the shaded rectangles 48 of the same edge region 41 or 42 do not intersect, allowing the covering to exist between the two rectangles 48 without being touched by the pressing finger 56 during the movement of the pressing finger. Therefore, in a variation not shown, the carrying object has two front edge regions and / or two rear edge regions as described above, but each edge region has a smaller extension dimension in the left-right direction, and the covering is positioned between the two front edge regions and / or the two rear edge regions.

[0134] Figure 13 The carrier 70 shown is similar to the carrier 40, except that the carrier 70 includes only two separate compartments 14, and in this case, the distance between the axes 38 of these compartments is equal to 15.84 mm.

[0135] Figure 8 This shows the loading / unloading configuration of the system 100 obtained after the stage 52 is separated from the pressure plate 53, so that the user can approach the carrier 40 to remove it from the stage 52, and optionally replace it with another similar carrier to analyze the sample contained in the compartment of that other carrier.

[0136] Conversely, in order to switch the system 100 from a loading / unloading configuration to an observation configuration, the stage 52 carrying the object 40 is moved closer to the pressure plate 53 until each pressing finger 56 contacts the edge regions 41 and 42 as previously described.

[0137] Now we will use Figures 9 to 12 A more detailed description of system 100 is provided.

[0138] The frame here includes a mounting wall 60, and the base 55 includes two arms 61, each arm projecting laterally from the mounting wall 60. The two arms 61 extend facing each other, defining an opening 58.

[0139] Here, each arm 61 is equipped with two pressing fingers 56, when the system 100 is in the observation configuration ( Figure 12 One of the pressing fingers 56 contacts the front edge region 41, and the other pressing finger contacts the rear edge region 42.

[0140] from Figure 10 As can be clearly seen, the magnets 54 of the clamping member are housed in the arms 61, and each arm 61 is provided with three cylindrical magnets 54. For each arm 61, the magnets 54 are distributed between the pressing fingers 56 of that arm 61.

[0141] Here, the pressure plate 53 has a guide groove 62 disposed in the base 55 on the same side as the pressing finger 56. Each guide groove 62 is configured to receive the end of a corresponding pin of one of the front positioning stop 33, the left positioning stop 34 and / or the right positioning stop 35 forming the stage 52 when the system 100 is in the observation configuration. When the stage 52 moves in the left and right direction, the end of the pin moves in the corresponding guide groove.

[0142] The guide groove 62 thus participates in the correct positioning of the stage 52 along the front-back direction of the object 40, so that when the stage 52 moves, the pressing finger 56 remains in contact only with the edge areas 41 and 42.

[0143] The stage 52 also has a ramp 63, and the pressure plate 53 has a protrusion 64. The protrusion is configured to engage with the ramp 63 during the lateral translational movement of the stage 52 along the object 40, so as to overcome the force applied by the clamping members to separate the stage 52 from the pressure plate 53, until the stage reaches... Figure 11 The location shown is where system 100 is in its loading / unloading configuration.

[0144] The stage 52 is mechanically connected to the frame via a sliding pivot connector, which includes a cylindrical guide 65 and a cylinder 66 mounted on the guide 65 to slide. When the object is received on the stage 52 in its predetermined position, the cylinder is longitudinally oriented in a direction parallel to the left-right direction of the object 40.

[0145] This mechanical connection allows the stage 52 to move further away from the pressure plate and closer to the pressure plate 53, and allows the stage 52 to move in the left and right directions.

[0146] The guide 65 is attached to the mounting wall 60, while the cylinder 66 is fixed to the stage 52.

[0147] The inner diameter of the cylinder 66 is slightly larger than the outer diameter of the guide 65, so that there is a gap between them, which allows the stage 52 to pivot to a certain extent in both directions on the left-right axis and in both directions on the front-back axis.

[0148] This pivoting capability of the stage 52 allows adjustment of the tilt of the upper surface 17 of the slide 11 relative to the pressing fingers 56 to ensure that all pressing fingers are in contact with the upper surface 17 and that the pressure is evenly distributed among all pressing fingers 56.

[0149] To move the stage 52 in the left-right direction, the microscope 50 also includes an electric mechanism (not shown) controlled from the user interface. In one variant, this mechanism is not electric but manually actuated by a knob.

[0150] The system 100 described above is particularly suitable for an analysis that includes the step of counting sperm present in an animal semen sample to determine its concentration.

[0151] In a variant not shown, system 100 is coupled to a computer configured to automatically analyze the images captured by the lens in order to perform a counting step and determine the concentration.

[0152] In a variant not shown, the focal point of the lens is not in the center of the individual compartment, but is located at a predetermined distance from the center of the individual compartment along the downward-upward direction.

[0153] In a variant not shown, the lens is mounted so that it can move relative to the frame of the microscope between a plurality of fixed, predetermined positions, in each of the plurality of predetermined positions the lens is located at a different corresponding distance from the object being carried.

[0154] Of course, in this application, the term "lens" must be understood in a broad sense, and can specifically refer to simple electronic image sensors, such as CMOS sensors, which are used, for example, in so-called "lensless" microscopes employing the principle of in-line holography. Thus, in one variant, the aforementioned analytical system is equipped with a camera equipped with such an electronic sensor, which is configured to record and process a series of images to analyze biological material, for example, to determine the motility of sperm contained in an animal semen sample.

[0155] In another variation not shown, the front edge strip and the rear edge strip are located at a short distance from the front surface 21 or the rear surface 22, respectively, i.e., for example, a cover or a portion of a cover is located on the upper surface 17 between the front surface 21 and the front edge region and / or between the rear surface 22 and the rear edge region.

[0156] In another variation not shown, the pressure plate includes three pressing fingers, two of which contact only one of the front and rear edge regions when the system is in the holding configuration, and the last pressing finger contacts only the other of the front and rear edge regions.

[0157] In another variation, not shown, the clamping member includes a magnet fixed to the stage, while the pressure plate is magnetic.

[0158] In another variation, not shown, the positioning stop differs from the pin protruding from the upper surface 37 of the stage 52. The positioning stop is formed, for example, by the edge of a recess provided in the upper surface 37, which is configured to accommodate the glass slide 11.

[0159] In other variations not shown, the recesses and vents are arranged in a manner different from that described above. For example, all recesses face the front surface and all vents face the rear surface, or from one compartment to another, the recesses and vents alternately face the front surface and then the rear surface.

[0160] In other variants not shown:

[0161] - The lens has a variable focal length instead of a fixed focal length;

[0162] -The number of individual compartments is different from four and two, for example, three, five or more;

[0163] - The clamping member includes a number of magnets different from the three magnets in each arm, such as one, two, four or more magnets per arm, and / or the magnets have a shape different from a cylinder, such as a parallelepiped;

[0164] - The clamping member differs from the magnet housed in the arm of the pressure plate, for example including a spring disposed on the stage side facing away from the pressure plate, the spring being configured to push the stage toward the pressure plate; and / or

[0165] - The inclined platform 63 is supported by the pressure plate, and the protrusion 64 is supported by the platform.

[0166] Many other variations are possible depending on the circumstances, and therefore it is stated that the invention is not limited to the embodiments described and shown.

Claims

1. A carrier for analyzing liquid-based biological material by microscopy, the carrier comprising a slide (11), a coverslip (12), and an adhesive thread (13) disposed between the slide (11) and the coverslip (12) to secure the slide and coverslip to each other, while defining a separate compartment (14) for receiving a sample of biological material on the periphery; the slide (11) is a parallelepiped having two main surfaces, namely an upper surface (17) and a lower surface (18), and side surfaces extending from one of the two main surfaces to the other, namely a left surface (19), a right surface (20), a front surface (21), and a back surface (22). The distance between the left surface (19) and the right surface (20) is greater than the distance between the front surface (21) and the rear surface (22), and the distance between the front surface and the rear surface is itself greater than the distance between the lower surface (18) and the upper surface (17); the carrying object (40) has the following characteristics: in the left-right direction, it is transverse to the left surface (19) and the right surface (20), and parallel to the front surface (21), the rear surface (22) and the two main surfaces; in the front-back direction, it is transverse to the front surface (21) and the rear surface (22), and parallel to the left surface (19), the right surface (20) and the two main surfaces; in the up-down direction, it is transverse to the two main surfaces and parallel to the side surfaces; the cover glass (12) is flat. The coverslip is a hexahedron with two main surfaces, namely the upper surface (23) and the lower surface (24), and side surfaces extending from one of the two main surfaces of the coverslip to the other, namely the left surface (25), the right surface (26), the front surface (27), and the back surface (28); the lower surface (24) of the coverslip (12) faces the upper surface (17) of the slide (11), while the front surface (27), the back surface (28), the left surface (25), and the right surface (26) of the coverslip (12) face the front surface (21), the back surface (22), the left surface (19), and the right surface (20) of the slide (11) respectively and are separated from them by a distance; each single Each individual compartment (14) is defined in the upper direction by the upper surface (17) of a glass slide (11) and the lower surface (24) of a coverslip (12), and is defined peripherally by an adhesive line (13). Each individual compartment (14) has an opening to a groove (15) on only one side and an opening to a vent (16) on the other side. Each groove (15) and each vent (16) are defined by an adhesive line (13). Each groove (15) faces one of the front surface (21) and the rear surface (22) of the glass slide (11), and each vent (16) faces the other of the front surface (21) and the rear surface (22) of the glass slide (11). The characteristic feature is that... The upper surface (17) of the slide (11) is bare and uncovered on at least one front edge area (41) extending along the front surface (21) of the slide (11) and on at least one rear edge area (42) extending along the rear surface (22) of the slide (11), each of the front edge area (41) and the rear edge area (42) having: an extension in the front-rear direction at least equal to 2.5 mm; and an extension in the left-right direction at least equal to the distance between the respective axes (38) of the two separate compartments (14) farthest from each other and not greater than the distance between the left surface (19) and the right surface (20) of the slide.

2. The object of claim 1, wherein For at least one of the front edge area (41) and the rear edge area (42), the extension in the left-right direction is equal to the distance between the left surface (19) and the right surface (20) of the slide (11).

3. The object of claim 1 or 2, wherein Each recess (15) has an extension in the front-rear direction smaller than the extension in the front-rear direction of each of the front edge area (41) and the rear edge area (42).

4. The object of claim 1 or 2, wherein The distance between the left surface (19) and the right surface (20) of the slide (11) is between 74.8 mm and 75.2 mm, the distance between the front surface (21) and the rear surface (22) of the slide (11) is between 24.8 mm and 25.2 mm, the distance between the left surface (25) and the right surface (26) of the cover glass (12) is between 31.8 mm and 32.2 mm, the distance between the front surface (27) and the rear surface (28) of the cover glass (12) is between 16.8 mm and 16.2 mm.

5. The object of claim 1 or 2, wherein All the recesses (15) are directed towards one of the front surface (21) and the rear surface (22) of the slide (11) and all the vents (16) are directed towards the other one of the front surface (21) and the rear surface (22) of the slide (11).

6. A system for analyzing liquid-based biological matter, the system comprising a carrier object (40) according to any one of claims 1 to 5, further comprising a microscope (50), the microscope comprising: a lens (51); a stage (52) configured to receive the object carrier (40) in a first observation position in which the lower surface (18) of the slide (11) rests against the stage (52) and one of the separate compartments (14) is aligned with the lens (51); a frame on which the lens (51) is mounted in at least one fixed predetermined position and on which the stage (52) is mounted so as to be able to move in translation in the left-right direction of the object carrier (40) to be able to successively align the other separate compartments (14) with the lens (51); a presser plate (53) extending at least partially opposite the object carrier (40); and clamping means configured to urge the platen (53) and the object table (52) towards each other so that the object (40) is clamped between the platen (53) and the object table (52); the platen (53) is fixedly mounted on the frame and comprises a base (55) and pressing fingers (56) protruding from the base (55) towards the object table (52) and whose distal ends (57) are in contact only with the front edge region (41) and the rear edge region (42), wherein at least two pressing fingers (56) are in contact on one of the front edge region (41) and the rear edge region (42) and at least one pressing finger (56) is in contact on the other of the front edge region (41) and the rear edge region (42); the object table (52) has positioning stops (34, 35) for fixing the object (40) immovably relative to the object table (52) in the left-right direction.

7. The system of claim 6, wherein, The platen (53) comprises four pressing fingers (56), of which two are in contact with the front edge region (41) and two are in contact with the rear edge region (42).

8. The system of claim 6 or 7, wherein, One of the object table (52) and the platen (53) is magnetic and the clamping means comprise a magnet (54) fixed to the other of the object table (52) and the platen (53).

9. The system of claim 6 or 7, wherein, The base (55) has an opening (58) through which the lens (51) is directed towards the individual compartment (14) aligned with the lens (51) in the first viewing position, at least two of the pressing fingers (56) in contact with the same edge region being arranged on either side of the opening (58).

10. The system of claim 9, wherein, The frame comprises a mounting wall (60), the base (55) comprises two arms (61) each protruding transversely from the mounting wall (60), the two arms extending facing each other and defining said opening (58), each arm (61) being provided with two pressing fingers (56), of which one is in contact with the front edge region (41) and one is in contact with the rear edge region (42).

11. The system of claim 10, wherein, The clamping means comprise a magnet (54) housed in the arm (61).

12. The system of claim 6 or 7, wherein, One of the object table (52) and the platen (53) has a ramp (63) and the other of the object table (52) and the platen (53) has a protrusion (64) configured to cooperate with the ramp (63) during translational movement of the object table (52) in the left-right direction in order to separate the object table (52) from the platen (53) against the force exerted by the clamping means to a position in which the object (40) is accessible for removal from the object table (52).

13. The system of claim 6 or 7, wherein, The object table (52) is mechanically connected to the frame by a sliding pivot connection comprising a cylindrical guide (65) oriented longitudinally in the left-right direction and a barrel (66) mounted to slide on the guide (65), there being a gap between the barrel (66) and the guide (65).

14. The system of claim 6 or 7, wherein, The lens (51) has a fixed focal length.

15. Use of a system according to any one of claims 6 to 14, characterized in that, The liquid-based biological substance is animal semen and the analysis comprises a sperm count step. The platen (53) comprises four pressing fingers (56), of which two are in contact with the front edge region (41) and two are in contact with the rear edge region (42). One of the object table (52) and the platen (53) is magnetic and the clamping means comprise a magnet (54) fixed to the other of the object table (52) and the platen (53). The base (55) has an opening (58) through which the lens (51) is directed towards the individual compartment (14) aligned with the lens (51) in the first viewing position, at least two of the pressing fingers (56) in contact with the same edge region being arranged on either side of the opening (58). The frame comprises a mounting wall (60), the base (55) comprises two arms (61) each protruding transversely from the mounting wall (60), the two arms extending facing each other and defining said opening (58), each arm (61) being provided with two pressing fingers (56), of which one is in contact with the front edge region (41) and one is in contact with the rear edge region (42). The clamping means comprise a magnet (54) housed in the arm (61). One of the object table (52) and the platen (53) has a ramp (63) and the other of the object table (52) and the platen (53) has a protrusion (64) configured to cooperate with the ramp (63) during translational movement of the object table (52) in the left-right direction in order to separate the object table (52) from the platen (53) against the force exerted by the clamping means to a position in which the object (40) is accessible for removal from the object table (52). The object table (52) is mechanically connected to the frame by a sliding pivot connection comprising a cylindrical guide (65) oriented longitudinally in the left-right direction and a barrel (66) mounted to slide on the guide (65), there being a gap between the barrel (66) and the guide (65). The lens (51) has a fixed focal length. The liquid-based biological substance is animal semen and the analysis comprises a sperm count step.

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