Equipment and method for preparation of leaflets for bioprosthetic heart valves

Abstract

The equipment (100) for preparation of leaflets (L) for bioprosthetic heart valves (10) comprises a station (200) for digitization of patches (P) of animal tissue including 2D optical sensors (225, 235) for imaging the patch (P) with at least one of bright-field epi- illumination and transillumination means (220, 230) for at least one of detection defects and assessing a collagen density in the patch (P), 3D optical sensors (240) for obtaining a thickness map (250) of patch (P), and a small-angle light scattering (SALS) system (300) for obtaining a through thickness orientation and dispersion map (255) of collagen fibers in patch (P). A station (400) for automatic selection of optimal regions of patch (P) to form accepted leaflets (L) based on a leaflet model (L'), a leaflet cutting station (500), a station (600) for final inspection, acceptance, and certification of cut leaflets (L) and a grouping station (700) are also provided.

Description

[0001] Equipment and method for preparation of leaflets for bioprosthetic heart valves

[0002] The present disclosure relates to bioprosthetic heart valves. More specifically, the present disclosure relates to equipment for preparation of leaflets for the manufacture of bioprosthetic heart valves. The present disclosure further relates to a method for preparation of leaflets for bioprosthetic heart valves using said equipment.

[0003] BACKGROUND

[0004] Biological or bioprosthetic heart valves (BHVs) are currently widely used over mechanical heart valves. Mechanical heart valves are made from metallic alloys or plastic materials. Due to thrombogenicity, mechanical heart valves require anticoagulation therapies which involve bleeding risks.

[0005] Bioprosthetic heart valves (BHVs) may be divided into transcatheter aortic valves (TAVs) and surgical aortic valves (SAVs).

[0006] Surgical and transcatheter implant of prosthetic heart valves in patients with valvular heart diseases (VHD) has significantly increased over the last decades. One example of implant of prosthetic heart valve is Aortic Valve Replacement (AVR). It is a common intervention in which a native heart valve is replaced with a prosthetic heart valve.

[0007] Although SAVs have been usually applied in the treatment of aortic valve diseases, transcatheter aortic valve replacement, also known as percutaneous transcatheter aortic valve implant (TAVI), has become the treatment of choice for inoperable patients with severe aortic valve stenosis.

[0008] BHVs typically comprise valve elements, referred to as leaflets or cusps, assembled into a flexible frame of the BHV. The flexible frame allows the leaflets to work together replicating a native valve.

[0009] Said leaflets are usually made from animal tissue, such as bovine or porcine pericardium. Animal tissue includes collagen fibers that reinforce the leaflets providing structural integrity thereto. This makes them to be the best option for manufacturing BHVs. Although BHVs are being preferred over mechanical heart valves, when the BHVs are implanted, the leaflets undergo cyclic mechanical stresses due cyclic opening and closure of the valve. Said stresses lead to failures since the mechanical response of the leaflet tissue greatly depends on collagen fiber density, dispersion and orientation, as reported by S. Hamed Alavi et al. "Characterizing the collagen fiber orientation in pericardial leaflets under mechanical loading conditions", Ann Biomed Eng. 2013 Mar; 41(3): 547-61.

[0010] It has been found that, if a given orientation of the collagen fibers with respect to the flow is selected in the manufacture of BHVs, a good tissue response to flow-related shear stresses can be obtained. However, the leaflets inevitably degenerate, which may result in valve dysfunction. As a result, an average lifespan of BHVs is estimated at fifteen years in elderly patients. This is the reason why they are currently of choice for most patients over the age of 65.

[0011] In this respect, prior art BHVs have been focused to increasing durability. Attempts towards developing durable BHVs have thus been made, such as for example in US6245105B1 and US6553681B2. Their approach is selecting leaflets for producing multi-leaflet bioprosthetic heart valves to insure maximum functional compatibility. In WO 2013 / 003842 A1 , a method is disclosed for assessing and cutting a bioprosthetic tissue.

[0012] In known prior art devices and methods for processing leaflets for BHVs, the thickness of pericardium tissues is measured, and useful areas that will become leaflets are selected. This however is carried out in inefficient ways which renders the processes not sufficiently effective for making durable heart valves.

[0013] A need still exists for more efficient devices and methods for producing leaflets for making durable bioprosthetic heart valves (BHVs).

[0014] SUMMARY

[0015] In view of the above needs, the present disclosure aims to provide an equipment for efficient preparation of leaflets for the manufacture of bioprosthetic heart valves (BHVs). A method for preparation of leaflets for bioprosthetic heart valves using said equipment is also disclosed herein. As described later, through the method that will be described below, the present equipment is capable of efficiently analyzing animal pericardium tissue, automatically selecting optimal regions in patches of said analyzed tissue, and automatically cutting said selected regions to be used as leaflets for the manufacture of BHVs, with a final quality control, certification, and grouping of the cut leaflets.

[0016] To clarify some of the terms used throughout the present description and claims, the following definitions are given hereinbelow.

[0017] The term “leaflet” as used throughout the present description is to be understood as each of the elements or sections, also referred to as cusps, that form a bioprosthetic heart valve and provide its main opening and closing mechanism. The aortic valve normally has three leaflets.

[0018] The term “patch” refers herein to a piece of tissue, for example pieces of the pericardia, that have been cut to size for making the leaflets of a heart valve.

[0019] Within the context of the present disclosure, "tissue" refers to any animal material comprising a stack of layers of collagen fibers.

[0020] The term “automatically” is to be interpreted in the present disclosure as any action or step that is carried out with little or no direct human control.

[0021] Also within the meaning of the present disclosure, "preparation of leaflets" refers to arranging leaflets through method steps that will be described hereinbelow so that they are ready for making bioprosthetic heart valves.

[0022] As used herein "inspecting leaflets" refers to optically inspecting leaflets.

[0023] Within the meaning of the present disclosure, "through thickness dispersion and orientation" define an architecture of collagen fibers in a patch or in leaflet, that is, the average dispersion and dominant orientation of collagen fibers through all the layers of a patch or a leaflet.

[0024] The term “dispersion” in the present disclosure refers to a variation of an orientation of collagen fibers at the different layers of the pericardial tissue. The term "orientation" in the present disclosure refers to a direction in which the collagen fibers are arranged, and "dominant orientation" refers to a direction around which most of the collagen fibers are arranged in the overall thickness of the tissue.

[0025] As used herein, “fiber size” refers to a diameter of collagen fibers. Also, "interfiber spacing" refers herein to spaces between collagen fibers defining a separation there between.

[0026] Within the context of the present disclosure, “sensor” or “optical sensor” is understood to include at least one sensor camera, a corresponding sensor optics, and a suitable illumination means.

[0027] In a first aspect, an equipment for preparation of leaflets for bioprosthetic heart valves is provided. The present equipment is intended to process patches from animal pericardium tissue having maximum size of 120 x 120 mm. However, other values may be also envisaged as required.

[0028] The present equipment comprises a digitization station for digitization of at least one patch of animal pericardium tissue.

[0029] Said station for digitization of at least one patch of animal pericardium tissue includes at least one 2D optical sensor. Said 2D optical sensor is suitable for imaging a patch with at least one of bright-field epi-illumination means and transillumination means for at least one of detection of defects in the patch and assessing a collagen density in the patch. For assessing collagen density in the patch, the 2D optical sensor may include polarized light transillumination means.

[0030] The station for digitization of at least one patch of animal pericardium tissue further includes at least one 3D optical sensor. Said 3D optical sensor is suitable for obtaining a thickness map of the patch. For this purpose, the 3D optical sensor may be one of dynamic fringe projection, confocal, interferometric, and focus variation. In further examples, the 3D optical sensor may include a 5 megapixel camera with a lateral resolution better than 15 microns ( .m) and a vertical resolution better than 1 .m.

[0031] Within the context of the present disclosure and as it will be clear to those experienced in the art, a better lateral resolution of the optical sensor means a smaller distance resolved on the surface of the tissue. Thus, in the example above, a resolution better than 15 .m means a resolution having a value lower than 15 .m. Within the context of the present disclosure and as it will be clear to those experienced in the art, a better vertical resolution means a smaller size of the distance resolved on the direction perpendicular to the surface of the tissue. Thus, in the example above, a vertical resolution better than 1 .m means a resolution having a value lower than 1 .m.

[0032] The station for digitization of at least one patch of animal pericardium tissue further includes a small-angle light scattering (SALS) sensor. The present SALS sensor has been improved for obtaining a through thickness dispersion and orientation map of collagen fibers in the patch.

[0033] Specifically, the present SALS sensor has been improved to allow for rapid tissue analysis of features at a micron scale allowing fiber dispersion and orientation of collagen fibers to be rapidly characterized to provide an architectural mapping of collagen fibers, avoiding extensive manual intervention

[0034] The present SALS sensor provides an accurate method for rapid quantification of collagen fibers through the use of a laser with a wavelength within an order of magnitude of the fiber size and the interfiber spacing, such as a He-Ne laser with a wavelength of 632.8 nm, as reported by M. S. Sacks et al., “A small angle light scattering device for planar connective tissue microstructural analysis", Annals of Biomedical Engineering, Vol. 25, pp. 678-589, 1997. The present SALS sensor is also highly compact as compared with standard known SALS sensors in terms of short distance between the projection screen and the patch while maintaining the far- field diffraction condition. The present SALS sensor provides high-resolution information on local collagen fiber dispersion and orientation avoiding the need for expensive equipment and time-consuming tissue preparations while allowing a rapid analysis of large areas of tissue.

[0035] When a laser beam passes through the thickness of the pericardium tissue a characteristic image of the far-field diffraction pattern is obtained which corresponds to an ellipse. More in detail, the improved SALS sensor allows a far-field diffraction pattern to be obtained where diffraction patterns generated in each of the respective layers of collagen fibers in the pericardium tissue crossed by a focused laser beam with the above mentioned wavelength are superimposed. Such far-field diffraction pattern that is obtained by the improved SALS sensor is an ellipse. Said ellipse is characterised by an eccentricity value and an orientation of the major axis and the minor axis of said ellipse. The through thickness orientation of collagen fibers is coincident with the orientation of the minor axis of the ellipse, and the through thickness dispersion of collagen fibers is assessed from the eccentricity value of the ellipse.

[0036] It was found that the results of the eccentricity and the orientation of the major axis of the ellipse obtained with the improved SALS sensor are consistent with the architecture of the fibres observed through a second-harmonic generation (SHG) microscope.

[0037] The eccentricity of the ellipse is related to the dispersion of the collagen fibres of the patch. The more the fibres are arranged in the same orientation in the different layers of the patch, the sharper the aggregate diffraction pattern and the higher the eccentricity of the ellipse. A further relevant parameter from the diffraction pattern is the angle of the major axis of the ellipse which corresponds to a direction perpendicular to the direction of the dominant orientation of the fibres in the overall thickness of the tissue of the patch. These parameters obtained from the improved SALS sensor will allow the optimal selection of the angular orientation of the leaflets in a patch.

[0038] The present equipment further comprises a selection station. The selection station is configured for automatic selection of regions of the patch considered to be optimal to form accepted leaflets based on digital results obtained by the above mentioned digitization station and according to at least one leaflet model.

[0039] The equipment may further include a cutting station for cutting selected optimal regions of the patch to form leaflets. Said cutting station may comprise a laser cutting device suitable for cutting selected optimal regions of a patch as sated above. Preferably, the laser cutting device is a CO2 laser cutting device. However, other different configurations are not ruled out, such as for example, those in which the cutting station comprises a die cutting device. Other suitable cutting devices for cutting selected optimal regions of a patch to form leaflets are not ruled out.

[0040] The station for digitization of patches described above can also be configured for inspection of leaflets if required. However, it may be preferred that a separate leaflet inspection station is provided. Thus, in this latter case, the equipment may include, in addition to said station for digitization of patches, a station for inspection of leaflets.

[0041] Said inspection station is configured for performing dimensional verifications in the leaflet, measuring a thickness map of the leaflet, detecting critical defects in the leaflet, assessing a collagen density in the leaflet, and measuring through thickness dispersion and orientation of collagen fibers in the leaflet, and considering a leaflet as accepted or rejected when compared with a leaflet model.

[0042] A grouping station may be included in the present equipment. If provided, the grouping station allows for arranging accepted leaflets into groups of at least two leaflets to be incorporated into a BHV. For example, in the case of an aortic BHV the accepted leaflets will be grouped into groups of three leaflets.

[0043] Feeding of patches or leaflets from one station to another in the present equipment may be performed automatically, through suitable feeding means for automatically feeding patches or leaflets through the stations. The feeding means may include robots. Alternatively, feeding of patches or leaflets from one station to another may be performed manually by an operator. A combined operation of the present equipment both automatically and by an operator may be possible.

[0044] The present equipment may further include support means for the pericardium tissue, that is, for supporting patches or leaflets. Said support means behave as an optically diffusing surface on a dry condition and also as an optically transparent surface on a wet condition that is when in contact with a hydrated patch or leaflet.

[0045] It may be preferred that the above mentioned support means for the pericardium tissue comprise a plane-parallel glass plate. Such plane-parallel glass plate has an upper textured side and a lower polished side. The upper side of the plane-parallel glass plate is intended to receive a hydrated pericardium tissue of a patch or a leaflet.

[0046] In a further aspect of the present disclosure, a method for preparation of leaflets for BHVs through an equipment as described above is also provided herein.

[0047] The present method for preparation of leaflets for BHVs includes loading a patch of animal pericardium tissue in the digitization station described above for digitization of at least one patch of animal pericardium tissue. Through the present method, at least one of 2D bright-field image and a 2D transillumination image of the patch is obtained. A 3D thickness map of the patch and a through thickness orientation and dispersion map of collagen fibers in the patch are also obtained.

[0048] Regions of the patch that are considered optimal to form accepted leaflets are selected according to at least one leaflet model. Selecting one or more optimal regions in a patch is based on digital results obtained from the above mentioned digitization station that is intended for digitization of at least one patch of animal pericardium tissue. Selection is carried out by applying an optimization process based on a mathematical optimization technique. Optimally, the mathematical optimization technique is a metaheuristic simulated annealing probabilistic technique.

[0049] The above mentioned selecting step in which one or more regions of a patch considered optimal to form accepted leaflets according to a leaflet model are selected may involve at least one of two additional optimization variables resulting from a mean tissue thickness at commissure regions being above an average value of the tissue thickness of the leaflet and a collagen density at commissure regions being above an average value of the collagen density of the leaflet.

[0050] Selected regions of the patch considered optimal to form accepted leaflets are cut in a cutting step according to the present method. Said cutting step may further include forming holes in the leaflet for subsequent sewing of the leaflet to a frame in a BHV. The frame of the BHV is preferably configured as a metal structure.

[0051] The selecting step may further comprise excluding regions of the patch having critical defects or low density of collagen fibers. Said selecting step may further include determining the best positions for a leaflet that meet the through thickness dispersion and orientation of the collagen fibers and thickness parameters that have been previously defined in a leaflet model. Critical defects in regions to be excluded are detected from at least one of 2D bright-field and 2D transillumination images obtained from the above mentioned 2D optical sensors in the digitization station.

[0052] The present method for preparation of leaflets for BHVs may further comprise an inspection step wherein leaflets are inspected. Such inspection step involves performing dimensional verifications in the leaflet, measuring a thickness map of the leaflet, detecting critical defects in the leaflet, assessing collagen density, and measuring through thickness dispersion and orientation of collagen fibers in the leaflet. Finally, in the inspection step, a leaflet is considered as accepted or rejected when compared with a leaflet model. Accepted leaflets are arranged into groups of at least two leaflets to be incorporated into a BHV. If required, the method may further comprise rehydration of leaflets that have been cut before the inspection step.

[0053] As mentioned above, the present equipment and method involve digitization of patches, and automatic selection and cutting of leaflets, as well as inspection, validation, certification and grouping of leaflets for the manufacture of BHVs. As a result, an increase of quality and productivity of the final product is advantageously obtained. Since the present equipment provides automation for inspection and manufacturing of leaflets for making BHVs, it has been found that efficiency, durability, and quality are advantageously increased while costs are reduced. Improvements in terms of traceability during the manufacturing process are also achieved. All of this represents great benefits for patients requiring implantation of BHVs.

[0054] BRIEF DESCRIPTION OF THE DRAWINGS

[0055] An exemplary detailed description is given below intended to provide further explanation of the present disclosure. Other advantages and features will be apparent from the following description and drawings.

[0056] In the drawings:

[0057] Figure 1a shows a top plan view of a Sapien® bioprosthetic heart valve with the leaflets in closed position (diastole);

[0058] Figure 1b shows a top plan view of the Sapien® bioprosthetic heart valve shown in figure 1a with the leaflets in open position (systole);

[0059] Figure 1c shows an elevational view of the Sapien® bioprosthetic heart valve shown in figures 1a and 1b;

[0060] Figure 2 is a top plan view of a leaflet ready to be assembled into a bioprosthetic heart valve; Figure 3 is a block diagram diagrammatically showing stations of the present equipment for preparation of leaflets for bioprosthetic heart valves;

[0061] Figure 4 is a block diagram diagrammatically showing the station for digitization of patches of animal pericardial tissue of the equipment shown in figure 3;

[0062] Figure 5 is a block diagram diagrammatically showing the station for automatic selection of optimal regions of the patch of the equipment shown in figure 3;

[0063] Figure 6 is a block diagram diagrammatically showing the station for inspection and acceptance or rejection of leaflets of the equipment shown in figure 3;

[0064] Figure 7 is a schematic drawing of one example of a 2D optical sensor for imaging a patch or a leaflet with epi-illumination means;

[0065] Figure 8 is a schematic drawing of one example of the 2D optical sensor for imaging a patch or a leaflet with transillumination means;

[0066] Figure 9 is a schematic drawing of one example of a dynamic fringe projection 3D optical sensor for obtaining a thickness map of a patch or a leaflet;

[0067] Figure 10 is a block diagram diagrammatically showing the Small Angle Light Scattering (SALS) system of the equipment shown in figure 3; and

[0068] Figure 11 is a graph showing an ellipse corresponding to a far-field diffraction pattern as displayed in a screen when a laser beam of the improved SALS sensor is passed through the thickness of a pericardium tissue of a patch or a leaflet, and also showing the orientation of the major axis of the ellipse E with respect to a horizontal reference axis X.

[0069] DETAILED DESCRIPTION OF ONE EXAMPLE

[0070] Figures 3-11 of the drawings show one non-limiting example of the present equipment. In the drawings, the equipment is denoted, in general, by reference sign The equipment 100, that will be described in detail below, is intended for preparation of leaflets L for bioprosthetic heart valves 10. Bioprosthetic heart valves 10 are referred hereinafter to as BHVs 10. As it will be described in detail herein below, the present equipment 100 allows for the automatic manufacture and certification of leaflets L for BHVs 10 that meets the demands of heart valve manufacturers. For this purpose, the equipment 100 provides for digitization, automatic selection, cutting of leaflets, and validation, certification, and grouping of leaflets L for manufacturing BHVs 10.

[0071] The leaflets L, as shown in detail in figure 2 and also in figures 1a, 1b, and 1c of the drawings, are valve elements in a BHV 10. Each BHV 10 comprises a flexible metal frame 15 to which leaflets L, three in the BHVs 10 in figures 1a, 1b, 1c, are attached such that they are allowed to work together replicating a native valve.

[0072] Leaflets L are made from animal tissue, for example, bovine or porcine pericardium, which includes collagen fibers. Collagen fibers reinforce the leaflets L providing structural integrity thereto.

[0073] As shown in figure 2, each leaflet L comprises side taps or commissures 20, a coapting edge 21, and an arcuate cusp edge 22.

[0074] The equipment 100 for preparation of leaflets for BHVs 10 is of a modular and flexible nature and it is based on optical sensors 225, 235, 240 and 300, that will be described in detail further below.

[0075] Referring to figure 3 and figure 4 of the drawings, the equipment 100 comprises a digitization station 200 for digitization of at least one patch P.

[0076] As used herein, a patch P refers to a piece of animal pericardium tissue having a stack of layers of collagen fibers.

[0077] The digitization station 200 is configured for digitization of at least one patch P. The patch P to be digitalized is placed onto the upper side 951 of a plane-parallel glass plate 950 adapted to be driven through a XY motor, not shown in the drawings. Said glass plate 950 is part of support means 900 that will be described further below.

[0078] More in particular, an analysis of patches P is performed in said digitization station 200. From said analysis, optimal regions of the patch P are automatically selected which can be converted into leaflets L for manufacturing BHVs 10.

[0079] In order to perform said analysis of the patches P, the digitization station 200 includes 2D optical sensors 225, 235 as shown in figures 4, 7, and 8. The 2D optical sensors 225, 235 comprise respective sensor cameras 221, 231 and sensor optics 215, as shown in figures 7 and 8 of the drawings. The 2D optical sensor 225 also includes bright-field epi-illumination means 220, as shown in figure 7, and the 2D optical sensor 235 also includes transillumination means 230, as shown in figure 8 for imaging the patch P. The 2D optical sensor 225 is configured for obtaining a 2D bright-field image 226 of the patch P. The 2D optical sensor 235 is configured for obtaining at least one 2D transillumination image 236 of the patch P for detection of defects and assessing collagen density in the patch P. For assessing collagen density, the 2D optical sensor may include polarized light transillumination means.

[0080] Referring to figure 9, the digitization station 200 further includes a 3D optical sensor 240, as shown in figures 4 and 9. The 3D optical sensor 240 comprises a sensor camera 241 and sensor optics 245, as shown in figure 9. The 3D optical sensor 240 also includes 3D illumination means 246, as shown in figure 9. The 3D illumination means 246 is, in particular, a fringe projection illumination means 246 configured for illuminating said patch P placed onto the upper side 951 of the above mentioned plane-parallel glass plate 950 of the support means 900.

[0081] The 3D optical sensor 240 of the digitization station 200 is configured for obtaining a high resolution thickness map 250 of the patch P. To this effect, the 3D optical sensor 240 may be a dynamic fringe projection optical sensor as shown in the example in figure 9, although the optical sensor 240 may be other types of optical sensor such as confocal optical sensor, interferometric optical sensor, focus-variation optical sensor, etc. In the example shown in said figure 9, the sensor optics 245 of the 3D optical sensor 240 includes a telecentric optical system, a 5 megapixel camera 241, and the above mentioned fringe projection illumination means 246. As a result, a depth of focus greater than 1 mm, a lateral resolution better than 15 .m and a vertical resolution better than 1 .m are advantageously achieved. Such a depth of focus allows the measurement of the thickness map of pericardium tissues with thicknesses under 1 mm. Through the thickness map 250 of the patch P that is obtained through said 3D optical sensor 240 of the digitization station 200 a mean value and a standard deviation of the thickness of the tissue of the patch P can be determined. From said thickness map 250 of the patch P, it is possible to feed a selection algorithm that determines optimal positions of leaflets L in the patch P that meet the parameters set from a leaflet model L’. Reference will be made to said selection algorithm below.

[0082] The digitization station 200 includes a compact and automatable improved smallangle light scattering (SALS) system 300 as diagrammatically shown in figures 4 and 10 of the drawings. The improved SALS sensor 300 is an essential element for an automatic manufacturing process of leaflets L to be assembled into a BHV 10.

[0083] In the example shown in figure 10, the improved SALS sensor 300 includes a camera 301 , an imaging optics 302, a projection screen 303, a He-Ne laser source 304 suitable for emitting a laser beam 305 with a power of at least 5 mW, and a wavelength of 632.8 nm, a beam shaping optics 306, and a bending mirror 307. Such wavelength is within an order of magnitude of the fiber size and the interfiber spacing of collagen fibers in a pericardial tissue. The projected laser beam 305 is reflected by the bending mirror 307, after passing through the beam shaping optics 306. After passing through said bending mirror 307, the laser beam 305 passes through the plane parallel-glass plate 950 and through the patch P of pericardial tissue, and it is scattered by the fibrous structures thereof. A resultant far-field diffraction pattern is thus projected onto the above mentioned projection screen 303 and then imaged by the above mentioned imaging optics 302 onto said camera 301.

[0084] The above mentioned far-field diffraction pattern is obtained by the improved SALS sensor 300 as a result of the laser beam 305 passing through the patch P. This far- field diffraction pattern is the result of the superposition of diffraction patterns generated in each one of the layers of collagen fibers of the pericardial tissue that are traversed by the laser beam 305 as also reported by M. S. Sacks et al., “A small angle light scattering device for planar connective tissue microstructural analysis", Annals of Biomedical Engineering, Vol. 25, pp. 678-589, 1997. Said far-field diffraction pattern corresponds to an ellipse E that is projected in the screen 303 and imaged onto the camera 301 by the imaging optics 302, as shown in figure 11 of the drawings. From said ellipse E, through thickness orientation and dispersion of collagen fibers in the patch P are determined such that a through thickness orientation and dispersion map 255 of collagen fibers in the patch P is quickly and reliably obtained.

[0085] The ellipse E is characterized by a value of an eccentricity s, and an orientation of its major axis A and minor axis B. The value of the eccentricity s of the ellipse E refers to how round said ellipse E is. The greater the value of the eccentricity s the more flattened the ellipse E is, and the smaller the value or closer to zero the eccentricity s the more circular the ellipse E is. The eccentricity s ranges between one and zero and can be calculated as follows: wherein a and b are the respective lengths of the major axis A and the minor axis B of the ellipse E as shown in figure 11. The eccentricity s is a dimensionless parameter. Therefore, said lengths a and b can be expressed in Arbitrary Units (AU).

[0086] In the example shown in the image illustrated in figure 11 , the length a of the major axis A of the ellipse E is 371 AU, and the length b of the minor axis B of the ellipse E is 268 AU. The eccentricity s is thus in this case: _ __ 0.69

[0087] The through thickness dispersion of collagen fibers is assessed from the eccentricity s, which is obtained by means of the major axis A and the minor axis B of the ellipse E as stated above and as reported by A. Whelan et al. “Collagen fibre orientation and dispersion govern ultimate tensile strength, stiffness and the fatigue performance of bovine pericardium" where tissues with highly aligned fibers show an eccentricity s equal to or above 0.65, while tissues with highly dispersed fibers, i.e. fibres orientated in multiple directions, show an eccentricity s lower than 0.65. In general, the more the fibres of the patch P are arranged in the same direction in tissue layers thereof, the more flattened the aggregate diffraction pattern and the higher the eccentricity s of the ellipse E. In this case, an eccentricity value of s = 0.69 as in the example shown in figure 11, means that the patch P can be considered as a tissue with highly aligned fibers. A further relevant parameter obtained from the diffraction pattern is the angle of orientation a of the major axis A of the ellipse E. As shown in figure 11 , said angle of orientation a is defined by the major axis A of the ellipse E and a horizontal reference axis x. The orientation of said major axis A of the ellipse E corresponds to a direction perpendicular to that of the dominant orientation of the collagen fibres in the overall thickness of the tissue of the patch P. In the non-limiting example shown in figure 11 , the angle of orientation a of the major axis A of the ellipse E is a = 45°.

[0088] Through said two parameters, that is, the eccentricity s and the angle of orientation a of the major axis A of the ellipse E obtained from the SALS sensor 300, an optimal angular orientation of the leaflets L is determined. Furthermore, it has been found that the improved SALS sensor 300 in the digitization station 200 of the present equipment 100 is capable of fast and accurate measuring of fiber orientation in patches with thicknesses ranging from 100 .m to 1000 .m with an angular resolution better than 1 degree and a lateral resolution better than 500 .m.

[0089] With the 2D and 3D sensors 225, 235, 240 and the improved SALS sensor 300 included in the digitization station 200 of the equipment 100 as described above, necessary parameters for a good selection of leaflets L are quickly and reliably obtained. Therefore, an efficient non-contact inspection of pericardial tissue can be performed both for manufacturing of leaflets L and for performing final quality control and certification of leaflets L before sewing them to the metal frame 15 of the BHVs 10.

[0090] The present equipment 100 further comprises a selection station 400. The selection station 400 is configured for automatic selection of regions of the patch P that are considered optimal to form accepted leaflets L. Automatic selection of regions of patches P is based on digital results obtained by the digitization station 200 described above and according to at least one leaflet model L’.

[0091] Selection of optimal regions of a patch P to become leaflets L, that is, accepted leaflets L, is carried out through the use of suitable algorithms in a sequential approach consisting of the following steps. An initial step excluding areas in a patch P affected by critical defects or low collagen density that have been detected by the 2D transillumination images of the patch P is performed. A further step of the selection of optimal regions of a patch P involves optimal positioning of leaflets L from the thickness map obtained through the 3D optical sensor 240 and the optimal leaflet orientation obtained through the improved SALS sensor 300 according to at least one leaflet model L’ as previously described. This is carried out through an optimization algorithm of tissue thickness distribution according to the above mentioned 3D thickness map 250 of the patch P that has been obtained.

[0092] Selecting regions in the patch P considered optimal to form accepted leaflets L according to a leaflet model L’ involves at least two additional optimization variables. Said additional optimization variables result from a mean tissue thickness at commissure regions being above an average value of the tissue thickness of the leaflet L and a collagen density at commissure regions being above an average value of the collagen density of the leaflet L.

[0093] The present equipment 100 further comprises a cutting station 500 for cutting said selected optimal regions of the patch P to form leaflets L. In the non-limiting example shown in the drawings, the cutting station 500 comprises a laser cutting device 550. In this example, it is a CO2 laser cutting device 550. A low-power CO2 laser cutting device 550 capable of delivering a power of up to 10 W is preferred, in particular a VLS2.30 model laser cutting device 550 available from Universal Laser Systems Inc. including a beam focusing head with aspheric optics. Cutting of leaflets L is optimized while heat affected zones of tissue are advantageously minimized. However, in other examples, the cutting station 500 may comprise other types of cutting devices such as die cutting devices. In any case, the leaflets L are cut in the cutting station 500 at optimal positions according to information provided by the digitization station 200 and the selection station 400 as described above. Leaflets L are cut in the cutting station 500 lying on the above mentioned plane-parallel glass plate 950.

[0094] Contour lines of a patch P defining the boundaries for selected leaflets L are converted into a drawing file such as, for example, Portable Document Format (pdf), or into a vector file format, such as Corel Draw® format (cdr). Once said drawing files have been created, they are transferred to the laser cutting device 550 in order to perform the corresponding cutting operation to form the leaflets L. The information contained in said drawing files may further include locations corresponding to small holes to be made in the tissue of the patch P for subsequent sewing of the formed leaflets L to the metal frame 15 of the BHV 10. A number of operating parameters of the laser cutting device 550 may be set such as power and pulse frequency. They may be suitably adjusted to obtain the best qualityspeed ratio in the cutting operation of the pericardium tissue of a patch P. In particular, power and cutting speed of the laser cutting device 550 can be set for each leaflet L to be obtained since the thickness of the leaflets L to be cut is known.

[0095] As stated above, support means 900 are provided for supporting patches P or leaflets L of pericardium tissue ensuring an accurate positioning thereof. By using said support means 900, the patches P do not require to be handled and the leaflets L are obtained exactly at optimal positions.

[0096] Specifically, the support means 900 illustrated in the example shown in figures 7-10 comprise the above mentioned plane-parallel glass plate 950. Said plane-parallel glass plate 950 is 150 x 150 mm and includes an upper side 951 and a lower side 952. The upper side 951 of the plane-parallel glass plate 950 is slightly textured and is intended to receive a hydrated pericardium tissue of a patch P. The lower side 952 of the plane- parallel glass plate 950 is polished.

[0097] As a consequence of the above configuration, when a hydrated pericardium tissue of a patch P contacts the textured upper side 951 of the plane-parallel glass plate 950, said textured upper side 951 behaves as an optically transparent surface for the 2D optical sensor 235 with transillumination means 230.

[0098] One advantage of said plane-parallel glass plate 950 having a textured upper side 951 is that it completely absorbs the laser beam of the CO2 laser cutting device. As a result, reflections of the laser beam remaining after cutting the tissue are avoided, so that a good cutting quality in the cutting station 500 is achieved.

[0099] Referring to figure 6, the present equipment 100 further comprises an inspection station 600 for inspection of cut leaflets L. The leaflet inspection station 600 is configured for performing dimensional verifications, detecting critical defects in the leaflet L, assessing collagen density, and double checking a 3D thickness map of the leaflets L and the through-thickness orientation and dispersion of collagen fibers in the leaflets L through the above mentioned 2D optical sensors 225, 235 with bright-field epiillumination means 220, and transillumination means 230, respectively, the above mentioned 3D optical sensor 240 with 3D fringe projection illumination means 246, and the improved SALS sensor 300.

[0100] As a result of the optimal positioning of the leaflets L in the selection station 400, it is expected that the orientation of the dominant direction of the collagen fibers in the leaflet L is horizontal, that is, parallel to the coapting edge 21 of the leaflet L shown in figure 2.

[0101] The result of said operations in the leaflet inspection station 600 is the acceptance or rejection of leaflet L, according to at least one leaflet model L', minimizing risks and providing traceability. In case of acceptance of a leaflet L, said leaflet L may be certified, a reference may be assigned thereto, and a set of characteristic parameters is provided for suitably grouping leaflets L into sets of at least two units to be assembled into a BHV 10.

[0102] Final inspection, quality control and certification of leaflets L in said inspection station 600 are performed before leaflets L are grouped. Grouping of leaflets L is performed in a grouping station 700 that will be described below. The inspection station 600 may be left aside if the digitization station 200 described above is also configured for inspection of leaflets L.

[0103] The present equipment 100 further comprises a grouping station 700 as stated above. The grouping station 700 is configured for arranging accepted leaflets L into groups of at least two leaflets L, typically three, to be incorporated into a BHV 10. More in particular, accepted leaflets L are arranged in said grouping station 700 into sets of units, for example three units in a transcatheter aortic valve implant (TAVI), for being assembled into a metal frame 15 of a BHV 10 for optimal operation. Before grouping, certified leaflets L are appropriately stored and managed through a suitable software.

[0104] Feeding means 800 are also provided for feeding patches P or leaflets L through the stations of the equipment 100. The feeding means 800 are configured for transferring patches P or leaflets L onto the plane-parallel glass plate 950 from one station to the other. The transferring operation may be carried out manually, automatically, or combining manual and automatic transferring operations.

[0105] Manual transferring operations may comprise, for example, manually loading patches P or leaflets L on the plane parallel glass plate 950 of the support means 900. The present equipment 100 for preparation of leaflets L for bioprosthetic heart valves 10 preferably has a U-shaped cell layout. This is preferred over a linear layout for the transfer of patches P and leaflets L. U-shaped cell layout provides for a good scalability of productivity of the equipment 100 as well as a suitable integration of the cutting station 500. U-shaped cell layout can be operated flexibly by an operator or by a robot as required. Such preferred cell layout of the equipment 100 is advantageous in respect of accessing the laser cutting device 550 since currently available laser cutting devices operate in a closed cabin with no other access than a hinged top cover. With the present preferred U-shaped cell layout, a side access to a cutting platform is not required, so that a protective enclosure is not necessary to be removed or modified to gain access. With U-shaped cell layout the flow of patches P and leaflets L through stations is facilitated. Patches P come in at one end of the equipment 100, they are processed, that is, digitized, cut, etc., at different stations, moving around the ‘U’ space. At the other end, accepted and certified leaflets L leave the equipment 100. As a result, an optimal lean manufacturing process is obtained.

[0106] The present equipment 100 is suitably controlled by a computer program. Said computer program comprises instructions which, when executed by a computer, cause the computer to carry out the steps of the method that has been described above.

[0107] Said computer program includes software modules that include one or more of the following.

[0108] The computer program includes a digitization module for acquisition and analysis of pericardium tissue patches P, and selection of optimal regions to be converted into leaflets L. The digitization module operates from information provided by the 2D and 3D sensors 225, 235 and 240 depicted in figures 4,7-9, and also from information provided by an improved SALS sensor 300 depicted in figures 4 and 10 of the drawings.

[0109] The computer program further includes a simple leaflet cutting module for translating lines defining the boundaries of the selected leaflets L into computer information that is delivered in the above mentioned formats, that is, pdf or cdr formats, for example. Such information is sent through suitable means to the laser cutting device 550 of the cutting station 500 for cutting the patches P into leaflets L. A method for preparation of leaflets L for BHVs 10 using the equipment 100 as described above is disclosed herein with reference to figure 4 of the drawings.

[0110] Referring now back to figure 4 of the drawings, a patch P of an animal pericardium tissue is loaded 1000 in the digitization station 200 of the equipment 100. In step 1100, a 2D bright-field image 226 of the patch P is obtained through the 2D optical sensor 225. In step 1200, at least one 2D transillumination image 236 of the patch P is obtained through the 2D optical sensor 235. In step 1300, a 3D thickness map 250 of the patch P is obtained through the 3D optical sensor 240. In step 1400, a through thickness orientation and dispersion map 255 of collagen fibers in the patch P is obtained through the improved SALS sensor 300.

[0111] Regions of the patch P considered optimal to form accepted leaflets L are selected in step 1500. Selection of optimal regions of the patch P is performed in a selection station 400 based both on at least one leaflet model L’ and on digital results obtained from the digitization station 200. Selection is conducted by applying an optimization process based on a mathematical optimization technique. As shown in figure 5, as a result of applying an optimal positioning process based on the above mentioned leaflet model L’, optimal positions for potential accepted leaflets L are obtained.

[0112] Instead of algorithms such as gradient descent or branch-and-bound, said mathematical optimization technique used in the present method is a metaheuristic simulated annealing (SA) probabilistic technique. SA is a probabilistic technique using an algorithm for approximating the global optimum of a given function which is useful when finding an approximate global optimum is more important than finding a precise local optimum in a fixed amount of time. Specifically, SA performs an approximate global optimization in a large search space for an optimization problem.

[0113] The selection step 1500 further involves excluding regions of the patch P where critical defects or low collagen density have been found. Critical defects and low collagen density regions are detected from at least one of the above mentioned 2D bright-field and 2D transillumination images 226, 236 obtained through the above mentioned 2D optical sensors 225, 235, respectively.

[0114] Selection step 1500 further includes determining the best positions for a leaflet L that meet the through thickness orientation and dispersion of the collagen fibers and thickness parameters that have been previously defined in the leaflet model L’. This corresponds to positions of a leaflet L oriented according to a dominant direction of collagen fibers that has been identified from the through thickness orientation and dispersion map 255 obtained by the improved SALS sensor 300.

[0115] The SA probabilistic technique according to the present method involves at least one of two additional optimization variables resulting from a mean tissue thickness at side taps or commissures 20 of the leaflet L being above an average value of the tissue thickness of the leaflet L and a collagen density at side taps or commissures 20 of the leaflet L being above an average value of the collagen density of the leaflet L.

[0116] In step 1600, selected optimal regions of the patch P are cut by a laser cutting device 550 in a cutting station 500 to form leaflets L. The cutting step 1600 also involves forming holes in accepted leaflets L for subsequent sewing to the BHV frame 15. Cut leaflets are rehydrated before being transferred to a final inspection in step 1700.

[0117] In step 1700, leaflets L are inspected in an inspection station 600. Inspection involves performing dimensional verifications in leaflets L, detecting critical defects in the leaflet L, assessing collagen fiber density, measuring a leaflet thickness map 250, and measuring through thickness orientation and dispersion of collagen fibers in the leaflets L. Inspection further involves considering a leaflet L as accepted or rejected when compared with a leaflet model L’. Inspected and accepted leaflets L are then certified.

[0118] In step 1800, certified leaflets L are arranged into groups of at least two leaflets L, for example three in a transcatheter aortic valve implant (TAVI), to be incorporated into a BHV 10. This is performed in a grouping station 700.

[0119] The performance of the above described equipment 100 and method for preparation of leaflets L for BH s 10 was tested for assessing their effectiveness.

[0120] The technology that has been described herein above for measuring a thickness of patches P of pericardium tissue was duly validated. A test was carried out for comparing results measured with the 3D optical dynamic fringe projection sensor 240 described above with values obtained from mechanical contact measurements. Thickness measurements performed by said sensor 240 in the present equipment 100 were observed to be coincident with those involving a pressure exerted on a surface of the pericardium tissue of the patch P by a contact sensor, equal to 0.01 N / mm2. The test was carried out using a Litematic 0.01 N system from Mitutoyo, which is considered a reference for measuring thicknesses in soft samples.

[0121] Furthermore, as stated above, an ellipse E, as shown in figure 11 , corresponds to the far-field diffraction pattern obtained by the improved SALS sensor 300 of the present equipment 100 when a laser beam 305 is passed through the thickness of pericardium tissue of a patch P. The eccentricity s of said ellipse E is associated with a dispersion of the collagen fibers in the pericardium tissue of a patch P. In the tests carried out, the results of the eccentricity s and the orientation of the major axis A and the minor axis B of said ellipse E were found to match perfectly with the architecture of the fibers observed through a non-linear second harmonic generation (SHG) microscope.

[0122] More specifically, a SHG microscope was employed for validating the through thickness orientation and dispersion map 255 of the collagen fibers obtained by the improved SALS sensor 300 of the present equipment 100. The SHG microscope used in the test allowed for imaging individual collagen fibers in a patch P or in a leaflet L down to a depth of about 100 .m. It was found that the results of eccentricity s and the orientation of the major axis A of the ellipse E obtained with the SALS sensor 300 matched perfectly with the architecture of the fibres observed with the SHG microscope.

[0123] Gauging instrument’s accuracy was also checked through a gage repeatability and reproducibility (GR&R) test. Again, excellent results in the measurement of the thickness map of the patches P and the leaflets L were also obtained.

[0124] The advantages of the present equipment 100 and method for preparation of leaflets L for BHVs 10 are numerous.

[0125] The most important advantage is that the present equipment 100 has been found to significantly contribute to improving hemodynamic efficiency and durability of BHVs 10 for the benefit of patients requiring their implantation.

[0126] Furthermore, non-contact optical inspection, both for the leaflet manufacturing stage and for final quality control and certification of leaflets L before being sewn to heart valve frames 15 was found to be extremely efficient. The improved 2D and 3D optical sensors 225, 235, 240, make it possible to quickly and reliably obtain the parameters required to obtain a good selection of optimal leaflets L for the manufacture of BHVs 10. In particular, acquisition and analysis speed of the 2D, 3D optical sensors 225, 235, 240 and the improved SALS sensor 300 is optimized. In this respect, it has been found that digitization, analysis and cutting operations of the patches P can be performed in less than 5 minutes, which is the time during which no rehydration of the tissue is required. Rapid analysis of large tissue areas can be performed avoiding expensive equipment and time-consuming tissue preprocessing steps.

[0127] In addition, the extended digitization capabilities of the present equipment 100 through the development of new 2D, 3D optical sensors 225, 235, 240, and the improved SALS sensor 300 allows thickness map 250 of pericardial tissues to be reliably obtained, as well as detection of critical defects, collagen density, and architecture of collagen fibers in the patches P of pericardium tissue. These are essential elements that improve the quality of leaflets L, and thus the efficiency and durability of BHVs 10 implanted in patients affected by aortic stenosis.

[0128] The equipment 100 described above is intended for, but not limited to, managing patches P having a maximum size of 120 x 120 mm. Such values for the maximum size of the patches P to be processed has been found to allow the risk of dehydration of pericardium tissue patches P to be advantageously mitigated.

[0129] The present equipment 100 also provides improved productivity and performance making it possible to partially or completely automate manufacturing and inspection processes of BHVs 10. In this respect, it has been found that the leaflet processing capacity of the present equipment 100 is of the order of 20 certified leaflets L per hour, equivalent to 30,000 leaflets L per year in a single shift. This means that, with a single equipment 100, a heart valve manufacturer could produce enough leaflets L to produce 10,000 aortic heart valves in a year. In addition, productivity could be easily increased by virtue of the modular and scalable nature of the present equipment 100. This is advantageous in particular for the case of heart valve manufacturers. Also, the estimated cost of manufacturing each leaflet L, assuming a capital investment payback period of two years, is extremely low compared to current manual manufacturing costs.

[0130] In addition to the above mentioned competitive improvements in terms of cost, as well as intrinsic improvements in quality and reliability, it is to be noted that traceability during the manufacturing process is also improved, meeting the requirements of the Medical Device Regulation (MDR).

[0131] Although examples of the present equipment 100 and method for preparation of leaflets L for BHVs 10 have been disclosed herein, other alternatives, modifications, and / or equivalents thereof are possible. All possible combinations of the described examples are also covered. The scope of the present disclosure should not be limited by the examples disclosed herein but should be determined only by a fair reading of the claims that follow.

[0132] Reference signs related to drawings placed in parentheses in a claim are solely for attempting to increase the intelligibility of the claim and shall not be construed as limiting the scope of the claim.

Claims

CLAIMS1- Equipment (100) for preparation of leaflets (L) for bioprosthetic heart valves (10), the equipment (100) comprising:- a digitization station (200) for digitization of at least one patch (P) of animal pericardium tissue comprising a stack of layers of collagen fibers, the digitization station (200) including:- at least one 2D optical sensor (225) for imaging the patch (P) with at least one of bright-field epi-illumination means (220) and transillumination means (230) for at least one of detection of defects in the patch (P) and assessing a collagen density in the patch (P);- at least one 3D optical sensor (240) for obtaining a thickness map (250) of the patch (P); and- a small-angle light scattering (SALS) system (300) for obtaining a through thickness orientation and dispersion map (255) of collagen fibers in the patch (P), wherein the SALS sensor (300) is configured for obtaining a far-field diffraction pattern where diffraction patterns generated in each of the respective layers of collagen fibers in the pericardium tissue crossed by a focused laser beam (305) with a wavelength of the same order of magnitude as a diameter of the collagen fibers and an interfiber spacing are superimposed; and- a selection station (400) for automatic selection of regions of the patch (P) considered optimal to form accepted leaflets (L) based on digital results obtained by the digitization station (200) and according to at least one leaflet model (L’).

2. The equipment (100) of claim 1 , wherein it further includes one or more of:- a cutting station (500) for cutting selected optimal regions of the patch (P) to form leaflets (L);- a inspection station (600) for inspection of leaflets (L) configured for performing dimensional verifications in the leaflet, measuring a thickness map of the leaflet (L), detecting critical defects in the leaflet (L), assessing a collagen density in the leaflet (L) and measuring through thickness orientation and dispersion of collagen fibers in the leaflet (L), and considering a leaflet (L) as accepted or rejected when compared with a leaflet model (L’); and- a grouping station (700) for arranging accepted leaflets (L) into groups of at least two leaflets (L) to be incorporated into a bioprosthetic heart valve (10).

3. The equipment (100) of claim 2, wherein the cutting station (500) for cutting selected optimal regions of a patch (P) comprises a laser cutting device (550).

4. The equipment (100) of claim 3, wherein the laser cutting device (550) is a CO2 laser cutting device.

5. The equipment (100) of claim 2 or 3, wherein the cutting station (500) for automatic cutting selected optimal regions of a patch (P) comprises a die cutting device.

6. The equipment (100) of any preceding claim, wherein the digitization station (200) for digitization of patches (P) is also configured for inspection of leaflets (L).

7. The equipment (100) of any preceding claim, wherein the patches P (P) have maximum size of 120 x 120 mm.

8. The equipment (100) of any preceding claim, wherein if further includes feeding means (800) for feeding patches (P) or leaflets (L) through the stations.

9. The equipment (100) of any preceding claim, wherein the 3D optical sensor (240) is one of dynamic fringe projection, confocal, interferometric, and focus variation.

10. The equipment (100) of any preceding claim, wherein the 3D optical sensor (240) has a 5 megapixel camera with a lateral resolution better than 15 .m and a vertical resolution better than 1 .m.

11. The equipment (100) of any preceding claim, wherein the SALS sensor (300) includes a He-Ne laser with a wavelength of 632.8 nm.

12. The equipment (100) of any preceding claim, wherein it further includes support means (900) for the pericardium tissue, the support means (900), when in contact with hydrated pericardium tissue, behaving:- as an optically diffusing surface for the 3D optical sensor (240) and 2D optical sensor (225) with bright-field epi-illumination means (220); and- as an optically transparent surface for the 2D optical sensor (225) with transillumination means (230).

13. The equipment (100) of claim 12, wherein the support means (900) for the pericardium tissue comprises a plane-parallel glass plate (950) having a polished side, and also a textured side where the hydrated pericardium tissue is to be placed.

14. A method for preparation of leaflets (L) for bioprosthetic heart valves (10) through an equipment (100) according to any of the claims 1-13, the method comprising:- loading an animal pericardium tissue in the digitization station (200) for digitization of at least one patch (P) of animal pericardium tissue;- obtaining at least one of 2D bright-field image and a 2D transillumination image of the patch (P);- obtaining a 3D thickness map (250) of the patch (P);- obtaining a through thickness orientation and dispersion map (255) of collagen fibers in the patch (P); and- selecting regions of the patch (P) considered optimal to form accepted leaflets (L) according to at least one leaflet model (L’), the selecting step being based on digital results obtained from the digitization station for digitization by applying an optimization process based on a mathematical optimization technique.

15. The method of claim 14, wherein it further comprises:- cutting selected optimal regions of the patch (P) to form leaflets (L);- inspecting leaflets (L), the inspection step comprising performing dimensional verifications in the leaflet, measuring a thickness map (250) of the leaflet (L), detecting critical defects in the leaflet (L), measuring through thickness orientation and dispersion of collagen fibers in the leaflet, and considering a leaflet (L) as accepted or rejected when compared with a leaflet model (L’); and- arranging accepted leaflets (L) into groups of at least two leaflets (L) to be incorporated into a bioprosthetic heart valve (10).

16. The method of claim 14 or 15, wherein selecting regions in the patch considered optimal to form accepted leaflets (L) according to a leaflet model (!_’) comprises:- excluding regions of the patch (P) having critical defects; and- determining the best positions for a leaflet (L) that meet the through thickness orientation and dispersion of the collagen fibers and thickness parameters previously defined in the leaflet model (L’).

17. The method of any of the claims 14-16, wherein the far-field diffraction pattern obtained by the small-angle light scattering (SALS) system (300) is an ellipse (E), the ellipse (E) being characterised by a value of eccentricity (s) and an orientation of the major axis (A) and the minor axis (B) of the ellipse (E).

18. The method of claim 17, wherein the through thickness orientation of collagen fibers is coincident with the orientation of the minor axis (B) of the ellipse (E), and wherein the through thickness dispersion of collagen fibers is obtained from the eccentricity (s) of the ellipse (E).

19. The method of any of the claims 15-18, wherein it further comprises rehydration of cut leaflets (L).

20. The method of any of the claims 15-19, wherein the cutting step includes forming holes in the accepted leaflet (L) for subsequent sewing of the leaflet (L) to a frame (15) of the bioprosthetic heart valve (10).

21. The method of any of the claims 14-20, wherein selecting regions in the patch (P) considered optimal to form accepted leaflets (L) according to a leaflet model (L’) involves at least one of two additional optimization variables resulting from a mean tissue thickness at commissure regions being above an average value of the tissue thickness of the leaflet and a collagen density at commissure regions being above an average value of the collagen density of the leaflet.

22. The method of any of the claims 14-21 , wherein critical defects in said regions to be excluded are detected from at least one of the 2D bright-field and 2D transillumination images.

23. The method any of the claims 14-22, wherein the mathematical optimization technique is a metaheuristic simulated annealing probabilistic technique.

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