Microfluidic filtration chamber and microfluidic device, methods and uses thereof

Abstract

The present disclosure relates to a microfluidic filtration chamber and to a microfluidic flow device designed for separating and concentrating tumor cells or other target molecules / cells from a fluidic sample.

Description

D E S C R I P T I O NMICROFLUIDIC FILTRATION CHAMBER AND MICROFLUIDIC DEVICE, METHODS AND USES THEREOFTECH NICAL FIELD

[0001] The present disclosure relates to biomedical devices. In particular, the present disclosure relates to the field of microfluidics and more particularly to a microfluidic filtration chamber and to a microfluidic flow device for separating and concentrating fluidic samples to identify target molecules or cells.BACKGROUND

[0002] Circulating tumor cells, abbreviated as CTCs, originated from a primary tumor and travel via the bloodstream to various body regions, potentially infiltrating other organs and sparking metastasis. CTCs exhibit heterogeneity, varying in size and characteristics. Identifying CTCs offers critical insights into managing cancer patients, offering a real-time glimpse into their tumor load. However, detecting CTCs poses a challenge due to their exceedingly low presence, often found at concentrations of 1 to 10 CTCs per billion blood cells.

[0003] Numerous techniques exist for isolating and characterizing CTCs, yet each method presents several disadvantages, in particular lacking sensitivity and reliability, stability, adequate mechanical structure and suitability for large-scale production.

[0004] For instance, the prior art describes microscale cell filters for trapping cells, nonetheless, the devices described present inadequate flow stability and pressure, due to inadequate design and usage of inappropriate materials, which lead to a lack of efficiency on trapping CTCs with high sensitivity, selectivity and reproducibility. Additionally, the devices described in the prior art do not enable the quantification of trapped cells directly within the microscale cell filter. Instead, the cells must be transferred to another suitable support for quantification and / or characterization, which compromises precision, reproducibility, sensitivity, and overall reliability.

[0005] Thus, there's a demand for an analytical system capable of efficiently managing samples containing diverse cell sub-populations. Specifically, there is a need of a device able to provide an adequate filtration of the blood sample, and at the same time provide accurate results in concentrating and separating CTCs, in a fast and reliable way.

[0006] These facts are disclosed to illustrate the technical problem addressed by the present disclosure.GENERAL DESCRIPTION

[0007] The present disclosure relates to an improved microfluidic filtration chamber for separating and concentrating circulating tumour cells and to a microfluidic flow device comprising a plurality of said microfluidic filtration chambers.

[0008] An aspect of the present disclosure relates to a microfluidic filtration chamber for separating and concentrating (preferably, separating, concentrating and quantifying) circulating tumor cells from a fluidic sample, comprising: an inlet (1) and an outlet (2) for the entry and exit of the sample; a plurality of first post elements (3) for prefiltering the sample (to remove larger debris and nontarget cells, minimizing turbulence and preventing the formation of pressure peaks, that may damage the sample and increase risk for device failure); a plurality of second post elements (4) for filtering the sample; wherein: the plurality of first post elements (3) (pre filters) is arranged in a zigzag geometry between adjacent first post elements; the plurality of first post elements is interspaced, thereby forming a plurality of first gaps (5), each first gap being formed between two adjacent first post elements; the plurality of second post elements (4) is interspaced, thereby forming a plurality of second gaps (7), each second gap being formed in between two adjacent second post elements; the plurality of first post elements and the plurality of second post elements is arranged such that the flow of the sample from the inlet flow channel to the outlet flow channel passes through the plurality of first gaps and through the plurality of second gaps for separating and concentrating circulating tumor cells.

[0009] Surprisingly, the shape and distribution of the first post elements (prefilters) allow to distribute evenly the sample across the filter area, maximizing the filter capacity and the device throughput, and the volume of fluid that each device can process. Also, they ensure the uniformity of the flow and cell linear velocity across the second filter post elements, ensuring optimal trapping conditions (capture efficiency) across the device.

[0010] The design and disposition of the first post elements, with lozenge shape and arranged in a zigzag geometry, allows to obtain a stable flow inside the microfluidic filter chamber, avoiding pressure peaks and consequently cell damage and risk of device failure, and at the same time providing improved pre-filtration of potential debris or clots that could block the filter area.

[0011] Surprisingly, it was found that the specific structure of the microfluidic filtration chamber according of the present disclosure, in particular the design and disposition of the first post elements(lozenge / diamond shape with), allow to obtain flow uniformity and avoid pressure peaks. As depicted in Figures 9-11, while other structures (comparative examples 1-2) lead to variations in the flow across the microfluidic filtration chamber and flow above 1, the structure of the microfluidic filtration chamber of the present disclosure (Figure 9) provides flow uniformity and a flow below 1; preferably below 0.8.

[0012] The design and disposition of the second post elements, arranged in a zigzag geometry (and preferably with cylinder shape) provided improved results in selectively filtering and concentrating CTCs.

[0013] The microfluidic filtration chamber of the present disclosure allows to sustain higher flow rates and higher viscosity samples, while avoiding clogging and overpressure, which could lead to leaking and debonding. The microfluidic filtration chamber has optimized channel geometries for uniform flow distribution and pressure uniformity stability. The specific first post elements (pre-filtering area) with rhombus / diamond-shaped structures allow to maintain consistent pressure and minimize turbulence and the zigzag arrangement of second post elements (filtering area) allow to improve filtration capacity.

[0014] The arrangement of first post elements in the filtering area creates specific filter gaps for effective separation of circulating tumor cells from other sample components.

[0015] In an embodiment for better results, the width of each first gap ranges from 80 to 200 pm.

[0016] In an embodiment for better results, the aspect ratio (ratio between their height and width) of each first gap ranges from 1:2 to 1:7.

[0017] It was surprisingly found that the width (post-to-post distance) of the second gaps (gaps formed in between two adjacent second post elements) is crucial to allow proper selective trapping of CTCs. Second gaps width ranging from 6-7.5 pm (preferably 6.5 pm) and channel depth (channel height) ranging from 28-35 pm (preferably 30 pm) allows small RBCs to flow through seamlessly and force WBCs to deform in the vertical axis to fit and flow through, while trapping CTCs that are larger than the horizontal gap and cannot deform vertically.

[0018] The microfluidic filtration chamber of the present disclosure provides improved characteristics:Flow and pressure uniformity: The microfluidic filtration chamber is designed with optimized channel geometries that promote uniform flow distribution. The plinko arrangement (i.e., wherein each row of first post elements is laterally offset with respect to adjacent upstream and / or downstream rows) of the first post elements and zigzag geometry of the second post elements (in the prefiltering and filtering area, respectively) creates a controlled pathway for the sample, minimizing turbulence and preventing the formation of pressure peaks. In a preferred embodiment, the microfluidic filtration chamber has a lozenge / rhombus (diamond) shape. The lozenge / rhombus (diamond) shape of the microfluidic filtration chamber allows to maintain consistent pressure across the filtering elements;Sample storage: The microfluidic filtration chamber is manufactured using materials that are impermeable to oxygen and moisture, such as COP (cyclo olefin polymer), acrylic or PA (polyamide), with low gas permeability. These materials protect the internal components from oxidative degradation, thereby enhancing the shelf life of the device.Manufacturing for Large-Scale Production: The design of the microfluidic filtration chamber is optimized for scalability in manufacturing. The use of injection molding and hot embossing techniques allow for the mass production of the chamber components with high precision and repeatability.Sensitivity and reproducibility: The precise arrangement and design of the first and second post elements creates highly specific filter gaps that effectively separate CTCs from other components in the sample, in particular red blood cells (RBCs) and white blood cells (WBCs), in a sensitive and reproducible way.

[0019] The microfluidic filtration chamber of the present disclosure showed to be mechanically robust and supports high flow rates. It is stable, suitable for large-scale production, and exhibits high performance to separate and concentrate circulating tumor cells (CTCs), along with reliable precision, achieving results in less than one hour.

[0020] In a preferred embodiment, the first post elements are arranged in a plinko distribution, i.e., the plurality of first post elements is arranged in multiple rows.

[0021] In a preferred embodiment, each row of first post elements is laterally offset with respect to the adjacent upstream and / or downstream rows.

[0022] In a preferred embodiment, the plurality of first post elements (3) (pre filters) is arranged in a zigzag geometry between adjacent first post elements, forming an angle ranging from 30 to 60 degrees between adjacent first post elements.

[0023] In a preferred embodiment, the microfluidic filtration chamber further comprises a central post element (9) (a large prefilter) on the top of the filtering chamber, followed by a distribution of first post elements (smaller prefilters). It was surprisingly found that said central post element allows to improve the uniformity of flow and hence increasing filtration efficiency.

[0024] In a preferred embodiment, the central post element has a rhombus shape (losange / diamond shape), preferably with a horizontal diagonal ranging from 1000-900 pm and a vertical diagonal ranging from 450-550 pm.

[0025] In a preferred embodiment, the first post elements are rhombus shaped (losange / diamond shape).

[0026] In a preferred embodiment, the rhombus shaped first post elements have a horizontal diagonal ranging from 300-400pm and a vertical diagonal ranging from 150-250 pm.

[0027] In a preferred embodiment for better results, each row of first post elements is laterally offset with respect to adjacent upstream and / or downstream rows.

[0028] In a preferred embodiment, each second post element of the plurality of second post elements has a cylindrical shape, preferably having a diameter ranging from 25-40 pm, preferably 35 pm.

[0029] In a preferred embodiment, the plurality of second post elements is arranged in a zigzag geometry.

[0030] In a preferred embodiment, the plurality of second post elements is arranged in a zigzag geometry forms an angle ranging from 28°-32° between each adjacent second post element; preferably 309; to increase the filtering area.

[0031] In a preferred embodiment, the diameter of each second post element ranges from 28-40 pm; preferably 35 pm.

[0032] In a preferred embodiment, the height (depth) of each second post element ranges from 28-35 pm; preferably 30 pm.

[0033] In a preferred embodiment, the width of each second gap ranges from 6 to 7.5 pm (preferably 6.5 pm).

[0034] In an embodiment, each second gap has an aspect ratio between its height and width ranging from 3.7 to 5.8 (gap ratio=gap height / gap width).

[0035] In a preferred embodiment, the width of the second gap is 6.5 pm.

[0036] In a preferred embodiment, the height (depth) of each second gap ranges from 28-35 pm; preferably 30 pm. In a preferred embodiment, the filtering area depth (height) is 30 pm.

[0037] In a preferred embodiment, the width of each first gap ranges from 80 to 200 pm; preferably 200 pm.

[0038] In an embodiment, the plurality of second post elements is arranged in linear (straight) geometry, forming an angle of 90° between each adjacent second post element. Preferably the height of each second post element ranges from 28-35 pm.

[0039] In a preferred embodiment, the plurality of first post elements and the plurality of second post elements are arranged such that a flow of the sample flowing from the inlet flow channel to the outlet flow channel passes through the plurality of first gaps and the plurality of second gaps.

[0040] In an embodiment for better results, the materials used for manufacturing the microfluidic filtration chamber are impermeable to oxygen and moisture, and hold relevant optical properties.

[0041] In an embodiment for better results, the material of the microfluidic filtration chamber comprises a material selected from the group consisting of cyclo olefin polymer, cyclic olefin copolymer, acrylic, polyamide, or mixtures thereof; preferably the material of the microfluidic filtration chamber is a material selected from the group consisting of: cyclo olefin polymer, cyclic olefin copolymer, acrylic, polyamide, or mixtures thereof.

[0042] Preferably, the material is selected from cyclo olefin polymer (COP), cyclo olefin polymer (COC), acrylic, or polyamide (PA), pressure-sensitive adhesive materials (PSA) or mixtures thereof.

[0043] In a preferred embodiment, the material is cyclo olefin polymer (COP).

[0044] In an embodiment for better results, the chamber is optimized for scalability in manufacturing using injection molding or hot embossing techniques.

[0045] In an embodiment for better results, the microfluidic filtration chamber further comprises a substrate and a cover for sealing the microfluidic filter chamber.

[0046] In an embodiment for better results, the first post elements are disposed in a plinko distribution.

[0047] In an embodiment for better results, each first post element of the plurality of first post elements has a lozenge (rhombus / diamond) shape.

[0048] In an embodiment for better results, the inlet and the outlet of the microfluidic filtration chamber have a height ranging from 150-400 pm; preferably 190-210 pm; more preferably is 200 pm.

[0049] In an embodiment for better results, the height of the inlet and / or the height of the outlet ranges from 180-220 pm; preferably 190-210 pm; more preferably is 200 pm.

[0050] In an embodiment for better results, each first post element and each second post element is arranged in between the substrate and the cover.

[0051] In an embodiment for better results, each first post element has a height ranging from 29-31 pM; preferably 30 pm.

[0052] In an embodiment for better results, each second post element has a height ranging from 29-31 pm; preferably 30 pm.

[0053] In an embodiment for better results, the angle formed between two adjacent second post elements ranges from 29 to 31 °; preferably ranges from 29° to 30 °; more preferably is 29.83 °.

[0054] In an embodiment for better results, the diameter of each second post element ranges from 34- 36 pm; preferably is 35 pm.

[0055] In an embodiment for better results, the substrate and / or the cover is transparent

[0056] In an embodiment for better results, the fluidic sample is a body fluid sample; preferably a blood sample.

[0057] Another aspect of the present disclosure relates to a microfluidic flow device for separating and concentrating (preferably separating, concentrating and identifying) a target molecule or cell from a fluidic sample (preferably circulating tumor cells from a fluidic sample), comprising: a plurality of microfluidic filtration chambers as described in the present disclosure; an inlet and an outlet for the entry and exit of the sample; an inlet array of channels for directing the sample through the plurality of microfluidic filtration chambers, wherein the inlet channel is connected to at least two new channels to split the sample, with each of these channels subsequently connected to another set of two new channels, progressively decreasing the volume of the flow sample arriving at the chambers, thereby allowing the separation and concentration of the fluidic sample to identify a target molecule or cell.

[0058] The microfluidic flow device of the present disclosure allows to sustain higher flow rates and higher viscosity samples, while avoiding clogging and overpressure, leading to leaking and debonding. It provides high precision and repeatability in large-scale production and the filtration chambers within the device effectively separate and concentrate target molecules or cells, including circulating tumor cells, from other components in the fluidic sample.

[0059] The inlet array of channels ensures a uniform distribution of the fluidic sample across the plurality of microfluidic filtration chambers of the device of the present disclosure.

[0060] The microfluidic flow device of the present disclosure provides improved performance by avoiding the buildup of high pressures (and backpressure) due to its novel multilevel design. This structural feature allows to support elevated and sustained flow rates, preventing clogging, ensuring uninterrupted operation, and as consequence it enables accelerates processing.

[0061] Most importantly, the system can manage samples with higher viscosity, a crucial feature for applications involving whole blood samples. Maintaining flow and functionality without compromising accuracy or clogging is essential and it is assured by the device of the present disclosure.

[0062] Within the present disclosure, the term "multilevel" refers to a configuration in which the channels of the inlet area of the array of channels may have different depths (i.e., different heights) relative to the depths (heights) of the chambers containing the first and second post elements.

[0063] In a preferred embodiment, the depth of each inlet channel is superior to the depth of each of the plurality of microfluidic filtration chambers.

[0064] In a preferred embodiment, the microfluidic flow device further comprises an outlet array of channels for directing the sample to the outlet for the exit of the sample.

[0065] In a preferred embodiment, the depth of each inlet channel is at least 4 times superior to the depth of each of the plurality of microfluidic filtration chambers.

[0066] In a preferred embodiment, the depth of each inlet channel is 4- 7 times superior to the depth of each of the plurality of microfluidic filtration chambers.

[0067] In a preferred embodiment, the depth of each inlet channel is 6-7 times superior to the depth of each of the plurality of microfluidic filtration chambers.

[0068] The microfluidic flow device with multiple levels (multilevel microfluidic flow device) was optimized to reduce pressure build up within the chip. This reduces cell damage while simplifying sample processing as less force is needed to achieve a stable sample flow rate. Moreover this feature enables the use of chip within a larger consumable cartridge. Lower pressure buildup within the chip also reduces the design complexity associated with any capital equipment which is required for processing samples. In a preferred embodiment, depths of 200 pm in the inlet, in the outlet and in each channel of the inlet array of channels of the microfluidic flow device and depths of 30 pm in the microfluidic filtration chambers containing the first and second post elements have shown to display pressures lower than 0.4 psi during processing of whole blood samples. Flow uniformity leveraging feature sizing and feature geometry leads to better overall distribution and filtering of whole blood which creates less pressure build up, more homogeneity of cell capture across pillar features and reduces field of view required to image and identify cancerous cells. These features significantly reduced the risk of leakage / burst. Uniform flow provides smaller field of view required for fluorescent imaging.

[0069] In a preferred embodiment, the height (depth) of the inlet and an outlet for the entry and exit of the sample of the microfluidic flow device ranges from 150-400 pm depth; preferably is 200 pm.

[0070] In a preferred embodiment, the height (depth) of each channel (of the inlet and outlet array of channels) ranges from 150-400 pm depth (preferably 200 pm). The channels distributes the flow to independent microfluidic filtration chambers.

[0071] The dimensions of the channels decrease the total pressure of the device enabling higher flow rates while maintaining homogeneous pressure throughout the microfluidic flow device.

[0072] The multilevel microfluidic flow device provides low inlet pressure, a result of the specific aspect ratio of the first channel of the inlet array of channels— specifically, a depth of approximately 200 pm. This geometry enables efficient fluid entry while simultaneously preserving effective filtration performance, through reduction in depth within the filtration zone to 29-35 pm (preferably 30 pm).

[0073] The device also provides uniform flow and pressure distribution throughout the microfluidic filtration chambers, which significantly reduces the occurrence of pressure peaks that could compromise structural integrity, such as by causing leaks.

[0074] The materials and specific design of the microfluidic flow device of the present disclosure allows to obtain a stable flow inside the device, avoiding pressure peaks, cell damage and consequently reducing risks of failure.

[0075] The design of the microfluidic flow device also enhances manufacturability: larger bonding areas improve manufacturing and reproducibility and reduce the risk of device failure.

[0076] In an embodiment for better results, the filtration chambers within the device effectively separate and concentrate target molecules or cells, including circulating tumor cells (CTCs), from other components in the fluidic sample.

[0077] In an embodiment for better results, the blank area between chambers is increased to ensure adequate support for the channels, said blank area ranging from 50 pm to 400 pm, facilitating manufacture via increased surface for bonding and reducing risks in manufacture process as well as in sample processing and prevent leaking.

[0078] Within the present disclosure, blank area refers to the area between the microfluidic filtration chambers (not cojoined chambers) in the shorter end of the lozenge, in between two adjacent isolation chambers (comprised in a microfluidic flow device).

[0079] Connected chambers produce a large chamber section (more filter area), but compromise flow homogeneity. Separated chambers produce homogeneous flow, but with a smaller filter area, compromise isolation efficiency or increase processing times. This is since the same flow rates produce higher linear velocities of the cells in the filter area. A solution was found by maintaining separate chambers, that are wider, to maintain a large filter area, while still providing larger blank area for bonding and reduced probability of leaking / bursting.

[0080] In an embodiment for better results, the inlet, the outlet and the inlet and outlet array of channels of the microfluidic flow device are adjusted to have the same length to avoid pressure differences and ensure even flow distribution across the different chambers of the device.

[0081] In an embodiment for better results, the interface tubing / device connection uses technologies that sustain pressure and avoid compromising imaging of the filter area. These include a laser-welded, thermal gluing, Luer connectors and combinations thereof.

[0082] In a preferred embodiment, the fluidic sample is a body fluid sample (blood, urine, cerebrospinal fluid, bladder washes, saliva); preferably a blood sample.

[0083] In a preferred embodiment, the outlet array channels have a configuration substantially identical to the inlet array channels.

[0084] In a preferred embodiment, the height of the inlet channel and of the height of the outlet channel ranges from 150-400 pm, preferably 200 pm.

[0085] In an embodiment for better results, the microfluidic flow device comprises at least 4 microfluidic filtration chambers; preferably at least 8 microfluidic filtration chambers; more preferably at least 32 microfluidic filtration chambers; even more preferably 32 microfluidic filtration chambers.

[0086] In a preferred embodiment, the cross section area of each channel (of the inlet array of channels) is inferior in relation to the cross section area of predecessor channel as they approach to the microfluidic filtration chamber; preferably wherein the cross section area of each channel is at least 10% inferior of the cross section of the predecessor channel; more preferably wherein the cross section area of each channel is at least 20% inferior of the cross section of predecessor channel; even more preferably wherein the cross section area of each channel is at least 30% inferior of the cross section of predecessor channel; even more preferably wherein the cross section area of each channel is at least 50% inferior of the cross section of predecessor channel.

[0087] The cross section of a channel is the two-dimensional profile obtained by taking a plane perpendicular to the longitudinal axis (i.e., direction of flow) of the channel. This profile defines the shape and size of the internal flow passage and is typically characterized by parameters such as width and height (or depth)

[0088] In an embodiment for better results, the microfluidic filtration chamber is a single-use microfluidic filtration chamber

[0089] In an embodiment for better results, the microfluidic flow device is a single-use microfluidic flow deviceBRIEF DESCRIPTION OF THE DRAWINGS

[0090] The following figures provide preferred embodiments for illustrating the disclosure and should not be seen as limiting the scope of invention.

[0091] Figure 1: Schematic representation of example 1 of the microfluidic filtration chamber of the present disclosure. 1: inlet; 2: outlet; 3: first post element; 4: second post element.

[0092] Figure 2: Schematic representation of a zoomed view of a plurality of first post elements of example 1. 5: first gap.

[0093] Figure 3-Figure 5: Schematic representation of a zoomed view of a plurality of second post elements of example 1. 4: second post element; 7: second gap; 8: angle between two adjacent second post elements.

[0094] Figure 6: Schematic representation of example 2 of the microfluidic filtration chamber of the present disclosure. 9: central post element.

[0095] Figure 7. Design of a microfluidic filtration chamber according to comparative example 1.

[0096] Figure 8. Design of a microfluidic filtration chamber according to comparative example 2.

[0097] Figure 9. Analysis of flow CV (coefficient of variance) on a microfluidic filtration chamber according to the present disclosure (in particular, example 2 of the present disclosure).

[0098] Figure 10. Analysis of flow CV (coefficient of variance) of comparative example 1 (microfluidic filtration chamber without first post elements, Figure 7).

[0099] Figure 11. Analysis of flow CV (coefficient of variance) of comparative example 2 (microfluidic filtration chamber comprising first post elements, wherein said first post elements are placed parallelly to each other and said first post elements have a rectangular shape, Figure 8).

[0100] Figure 12. Microfluidic flow device comprising a plurality of microfluidic filter chambers according to the present disclosure.DETAILED DESCRIPTION

[0101] The present disclosure relates to a microfluidic filtration chamber and a microfluidic flow device designed for separating and concentrating tumor cells or other target molecules / cells from a fluidic sample. The microfluidic filtration chamber features optimized channel geometries and a zigzag arrangement of post elements, promoting uniform flow distribution, minimizing turbulence, and preventing pressure peaks.

[0102] The zigzag configuration or geometry in a device typically refers to the structural design or pattern that incorporates zigzag-shaped components or elements. This geometric arrangement is functional, providing an efficient pre-filtration of the sample. Additionally, the zigzag in the second posts - filtering area - increases the surface available for filtering.

[0103] The present disclosure relates to a microfluidic filtration chamber for separating and concentrating circulating tumor cells (CTCs) from a fluid sample, preferably blood.

[0104] The microfluidic filtration chamber and the microfluidic flow device described in the present disclosure are applicable in various biomedical fields, including cancer diagnostics, blood analysis, andmolecular biology research. The device of the present disclosure can precisely separate and concentrate target cells or molecules from fluid samples rendering it valuable for clinical and laboratory use.

[0105] In an embodiment for better results, the first post elements are arranged in a plinko distribution, i.e., wherein each row of first post elements is laterally offset with respect to adjacent upstream and / or downstream rows.

[0106] The microfluidic filtration chamber of the present disclosure allows to process a volume of at least 7.5 mL of whole blood, and to physically isolate CTCs from blood cells, based on their size and rigidity using a filter structure. Each mL of whole blood contains billions of red blood cells (RBCs), and millions of white blood cells (WBCs), while one a few (1-10) CTCs. Regarding their size, CTC are reported to have diameters from 15 to 30pm. RBCs have a disc shape with approximately 5pm in diameter, so they are significantly smaller, but WBCs range from 7 to 21 pm, so they can overlap in size with CTCs. In terms of their deformability, CTCs have a high nucleus to cytoplasm ratio (with the nucleus accounting for over 70% of the cell diameter), so they are reasonably rigid. On the other hand, the WBCs with larger sizes have a small nucleus or are polynucleated granulocytes, hence with a big capacity to deform. Hence, it was necessary to develop a microfluidic filtration chamber able to physically trap in a microfluidic filtration chamber the CTCs while allowing RBCs and WBCs to flow through.

[0107] Surprisingly, the microfluidic filtration chamber of the present disclosure allows to selectively trap the CTCs over the remaining blood cells, in particular RBCs and WBCs. The microfluidic filtration chamber of the present disclosure was designed containing a plurality of first post elements (for prefilter the sample) and a plurality of second post element (for filtering the sample) arranged in a line perpendicular to the direction of the flow, forcing the cells in the sample against them.

[0108] Regarding the second post elements, the gap between the two adjacent post elements and the height of the channel was specifically designed to allow small RBCs to flow through seamlessly and force WBCs to deform in the vertical axis to fit and flow through, while trapping CTCs that are larger that the horizontal gap and cannot deform vertically.

[0109] Another aspect to be considered is the inclusion in the microfluidic filtration chamber of a structure to pre-filter the sample to retain potential debris or clots that could block the filter area. Nonetheless, these structures frequently lead to turbulence and pressure peaks and / or pressure variation over the microfluidic filter chamber, which could cause a lack in the effectiveness of the microfluidic filtration chamber to trap CTCs, reproducibility and eventually clogging and loss of sample. The microfluidic filtration chamber of the present disclosure, in particular the plurality of first post elements, allow to efficiently prefilter the sample while avoiding such disadvantages.

[0110] The microfluidic filtration chamber of the present disclosure comprises the following characteristics (as exemplified in Figure 1):Inlet and Outlet: The inlet and outlet of the microfluidic filtration chamber are designed to facilitate the entry and exit of the sample at low fluid resistance. The heights of the inlet and outlet range from 150 to 400 pm, ensuring compatibility with standard microfluidic connections and stable flow and pressure.First Post Elements: The first post elements serve as a prefilter to remove larger particles and debris from the sample. These posts are arranged in a zigzag geometry. Preferably, each first post element has a lozenge shape. The specific gaps between the posts are critical for initial filtering, with each gap ranging from 80 to 200 pm.Second Post Elements: The second post elements are responsible for the accurate filtering of the sample, specifically targeting circulating tumor cells over other blood cells, as red blood cells (RBCs) and white blood cells (WBCs). The gaps between these posts (gap between two consecutive second post elements) are precisely controlled to range from 6 to 7.5 pm. These posts are also arranged in a zigzag pattern, forming angles between 28° and 32°. The height of each second post element ranges from 28 to 35 pm and the diameter ranges from 25-40 pm.

[0111] Operation: In operation, the sample enters the microfluidic filtration chamber through the inlet. It first encounters the first post elements, which prefilter the sample by removing larger particles and debris. The partially filtered sample then flows through the second post elements, where the circulating tumor cells are separated and concentrated due to the specific dimensions and arrangement of the second gaps. The filtered sample exits through the outlet, while the circulating tumor cells are concentrated and retained in the second post elements, ready for further analysis.

[0112] To maintain symmetry, facilitate manufacturing, and ensure a consistent flow, first post elements can also be found after the second post elements, as showed in Figure 1 and Figure 6.

[0113] The microfluidic filtration chamber of the present disclosure filter and retains circulating tumor cells (CTCs) based on size and deformability. This method does not require staining or antibodies, which are typically necessary in most existing methods. Cells captured on the filter can be extracted for subsequent analysis. The microfluidic filtration chamber can be integrated into a microfluidic flow device (microfluidic chip), allowing trapped cells to be analyzed directly within the chip.

[0114] For instance, circulating tumor cells are present in blood at very low concentrations, ranging from 1 to 10 cells per billion blood cells. CTCs typically have diameters greater than 5 micrometers and exhibit higher density and lower compressibility than other cells. The cell filter can detect CTCs in blood by allowing smaller and / or more deformable cells to pass through, while trapping the CTCs, thereby facilitating their analysis for accurate prognosis, personalized treatment, and therapy monitoring.

[0115] The microfluidic filtration chamber captures all CTCs regardless of their phenotype, including both epithelial and mesenchymal CTCs. It can be used to provide a real-time snapshot of the current tumor burden and for liquid biopsies of whole blood, urine, and other body fluids; preferably blood. Forexample, it can process whole blood directly without the need for sample preparation or pretreatment. The microfluidic filtration chamber of the present disclosure is cost-effective and easy to manufacture and can be easily integrated with a biosensor for rapid and cost-efficient in-situ phenotypic and molecular profiling of tumor cells.

[0116] The microfluidic filtration chamber can be made from materials such as COP (cyclo olefin polymer), acrylic or PA (polyamide), or any combination of these materials.

[0117] The term 'trapping' should be interpreted broadly to include both capturing cells and blocking them from passing through.

[0118] In a preferred embodiment, the first post elements have all the same height.

[0119] In a preferred embodiment, the second post elements have all the same height.

[0120] The microfluidic filtration chamber may further comprise a substrate, wherein the plurality of post elements is integrally formed with the substrate.

[0121] The cell filter may further comprise a cover, wherein the plurality of post elements is arranged in between the substrate and the cover.

[0122] The substrate and post elements can be made of COP (cyclo olefin polymer), acrylic or PA (polyamide), or any combination of these materials. These materials are biocompatible and ensures cell viability, as well as allow to maintain a stable flow and pressure inside the microfluidic filter chamber.

[0123] The microfluidic filtration chamber may also include a cover, with the post elements arranged between the substrate and the cover.

[0124] In a preferred embodiment, the device is transparent or translucid, allowing for optical imaging and visual observation of the filter's status.

[0125] In a preferred embodiment, the aspect ratio of the gaps refers to the proportion between the height (measured from the substrate to the cover in the area between adjacent post elements) and the width (measured as the distance between two adjacent post elements).

[0126] The dimensions of the second post elements are designed to create multiple gaps between them. These gaps are essential for sorting and retaining CTCs based on their size and deformability, while permitting the sample and unwanted cells to pass through the remaining open gaps.

[0127] Each of the first post elements have a lozenge (diamond) shape. It was surprisingly found that first post elements comprising a lozenge shape provide improved pre-filtration and allow to obtain a stable flow of the fluidic sample inside the microfluidic filter chamber, which is important to avoid peak pressure and to ensure proper pre-filtration and filtration.

[0128] Each of the second post elements are circular cylinders. Circular cylinders post elements facilitate easier manufacturing and more predictable trapping effects due to their consistent geometry. Additionally, cylindrical shapes reduce the contact surface along the flow channels, minimizing the risk of cell adhesion and maintaining proper flow pressure. Circular cylinders post elements also enhance the compressing effect on deformable cells, making it easier for larger deformable cells to pass through the filter. The second post elements are arranged in a zigzag geometry, forming an angle ranging from 28°- 32° between two adjacent post elements, improving the selective trapping of circulating tumor cells.

[0129] The microfluidic filtration chamber may include a flow generator configured to drive the sample through the filter. This flow generator can be connected directly or indirectly to the inlet flow channel via tubes, channels, capillaries, or combinations thereof. The generator can provide various flow types, including peristaltic, continuous, or periodic flows, at different rates and intervals. It can be a manual or powered pump, such as a syringe pump, peristaltic pump, or pressure pump.

[0130] In respect to the multilevel microfluidic flow device (multilevel microfluidic flow device), its deeper inlet and outlet channels together with the chamber's lozenge shape ensures consistent and low pressure across the filtering elements, while the use of impermeable materials enhances sample storage by protecting the sample against oxidative degradation. The device is optimized for large-scale production using injection molding or hot embossing techniques. The precise arrangement of post elements creates specific filter gaps that effectively separate circulating tumor cells from other components such as red blood cells and white blood cell. The present disclosure addresses challenges related to flow uniformity, pressure control, leakage, shelf-life, and manufacturing scalability, providing a reliable and efficient solution for diagnostic and research applications.

[0131] The present disclosure relates to a microfluidic filtration chamber and a microfluidic device designed for separating and concentrating tumor cells or other target molecules / cells from a fluidic sample. The microfluidic filtration chamber features optimized channel geometries and a zigzag arrangement of post elements, promoting uniform flow distribution, minimizing turbulence, and preventing pressure peaks. The chamber's lozenge shape ensures consistent pressure across the filtering elements, while the use of impermeable materials enhances sample storage by protecting against oxidative degradation. The device is optimized for large-scale production using injection molding and hot embossing techniques. The precise arrangement of post elements creates specific filter gaps that effectively separate circulating tumor cells from other components such as red blood cells and white blood cell. The present disclosure addresses challenges related to flow stability, pressure control, leakage, shelf-life, and manufacturing scalability, providing a reliable and efficient solution for diagnostic and research applications.EXAMPLE 1

[0132] In an embodiment, the microfluidic filtration chamber of the present disclosure comprises the structure as depicted in Figure 1-3.

[0133] The microfluidic filtration chamber for separating and concentrating circulating tumor cells from a fluid sample according to example 1 (and depicted in Figure 1, Figure 2 and Figure 3) comprises: an inlet (1) and an outlet (2) for the entry and exit of the sample, respectively; a plurality of first post elements (3) for prefiltering the sample; a plurality of second post elements (4) for filtering the sample; a substrate and a cover for sealing the microfluidic filter chamber; wherein: the inlet and outlet have a diameter of 200 pm; the plurality of first post elements is arranged in a zigzag geometry to each other; each first post element of the plurality of first post elements has a lozenge (diamond) shape; the plurality of first post elements is interspaced, thereby forming a plurality of first gaps (5), each first gap being formed in between two adjacent first post elements, wherein each first gap has 200 pm; the plurality of second post elements are arranged in a zigzag geometry, forming an angle (8) of 29.83 ° between each adjacent second post element; each second post element of the plurality of second post elements has a circular cylindrical, wherein the width of each second post element is 35 pm; the plurality of second post elements is interspaced, thereby forming a plurality of second gaps (7), each second gap being formed in between two adjacent second post elements, wherein the size of each second gap is 6.5 pm and each second gap has an aspect ratio between its height and width of 5.4; the plurality of first post elements and the plurality of second post elements are arranged such that a flow of the sample flowing from the inlet flow channel to the outlet flow channel passes through the plurality of first gaps and the plurality of second gaps for separating and concentrating circulating tumor cells.EXAMPLE 2

[0134] In an embodiment, the microfluidic filtration chamber of the present disclosure comprises the characteristics as depicted in Figure 6, wherein the microfluidic filtration chamber comprises: an inlet (1) and an outlet (2) for the entry and exit of the sample, respectively;a central distribution post element (central post element) (9) for distributing the fluid sample; a plurality of first post elements (3) for prefilter the sample; a plurality of second post elements (4) for filtering the sample; a substrate and a cover for sealing the microfluidic filter chamber; wherein: the inlet and outlet have a height of 200 pm; the central distribution post element (9) has a lozenge (diamond) shape and is positioned downstream of the inlet and upstream of the plurality of first post elements; the plurality of first post elements is arranged in a zigzag geometry in respect to each other; the plurality of first post elements is arranged in multiple rows, wherein each row of first post elements is laterally offset with respect to the adjacent upstream and / or downstream rows (plinko distribution); each first post element of the plurality of first post elements has a lozenge (diamond) shape; the plurality of first post elements is interspaced, thereby forming a plurality of first gaps (5), each first gap being formed in between two adjacent first post elements, wherein each first gap has a size of 200 pm; the plurality of second post elements are arranged in a zigzag geometry, forming an angle (8) of 29.83° between each adjacent second post element; each second post element of the plurality of second post elements has a circular cylindrical, wherein the diameter of each second post element is 35 pm; the plurality of second post elements is interspaced, thereby forming a plurality of second gaps (7), each second gap being formed in between two adjacent second post elements, wherein the size of each second gap is 7 pm and each second gap has an aspect ratio between its height and width of 5; the plurality of first post elements and the plurality of second post elements are arranged such that a flow of the sample flowing from the inlet flow channel to the outlet flow channel passes through the plurality of first gaps and the plurality of second gaps for separating and concentrating circulating tumor cells.

[0135] The following table summarizes the results obtained with the device according to Example 2.Comparative data in relation to comparative examples 1-2 are also provided.Table 1. Results obtained with example 2 and comparative examples 1-2.CV: coefficient of variance, calculated by level of flow uniformity.

[0136] Surprisingly, as depicted in Table 1, it was found that the specific structure of the microfluidic filtration chamber of the present disclosure, in particular the design and disposition of the first post elements (lozenge / diamond shape arranged in a zigzag geometry), allow to obtain improved flow uniformity and avoid pressure peaks, as well as improved capture efficiency. As depicted in Figures 9-11, while other structures (comparative examples 1-2) lead to variations in the flow across the microfluidic filtration chamber and flow above 1, the structure of the microfluidic filtration chamber of the present disclosure (Figure 9) provides flow uniformity and a flow below 1; preferably below 0.8.EXAMPLE 3Microfluidic flow device (microfluidic chip)

[0137] The microfluidic flow device according to the present disclosure (example 3) is depicted in Figure 12.

[0138] The microfluidic flow device of the present disclosure was designed to split the blood equally in 8 different modules, and each module comprising 32 microfluidic filter chambers. Each module can process a maximum of 1 ml of whole blood

[0139] The microfluidic flow device (microfluidic chip) of the present disclosure also comprises the following improved characteristics:Position of inlet and outlet: inlet and outlet have been moved to ensure that they are not too close to the border of the device or to the channels (risking leaking), and that they are not too close to the filter area, which could compromise imaging;Blank area between channels: Blank area between channels has been increased, ranging from 50 pm to 400 pm to ensure enough support to the channels and prevent leaking.Channel length: Inlet and outlet channels distribute the flow across the microfluidic filtration chambers. They are adjusted to have the same length, so that there are no pressure differences and there is no preferential path for the sample, so that the flow is evenly distributed across the different chambers of the devices.Connectors / interface: different solutions for the interface tubing / device were explored to ensure a good connection, with no leaks, that sustains the pressure and that is thin enough not to compromise imaging of the filter area. Current solution uses a laser-welded chimney.Device fabrication

[0140] In a preferred embodiment, the microfluidic filtration chamber of the present disclosure is fabricated through injection molding and / or hot embossing, using thermoplastics that are hard and rigid polymers at room temperature, preferably a thermoplastic selected from Cyclic olefin copolymer (COP), Cyclic olefin copolymer (COC), or mixtures thereof.

[0141] In a preferred embodiment for better results, the microfluidic filtration chamber of the present disclosure comprises a chamber height of 30 pm, second post elements with a diameter of 35 um, and second gaps width ranging from 6 -7.5 pm.

[0142] In a preferred embodiment, the ratio height / diameter of the second post elements ranges from 0.7 to 1.1.

[0143] In a preferred embodiment for better results, the first post elements and the second post elements are perfectly vertical to ensure that the width of the first gap and the width of the second gap is always the same across the height of each post element.

[0144] In a preferred embodiment for better results, the inlet and outlet have a diameter ranging from 150-250 pm; preferably 200 pm, to provide adequate flow of the fluid sample and avoid pressure peaks, which could compromise the sealing and ultimately leaking.Device priming and coating

[0145] In a preferred embodiment, surface priming and coating are important in both cell isolation and retrieval, as it is crucial to prevent cell attachment to maximize cell purity and recovery.

[0146] Therefore, in a preferred embodiment, the surfaces of the post elements can be coated with a surfactant, such as non-ionic copolymer surfactants (e.g., Pluronic F-127) to facilitate smooth cell transition from the inlet to the outlet. This treatment prevents unwanted cell adhesion, simplifies the isolation of desired cells, and makes it easier to harvest trapped cells by reversing the flow through the filter.

[0147] Devices are connected to a pump and filled with diluted ethanol to wet the channels, then devices are rinsed with PBS, and later treated with 1% Pluronic F-127 to avoid unspecific attachment of cells onto the channel surface.Sample analysis

[0148] Whole blood samples (7.5 mL) are collected in EDTA-coated tubes, shipped at room temperature and processed a maximum of 24h after collection. Samples are not fixed to maintain cell viability and to keep the mechanical properties of the cells intact.

[0149] The whole blood samples are injected in the microfluidic filtration chamber of the present disclosure at the optimized flow rate (depending on cancer type, preferably between 80 and 140 pL / min) using a pump. Trapped cells are rinsed with 2% BSA.

[0150] According to the downstream analysis selected, cells can be recovered by switching inlets and outlets, or fixed and stained for immunocytochemistry. Cells can also be lysed, and genetic material recovered for molecular analysis.

[0151] For immunocytochemistry, cells are fixed, permeabilized and stained with a cocktail of antibodies.Imaging and analysis

[0152] In a preferred embodiment, following spiked sample processing, a fluorescence microscopy analysis of the trapped cells can be performed using a fluorescence microscope. Target cells are identified by the presence of positive markers and absence of exclusion markers.

[0153] Efficiency is calculated in model experiments comparing number of CTCs trapped versus number of cells spiked. Sample purity is calculated comparing number of CTCs trapped against total number of cells trapped.

[0154] Study of protein expression in the CTCs is possible by adding labelled antibodies that are specific for the protein of interest.

[0155] For proper analysis of the cells, relatively high-resolution microscopy is necessary. For this, it is important that cells are trapped in a defined area (so that image scan is circumscribed to a small area), that the substrate is optically transparent (to allow brightfield and fluorescence imaging), and that the substrate is thin, so that magnification is at least 10-20x. It is also important that the substrate is flat, so that roughness does not interfere in the imaging. Higher magnification could be required for detailed single-cell analysis. It is also important that cells are in the same (or very similar) focal plane, so that the images obtained capture the cell in focus and that the scanning

[0156] In a preferred embodiment, the microfluidic filtration chamber of the present disclosure comprises a dimension of 75 x 25 mm and allows multicolor microscopy (brightfield and 4 fluorescencechannels), at 20x magnification (and higher), and a multilayer z-scan over the 30pm chamber height for each field of view, of the whole 2 filter areas in 30-60 min automatically.

[0157] As used in the specification and claims, the singular forms "a", "an" and "the" include plural references unless the context clearly dictates otherwise. For example, the term "a sample" includes a plurality of samples, including mixtures thereof.

[0158] Whenever the term "at least," "greater than," or "greater than or equal to" precedes the first numerical value in a series of two or more numerical values, the term "at least," "greater than" or "greater than or equal to" applies to each of the numerical values in that series of numerical values. For example, greater than or equal to 1, 2, or 3 is equivalent to greater than or equal to 1, greater than or equal to 2, or greater than or equal to 3.

[0159] The terms "determining," "measuring," "evaluating," "assessing," "assaying," and "analyzing" are often used interchangeably herein to refer to forms of measurement. The terms include determining if an element is present or not (for example, detection). These terms can include quantitative, qualitative or quantitative and qualitative determinations. Assessing can be relative or absolute. "Detecting the presence of" can include determining the amount of something present in addition to determining whether it is present or absent depending on the context.

[0160] Where ranges are given, endpoints are included. Furthermore, it is to be understood that unless otherwise indicated or otherwise evident from the context and / or the understanding of one of ordinary skill in the art, values that are expressed as ranges can assume any specific value within the stated ranges in different embodiments of the invention, to the tenth of the unit of the lower limit of the range, unless the context clearly dictates otherwise. It is also to be understood that unless otherwise indicated or otherwise evident from the context and / or the understanding of one of ordinary skill in the art, values expressed as ranges can assume any subrange within the given range, wherein the endpoints of the subrange are expressed to the same degree of accuracy as the tenth of the unit of the lower limit of the range.

[0161] The term "comprising" whenever used in this document is intended to indicate the presence of stated features, integers, steps, components, but not to preclude the presence or addition of one or more other features, integers, steps, components or groups thereof.

[0162] The disclosure should not be seen in any way restricted to the embodiments described and a person with ordinary skill in the art will foresee many possibilities to modifications thereof. The abovedescribed embodiments are combinable.

[0163] The following dependent claims further set out particular embodiments of the disclosure.

Claims

C L A I M S1. Microfluidic filtration chamber for separating and concentrating tumor cells from a fluidic sample comprising: an inlet and an outlet for the entry and exit of the sample; a plurality of first post elements for prefiltering the sample; a plurality of second post elements for filtering the sample; wherein: the plurality of first post elements is arranged in a zigzag geometry between adjacent first post elements; the plurality of first post elements is interspaced, thereby forming a plurality of first gaps, each first gap being formed in between two adjacent first post elements; the plurality of second post elements is interspaced, thereby forming a plurality of second gaps, each second gap being formed in between two adjacent second post elements; the plurality of first post elements and the plurality of second post elements is arranged such that the flow of the sample flowing from the inlet flow channel to the outlet flow channel passes through the plurality of first gaps and through the plurality of second gaps for separating and concentrating tumor cells.

2. Microfluidic filtration chamber according to the previous claim wherein each the plurality of first post elements is arranged in multiple rows.

3. Microfluidic filtration chamber according to the previous claim wherein each row of first post elements is laterally offset with respect to the adjacent upstream and / or downstream rows.

4. Microfluidic filtration chamber according to any of the previous claims wherein the first post elements are rhombus shaped.

5. Microfluidic filtration chamber according to the previous claim the rhombus shaped of the first post elements has a horizontal diagonal ranging from 300-400 pm and a vertical diagonal ranging from 150-250 pm.

6. Microfluidic filtration chamber according to any of the previous claims further comprising a central post element for distributing the fluid sample.

7. Microfluidic filtration chamber according to the previous claim wherein the central post element has a rhombus shape.

8. Microfluidic filtration chamber according to the previous claim the rhombus shaped the central post element has a horizontal diagonal ranging from 1000-900 pm and a vertical diagonal ranging from 450-550 pm.

9. Microfluidic filtration chamber according to any of the previous claims wherein each second post element of the plurality of second post elements has a cylindrical shape.

10. Microfluidic filtration chamber according to any of the previous claims wherein the plurality of second post elements is arranged in a zigzag geometry.

11. Microfluidic filtration chamber according to any of the previous claims wherein the plurality of second post elements arranged in a zigzag geometry forms an angle ranging from 28°-32° between each adjacent second post element.

12. Microfluidic filtration chamber according to any of the previous claims wherein the diameter of each second post element ranges from 28-40 pm; preferably 28-35 pm; more preferably 35 pm.

13. Microfluidic filtration chamber according to any of the previous claims wherein the width of each second gap ranges from 6 to 7.5 pm; preferably 6.5 pm.

14. Microfluidic filtration chamber according to any of the previous claims wherein the width of each first gap ranges from 80 to 200 pm; preferably 200 pm.

15. Microfluidic filtration chamber according to any of the previous claims wherein the aspect ratio (ratio between their height and width) of each first gap ranges from 1:2 to 1:7.

16. Microfluidic filtration chamber according to any of the previous claims wherein the plurality of first post elements is arranged in a zigzag geometry between adjacent first post elements, forming an angle ranging from 30 to 60 degrees between adjacent first post elements.

17. Microfluidic filtration chamber according to any of the previous claims wherein each second gap has an aspect ratio between its height and width ranging from 3.7 to 5.8.

18. Microfluidic filtration chamber according to any of the previous claims wherein:the plurality of second post elements is arranged in linear geometry, forming an angle of 90° between each adjacent second post element; the width of each second post element ranges from 28-35 pm; each second gap ranges from 6 to 7.5 pm.

19. Microfluidic filtration chamber according to any of the previous claims wherein the plurality of first post elements and the plurality of second post elements are arranged such that a flow of the sample flowing from the inlet flow channel to the outlet flow channel passes through the plurality of first gaps and the plurality of second gaps.

20. Microfluidic filtration chamber according to any of the previous claims wherein the material of the microfluidic filtration chamber comprises a material selected from the group consisting of cyclo olefin polymer, cyclic olefin copolymer, acrylic, polyamide, or mixtures thereof; preferably is a material selected from the group consisting of cyclo olefin polymer, cyclic olefin copolymer, acrylic, polyamide, or mixtures thereof.

21. Microfluidic filtration chamber according to the previous claim wherein the material of the microfluidic filtration chamber is a cyclo olefin polymer.

22. Microfluidic filtration chamber according to any of the previous claims further comprising a substrate and a cover for sealing the microfluidic filter chamber.

23. Microfluidic filtration chamber according to any of the previous claims wherein the first post elements are disposed in a plinko distribution.

24. Microfluidic filtration chamber according to any of the previous claims wherein each first post element of the plurality of first post elements has a lozenge shape.

25. Microfluidic filtration chamber according to any of the previous claims wherein the height of the inlet and / or the height of the outlet ranges from 150-400 pm; preferably 190-210 pm; more preferably is 200 pm.

26. Microfluidic filtration chamber according to any of the previous claims wherein each first post element and each second post element is arranged in between the substrate and the cover.

27. Microfluidic filtration chamber according to any of the previous claims wherein each first post element has a height ranging from 29-31 pm; preferably 30 pm.

28. Microfluidic filtration chamber according to any of the previous claims wherein each second post element has a height ranging from 29-31 pm; preferably 30 pm.

29. Microfluidic filtration chamber according to any of the previous claims wherein the angle formed between two adjacent second post elements ranges from 29 to 31 °; preferably ranges from 29° to 30 °; more preferably is 29.83 °.

30. Microfluidic filtration chamber according to any of the previous claims wherein the diameter of each second post element ranges from 34-36 pm; preferably is 35 pm.

31. Microfluidic filtration chamber according to any of the previous claims wherein the substrate and / or the cover is transparent.

32. Microfluidic filtration chamber according to any of the previous claims wherein the fluidic sample is a body fluid sample; preferably a blood sample.

33. Microfluidic flow device for separating and concentrating a target molecule or cell from a fluidic sample comprising a plurality of microfluidic filtration chambers according to any of the previous claims, an inlet and an outlet for the entry and exit of the sample; an inlet array of channels for flow the sample through the plurality of microfluidic filtration chambers, wherein said inlet channel is connected to at least two new channels to split the sample, with each of these channels subsequently connected to another set of two new channels, progressively decreasing the volume of the flow sample arriving at the chambers, thereby allowing the separation and concentration of the fluidic sample to identify a target molecule or cell.

34. Microfluidic flow device according to the previous claim wherein the outlet array channels have a configuration substantially identical to the inlet array channels.

35. Microfluidic flow device according to any of the previous claims 33-34 wherein the depth of each inlet channel is superior to the depth of each of the plurality of microfluidic filtration chambers.

36. Microfluidic flow device according to any of the previous claims 33-35 wherein the depth of each inlet channel is at least 4 times superior to the depth of each of the plurality of microfluidic filtration chambers.

37. Microfluidic flow device according to any of the previous claims 33-36 wherein the depth of each inlet channel is 4- 7 times superior to the depth of each of the plurality of microfluidic filtration chambers.

38. Microfluidic flow device according to any of the previous claims 33-37 wherein the depth of each inlet channel is 6-7 times superior to the depth of each of the plurality of microfluidic filtration chambers.

39. Microfluidic flow device according to any of the previous claims 33-38 wherein the height of the inlet channel and the height of the of the outlet channel ranges from 150-400 pm, preferably 200 pm.

40. Microfluidic flow device according to any of the previous claims 33-39 wherein the height of each channel of the inlet array of channels ranges from 150-400 pm, preferably is 200 pm.

41. Microfluidic flow device according to any of the previous claims 33-40 comprising at least 4 microfluidic filtration chambers; preferably at least 8 microfluidic filtration chambers; more preferably at least 32 microfluidic filtration chambers; even more preferably 32 microfluidic filtration chambers.

42. Microfluidic filtration chamber according to any of the previous claims 1-32 or microfluidic flow device according to any of the previous claims 33-41 wherein the microfluidic filtration chamber is a single-use microfluidic filtration chamber and / or the microfluidic flow device is a single-use microfluidic flow device.

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