Patent Description:
<CIT> discloses a discharge device (<NUM>) includes a discharge head (<NUM>) that discharges a liquid material (<NUM>) from a nozzle opening (<NUM>) connected to the cavity (<NUM>) by varying the internal pressure of a cavity (<NUM>) using an actuator (<NUM>). The discharge head (<NUM>) includes a monitoring portion (<NUM>) provided between the cavity (<NUM>) and the nozzle opening (<NUM>) and the discharge device (<NUM>) further includes a detection apparatus (<NUM>) that detects the number and/or form of the particle-like bodies (<NUM>) included in the liquid material (<NUM>) in the monitoring portion (<NUM>) of the discharge head (<NUM>) and a control unit (<NUM>) that drives the actuator (<NUM>) according to the detection result (7a) of the detection unit (<NUM>) to change the state of the particle-like bodies (<NUM>) included in the liquid material (<NUM>) of the monitoring portion (<NUM>).

An enrichment process is the first step in conventional research for biological particles, but the conventional enrichment process still requires a significant amount of manpower and has a problem that biological particles can be adhered to a pipe wall that is used in the process. Accordingly, how apparatuses or devices can be used to quickly and accurately implement the enrichment process has been one of the important areas of research and development in the relevant field.

In response to the above-referenced technical inadequacies, the present disclosure provides a biological particle enrichment apparatus and a pico-droplet generator thereof to effectively improve on the issues associated with conventional enrichment processes. According to the invention, a pico-droplet generator according to claim <NUM> and a biological particle enrichment apparatus according to claim <NUM> are provided. Some examples of said apparatus are defined in claims <NUM> to <NUM>.

In order to solve the above-mentioned problems, one of the technical aspects adopted by the present disclosure is to provide a biological particle enrichment apparatus, which includes a pico-droplet generator and a biochip corresponding in position to the pico-droplet generator. The pico-droplet generator is configured to output a pico-droplet from a liquid specimen. The pico-droplet generator includes a container, a hollow needle, a first piezoelectric member, and a second piezoelectric member. The container is configured to receive the liquid specimen having a plurality of biological particles, and the container has a bottom side and a surrounding lateral side that is connected to the bottom side. The hollow needle includes a connection end and a free end that is opposite to the connection end. The connection end of the hollow needle is connected to the bottom side of the container so as to establish a fluid communication between the hollow needle and the container. Moreover, an inner diameter of the container is within a range from <NUM> times to <NUM> times of an inner diameter of the hollow needle. The first piezoelectric member has a ring-shaped arrangement and is disposed on the surrounding lateral side of the container. The first piezoelectric member is configured to enable the biological particles in the container to be moved along a direction away from the surrounding lateral side by vibrating the container. The second piezoelectric member is disposed on an outer surface of the hollow needle. The second piezoelectric member is configured to squeeze the hollow needle, so that the liquid specimen flows outwardly and passes through the free end to form the pico-droplet. The biological particles include at least one target biological particle, and the pico-droplet having the at least one target biological particle is defined as a target pico-droplet. The biochip is configured to carry the target pico-droplet and to capture the at least one target biological particle in the target pico-droplet.

In order to solve the above-mentioned problems, another one of the technical aspects adopted by the present disclosure is to provide a pico-droplet generator of a biological particle enrichment apparatus. The pico-droplet generator includes a container, a hollow needle, a first piezoelectric member, and a second piezoelectric member. The container is configured to receive a liquid specimen having a plurality of biological particles. The container has a bottom side and a surrounding lateral side that is connected to the bottom side. The hollow needle includes a connection end and a free end that is opposite to the connection end. The connection end of the hollow needle is connected to the bottom side of the container so as to establish a fluid communication between the hollow needle and the container. Moreover, an inner diameter of the container is within a range from <NUM> times to <NUM> times of an inner diameter of the hollow needle. The first piezoelectric member has a ring-shaped arrangement and is disposed on the surrounding lateral side of the container. The first piezoelectric member is configured to enable the biological particles in the container to be moved along a direction away from the surrounding lateral side by vibrating the container. The second piezoelectric member is disposed on an outer surface of the hollow needle. The second piezoelectric member is configured to squeeze the hollow needle, so that the liquid specimen flows outwardly and passes through the free end to form a pico-droplet.

Therefore, any one of the biological particle enrichment apparatus and the pico-droplet generator thereof in the present disclosure can be provided to enable the biological particles to be moved toward a center of the container through a vibration of the first piezoelectric member, thereby preventing the biological particles from being adhered to inner walls of the container and completing the enrichment process of the target biological particle.

These and other aspects of the present disclosure will become apparent from the following description of the embodiment taken in conjunction with the following drawings and their captions, although variations and modifications therein may be affected without departing from the scope of the novel concepts of the disclosure.

As used in the description herein and throughout the claims that follow, unless the context clearly dictates otherwise, the meaning of "a," "an" and "the" includes plural reference, and the meaning of "in" includes "in" and "on. " Titles or subtitles can be used herein for the convenience of a reader, which shall have no influence on the scope of the present disclosure.

Numbering terms such as "first," "second" or "third" can be used to describe various components, signals or the like, which are for distinguishing one component/signal from another one only, and are not intended to, nor should be construed to impose any substantive limitations on the components, signals or the like.

Referring to <FIG>, a first embodiment of the present disclosure is provided. As shown in <FIG> and <FIG>, the present embodiment provides a biological particle analysis method S100 and an identifying system <NUM>. The identifying system <NUM> is applied to the biological particle analysis method S100, and the specific configuration of the identifying system <NUM> can be adjusted or changed according to design requirements, but the present disclosure is not limited thereto. For example, in other embodiments of the present disclosure not shown in the drawings, the biological particle analysis method S100 can be implemented through a structure other than the identifying system <NUM>.

In the present embodiment, the identifying system <NUM> includes a multi-fluorescence staining apparatus <NUM>, a pico-droplet generator <NUM>, a camera device <NUM>, and a biochip <NUM> that is provided to connect an operation of the multi-fluorescence staining apparatus <NUM> and an operation of the pico-droplet generator <NUM>. The multi-fluorescence staining apparatus <NUM> includes a staining device <NUM>, a washing device <NUM> arranged adjacent to the staining device <NUM>, and a recording device <NUM> (e.g., a camera) that corresponds in position to the staining device <NUM>, but the present disclosure is not limited thereto.

As shown in <FIG> and <FIG> to <FIG>, the biological particle analysis method S100 in the present embodiment sequentially includes a staining step S110, an analyzing step S120, a capturing step S130, a washing step S140, and a characterization expressing step S150, thereby effectively completing an enrichment process and multiple biological characterization expressions for at least one target biological particle 201a that is selected from a liquid specimen <NUM> having a plurality of biological particles <NUM>. In other words, any method not completely implementing the above steps in sequence is different from the biological particle analysis method S100 of the present embodiment.

As shown in <FIG>, <FIG>, and <FIG>, the staining step S110 is implemented by fluorescence staining the liquid specimen <NUM> through a fluorescence staining process, so that at least one of the biological particles <NUM> becomes a fluorescence color and is defined as the at least one target biological particle 201a. In other words, the biological particles <NUM> are divided into the at least one target biological particle 201a and other non-target biological particles 201b.

In the present embodiment, the liquid specimen <NUM> is received in the specimen container <NUM>, and the fluorescence staining process is implemented to the liquid specimen <NUM> in the specimen container <NUM> through the staining device <NUM> of the multi-fluorescence staining apparatus <NUM>, but the present disclosure is not limited thereto. For example, in other embodiments of the present disclosure not shown in the drawings, the liquid specimen <NUM> in the specimen container <NUM> can be stained in other manners without using the staining device <NUM>. Accordingly, the liquid specimen <NUM> being fluorescence stained is received in the specimen container <NUM>.

Moreover, the liquid specimen <NUM> can be a body fluid from an animal (e.g., e.g., blood, lymph, saliva, ascites, or urine), and the at least one target biological particle 201a can be a specific type of cell, such as circulating tumor cells (CTCs), fetal nucleated red blood cells (FNRBCs), exosome, virus, or bacterium, but the present disclosure is not limited thereto. For example, in other embodiments of the present disclosure not shown in the drawings, the liquid specimen <NUM> can be chosen from plants.

As shown in <FIG> and <FIG> to <FIG>, the analyzing step S120 is implemented by accommodating the liquid specimen <NUM> being fluorescence stained into the pico-droplet generator <NUM>, and using the camera device <NUM> to take a real-time image of the liquid specimen <NUM> in the pico-droplet generator <NUM>. In the present embodiment, the pico-droplet generator <NUM> can be used to receive the liquid specimen <NUM> therein from the specimen container <NUM>, and an immediate position of the at least one target biological particle 201a having the fluorescence color can be obtained by using the camera device <NUM> to take the real-time image.

As shown in <FIG> and <FIG> to <FIG>, the capturing step S130 is implemented by using the pico-droplet generator <NUM> to output a target pico-droplet 202a having the at least one target biological particle 201a onto the biochip <NUM> according to the real-time image. In other words, pico-droplets <NUM> generated from the liquid specimen <NUM> in the pico-droplet generator <NUM> in the present embodiment are divided into the target pico-droplet 202a and an abandoned pico-droplet 202b not having the target biological particle 201a. It should be noted that the pico-droplet <NUM>, which has the non-target biological particle 201b but does not have the target biological particle 201a, is also defined as the abandoned pico-droplet 202b.

Specifically, whether the pico-droplet <NUM> outputted from the pico-droplet generator <NUM> has the at least one target biological particle 201a, which can be confirmed according to the real-time image, so that the pico-droplet <NUM> can be accurately placed onto the biochip <NUM> each time for enabling the at least one target biological particle 201a in the target pico-droplet 202a to be captured by the biochip <NUM>, and the abandoned pico-droplet 202b is placed into an abandoned container <NUM>, thereby effectively completing the enrichment process of the at least one target biological particle 201a.

It should be noted that the biochip <NUM> in the present embodiment is manufactured through a semi-conductor process, and the specific structure of the biochip <NUM> can be adjusted or changed according to design requirements. In order to clearly describe the biochip <NUM>, the following description describes one kind of the biochip <NUM> having a better capturing effect, but the present disclosure is not limited thereto.

Specifically, as shown in <FIG> and <FIG>, the biochip <NUM> is configured to carry the target pico-droplet 202a, and is capable of capturing the at least one target biological particle 201a in the target pico-droplet 202a. The biochip <NUM> includes a bottom layer <NUM>, a plurality of capturing arms <NUM> connected to the bottom layer <NUM> and spaced apart from each other, and a surface modification layer <NUM> that is formed on ends of the capturing arms <NUM>. In the capturing step S <NUM>, the biochip <NUM> is configured to capture the at least one target biological particle 201a through the capturing arms <NUM> and the surface modification layer <NUM>.

Specifically, the surface modification layer <NUM> is preferably an arginylglycylaspartic acid (RGD) peptide layer, thereby facilitating capturing of the at least one target biological particle 201a in a chemical bonding manner, but the present disclosure is not limited thereto. Moreover, as shown in <FIG>, at least two of the capturing arms <NUM> of the biochip <NUM> are elastically swingable with respect to the bottom layer <NUM> and are capable of clamping (or pinching) the at least one target biological particle 201a, thereby facilitating capturing of the at least one target biological particle 201a in a physical manner.

As shown in <FIG>, <FIG>, and <FIG>, the washing step S140 is implemented by removing the fluorescent color of the at least one target biological particle 201a captured by the biochip <NUM> through a washing process. In the present embodiment, the washing process is implemented to the at least one target biological particle 201a captured by the biochip <NUM> through the washing device <NUM> of the multi-fluorescence staining apparatus <NUM>.

As shown in <FIG>, <FIG>, and <FIG>, the characterization expressing step S150 is implemented by fluorescence staining the at least one target biological particle 201a captured by the biochip <NUM> for N number of times through the fluorescence staining process and the washing process, and using the recording device <NUM> to obtain a plurality of fluorescence images respectively corresponding to N kinds of biological characterization expressions. In other words, each fluorescence staining of the at least one target biological particle 201a is related to (or corresponds to) one of the kinds of the biological characterization expressions. In the present embodiment, N is a positive integer within a range from <NUM> to <NUM>, and N is preferably within a range from <NUM> to <NUM>, but the present disclosure is not limited thereto.

In addition, the biochip <NUM> can be moved among different devices (e.g., the staining device <NUM>, the washing device <NUM>, and the carrying platform <NUM>) through manpower or automation (e.g., by a robotic arm), but the present disclosure is not limited thereto.

In summary, the capturing step S130 of the biological particle analysis method S100 in the present embodiment can be implemented to achieve the enrichment effect by being cooperated with the staining step S110 and the analyzing step S <NUM>. Moreover, the biological particle analysis method S <NUM> in the present embodiment can be implemented to effectively connect the capturing step S <NUM>, the washing step S <NUM>, and the characterization expressing step S150 by using the biochip <NUM> to firmly capture the at least one target biological particle 201a. Accordingly, the fluorescence images respectively corresponding to the N kinds of biological characterization expressions can be obtained from the liquid specimen <NUM> through the biological particle analysis method S100, thereby facilitating any evaluation and determination of the at least one target biological particle 201a. In addition, the biological particle analysis method S100 can be implemented such that the fluorescence images are overlapped with each other, thereby obtaining a biological characterization of the at least one target biological particle 201a.

As shown in <FIG>, <FIG>, and <FIG>, the above description describes the steps S110 - S150 of the biological particle analysis method S100 provided by the present embodiment, and in order to more clearly illustratee the present embodiment, a biological particle enrichment apparatus <NUM> that is cooperated with the multi-fluorescence staining apparatus <NUM> for jointly implementing the biological particle analysis method S <NUM> is further described, but the present disclosure is not limited thereto. For example, in other embodiments of the present disclosure not shown in the drawings, the biological particle enrichment apparatus <NUM> can be independently used (e.g., sold) or can be used in cooperation with other components.

In the present embodiment, the biological particle enrichment apparatus <NUM> includes the pico-droplet generator <NUM>, the biochip <NUM> and the camera device <NUM> both corresponding in position to the pico-droplet generator <NUM>, a controlling device <NUM> electrically coupled to the pico-droplet generator <NUM> and the camera device <NUM>, a carrying platform <NUM> being capable of carrying the biochip <NUM> and corresponding in position to the pico-droplet generator <NUM>, the specimen container <NUM> and the abandoned liquid container <NUM> both disposed on the carrying platform <NUM>, and a pressure balance mechanism <NUM> that is connected to the pico-droplet generator <NUM>, but the present disclosure is not limited thereto.

For example, in other embodiments of the present disclosure not shown in the drawings, types and a quantity of interior components of the biological particle enrichment apparatus <NUM> can be adjusted or changed according to design requirements, and the pico-droplet generator <NUM> can be independently used (e.g., sold) or can be used in cooperation with other components (e.g., the biochip <NUM>).

The following description describes the structure and connection relationship of each component of the biological particle enrichment apparatus <NUM>. In addition, part of the components (e.g., the biochip <NUM>) of the biological particle enrichment apparatus <NUM> is described in the above description and will be omitted herein for the sake of brevity.

The pico-droplet generator <NUM> includes a container <NUM>, a hollow needle <NUM> being in fluid communication with the container <NUM>, a first piezoelectric member <NUM> disposed on the container <NUM>, and a second piezoelectric member <NUM> that is disposed on the hollow needle <NUM>. The container <NUM> is configured to receive the liquid specimen <NUM> and to transmit the liquid specimen <NUM> into the hollow needle <NUM>, so that the hollow needle <NUM> can receive the liquid specimen <NUM> therein.

Specifically, the container <NUM> includes a bottom side <NUM> and a surrounding lateral side <NUM> that is connected to the bottom side <NUM>. The hollow needle <NUM> includes a connection end <NUM> and a free end <NUM> that is opposite to the connection end <NUM>. The connection end <NUM> of the hollow needle <NUM> is (perpendicularly) connected to the bottom side <NUM> of the container <NUM> so as to establish a fluid communication between the hollow needle <NUM> and the container <NUM>. Moreover, an inner diameter D211 of the container <NUM> is within a range from <NUM> times to <NUM> times of an inner diameter D212 of the hollow needle <NUM>. Specifically, the inner diameter D212 of the hollow needle <NUM> in the present embodiment that is measured is a portion other than the free end <NUM> and is preferably within a range from <NUM> to <NUM>, and the free end <NUM> of the hollow needle <NUM> in the present embodiment has an inner diameter D2122 being within a range from <NUM> to <NUM>, but the present disclosure is not limited thereto.

The first piezoelectric member <NUM> has a ring-shaped arrangement and is (annularly) disposed on the surrounding lateral side <NUM> of the container <NUM>, and the first piezoelectric member <NUM> in the present embodiment can be referred to as a top piezoelectric member <NUM>. The first piezoelectric member <NUM> is configured to vibrate the container <NUM> for enabling the biological particles <NUM> in the container <NUM> to be moved along a direction away from the surrounding lateral side <NUM>. In other words, the first piezoelectric member <NUM> is configured to vibrate the container <NUM> for enabling the biological particles <NUM> in the container <NUM> to be arranged along a predetermined path.

Specifically, the predetermined path is preferably located along a central axis L of the hollow needle <NUM> (and a direction of gravity). Along a direction parallel to the central axis L, the first piezoelectric member <NUM> is spaced apart from the connection end <NUM> by a distance D213 that is within a range from <NUM> to <NUM>. Moreover, the first piezoelectric member <NUM> is connected to and covers <NUM>% to <NUM>% of an area of the surrounding lateral side <NUM> of the container <NUM>. In addition, according to design requirements, the first piezoelectric member <NUM> can be a single one-piece structure having an annular shape or can be a structure having multiple components in an annular arrangement.

The second piezoelectric member <NUM> is disposed on an outer surface of the hollow needle <NUM>, and the second piezoelectric member <NUM> in the present embodiment can be referred to as a bottom piezoelectric member <NUM>. The second piezoelectric member <NUM> is configured to squeeze the hollow needle <NUM>, so that the liquid specimen <NUM> flows outwardly and passes through the free end <NUM> to form the pico-droplet <NUM>. The pico-droplet <NUM> generated by the pico-droplet generator <NUM> can be further defined as a target pico-droplet 202a having the at least one target biological particle 201a or an abandoned pico-droplet 202b not having the target biological particle 201a.

Specifically, along a direction parallel to the central axis L, the second piezoelectric member <NUM> is spaced apart from the free end <NUM> by a distance D214 that is within a range from <NUM> to <NUM>. The second piezoelectric member <NUM> is connected to and covers <NUM>% to <NUM>% of an area of the outer surface of the hollow needle <NUM>. In addition, according to design requirements, the second piezoelectric member <NUM> can be a single one-piece structure having an annular shape or can be a structure having multiple components in an annular arrangement.

Furthermore, the pressure balance mechanism <NUM> is connected to the container <NUM>, and the pressure balance mechanism <NUM> is configured to enable the liquid specimen <NUM> in the container <NUM> and the hollow needle <NUM> to be maintained at a predetermined pressure. The pressure balance mechanism <NUM> in the present embodiment is described as follows, but the present disclosure is not limited thereto. The pressure balance mechanism <NUM> includes an air pump <NUM>, a switch <NUM> connected to the air pump <NUM>, a pressure balance bottle <NUM> being in fluid communication with the air pump <NUM> and the switch <NUM>, and a liquid injection bottle <NUM> that is in fluid communication with the switch <NUM> and the container <NUM>.

The biochip <NUM>, the specimen container <NUM>, and the abandoned liquid container <NUM> are disposed on the carrying platform <NUM>, and the carrying platform <NUM> and the pico-droplet generator <NUM> are relatively movable to each other (e.g., the carrying platform <NUM> can be assembled with a multi-axis movable mechanism). Accordingly, the pico-droplet generator <NUM> (in the analyzing step S120) is moveable relative to the carrying platform <NUM> and is capable of sucking the liquid specimen <NUM> from the specimen container <NUM> through the free end <NUM> of the hollow needle <NUM> (e.g., the liquid specimen <NUM> is sucked into the hollow needle <NUM> and the container <NUM>). Moreover, the pico-droplet generator <NUM> is moveable relative to the carrying platform <NUM> so as to output the target pico-droplet 202a onto the biochip <NUM> and output the abandoned pico-droplet 202b into the abandoned liquid container <NUM>.

The camera device <NUM> corresponds in position to the hollow needle <NUM>, and the camera device <NUM> (in the analyzing step S120) is configured to take a real-time image of the liquid specimen <NUM> in the free end <NUM>. Moreover, the controlling device <NUM> is electrically coupled to the second piezoelectric member <NUM> and the camera device <NUM>. According to the real-time image, the controlling device <NUM> (in the capturing step S130) is configured to drive the second piezoelectric member <NUM> when the at least one target biological particle 201a is located in the free end <NUM>, so that the second piezoelectric member <NUM> is driven to squeeze the hollow needle <NUM> to enable the liquid specimen <NUM> to flow outwardly and pass through the free end <NUM> of the hollow needle <NUM> to form the target pico-droplet 202a.

In summary, the biological particle enrichment apparatus <NUM> of the present embodiment can be provided to enable the biological particles <NUM> to be moved toward a center of the container <NUM> through a vibration of the first piezoelectric member <NUM> (i.e., the top piezoelectric member <NUM>), thereby preventing the biological particles <NUM> from being adhered to inner walls of the container <NUM> and completing the enrichment process of the target biological particle 201a.

Specifically, the biological particle analysis method S100 or the biological particle enrichment apparatus <NUM> in the present embodiment is provided with the cooperation between the camera device <NUM> and the second piezoelectric member <NUM> (i.e., the bottom piezoelectric member <NUM>), so that according to the real-time image of the liquid specimen <NUM> in the free end <NUM>, the second piezoelectric member <NUM> can be driven to form the target pico-droplet 202a when the at least one target biological particle 201a is located in the free end <NUM>, thereby effectively completing the enrichment process of the target biological particle 201a.

In conclusion, the capturing step of the biological particle analysis method in the present disclosure can be implemented to achieve the enrichment effect by being cooperated with the staining step and the analyzing step. Moreover, the biological particle analysis method in the present disclosure can be implemented to effectively connect the capturing step, the washing step, and the characterization expressing step by using the biochip to capture the at least one target biological particle. Accordingly, the fluorescence images respectively corresponding to multiple kinds of biological characterization expressions can be obtained from the liquid specimen through the biological particle analysis method, thereby facilitating any evaluation and determination of the at least one target biological particle.

In addition, the biological particle enrichment apparatus of the present disclosure can be provided to enable the biological particles to be moved toward a center of the container through a vibration of the first piezoelectric member (i.e., the top piezoelectric member), thereby preventing the biological particles from being adhered to inner walls of the container and completing the enrichment process of the target biological particle.

Specifically, the biological particle analysis method or the biological particle enrichment apparatus in the present disclosure is provided with the cooperation between the camera device and the second piezoelectric member (i.e., the bottom piezoelectric member), so that according to the real-time image of the liquid specimen in the free end, the second piezoelectric member can be driven to form the target pico-droplet when the at least one target biological particle is located in the free end, thereby effectively completing the enrichment process of the target biological particle.

Claim 1:
A pico-droplet generator of a biological particle enrichment apparatus, comprising:
a container (<NUM>) configured to receive a liquid specimen (<NUM>) having a plurality of biological particles (<NUM>), wherein the container (<NUM>) has a bottom side (<NUM>) and a surrounding lateral side (<NUM>) that is connected to the bottom side (<NUM>);
a hollow needle (<NUM>) including a connection end (<NUM>) and a free end (<NUM>) that is opposite to the connection end (<NUM>), wherein the connection end (<NUM>) of the hollow needle (<NUM>) is connected to the bottom side (<NUM>) of the container (<NUM>) so as to establish a fluid communication between the hollow needle (<NUM>) and the container (<NUM>), and wherein an inner diameter (D211) of the container (<NUM>) is within a range from <NUM> times to <NUM> times of an inner diameter (D212) of the hollow needle (<NUM>); and
a second piezoelectric member (<NUM>) disposed on an outer surface of the hollow needle (<NUM>), wherein the second piezoelectric member (<NUM>) is configured to squeeze the hollow needle (<NUM>), so that the liquid specimen (<NUM>) flows outwardly and passes through the free end (<NUM>) to form a pico-droplet (<NUM>);
characterized by a first piezoelectric member (<NUM>) having a ring-shaped arrangement and disposed on the surrounding lateral side (<NUM>) of the container (<NUM>), wherein the first piezoelectric member (<NUM>) is configured to enable the biological particles (<NUM>) in the container (<NUM>) to be moved along a direction away from the surrounding lateral side (<NUM>) by vibrating the container (<NUM>).