CN110938530A - Adaptor, device and method for continuously acquiring micro tissue blocks - Google Patents
Adaptor, device and method for continuously acquiring micro tissue blocks Download PDFInfo
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- CN110938530A CN110938530A CN201911257890.3A CN201911257890A CN110938530A CN 110938530 A CN110938530 A CN 110938530A CN 201911257890 A CN201911257890 A CN 201911257890A CN 110938530 A CN110938530 A CN 110938530A
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- C12M33/00—Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus
- C12M33/12—Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus by pressure
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Abstract
The invention provides an adaptor, a device and a method for continuously acquiring a trace tissue block, and relates to the technical field of section sampling. The device comprises a piston rod connecting part, a needle head connecting part and a sampling needle, wherein the sampling needle is connected to the end part of a cylinder in the device through the adapter. Through the device earlier take a sample, after the sample, place collection device under the sampling needle, push away a kind mouth through syringe needle connecting portion lateral wall simultaneously and to sampling needle air feed or supply liquid, need not dismouting sampling needle and can realize pushing away a kind collection voluntarily after the sample is ended, realized taking a sample automatically in succession.
Description
Technical Field
The invention relates to the technical field of section sampling, in particular to an adapter, a device and a method for continuously acquiring a micro tissue block.
Background
In order to obtain spatial and omic information of biological tissues simultaneously, it is necessary to image tissue slices, precisely locate a target tissue region according to the imaging result, and then use a sampling device to obtain tissues in the region for subsequent omic analysis. There are three major types of sampling techniques for tissue sections in the prior art, one is manual poking, one is sampling by negative pressure suction, and the other is sampling by laser cutting. Wherein, the manual mode of stabbing of getting can't accurate control stab the atress when getting, and the sample result is uncontrollable. The laser cutting technology utilizes ultraviolet light or infrared light to cut tissues, so that heat deposition is easily caused, the properties of substances such as RNA (ribonucleic acid), protein and the like in the tissues are influenced, and the conclusion of follow-up omics analysis is influenced. The negative pressure sucking technique can only sample fresh/frozen brain tissue slices, and is difficult to sample tissue slices with excessive adhesion between cells, and target breakage is easily caused in the sampling process.
In the methods, the sample in the sampling needle needs to be removed or the sampling needle needs to be replaced after each sampling is finished, the preparation work needs to be repeated, the consumed time is long, the sampling success rate is low, and the method is very inconvenient to use when continuous batch sampling is needed.
Disclosure of Invention
The invention aims to provide an adapter and a device and a method for continuously acquiring micro tissue blocks, so as to solve the problem that continuous sampling cannot be performed with high success rate in the prior art.
An adapter for connecting a piston rod of an air cylinder and a sampling needle comprises a piston rod connecting part and a needle head connecting part, wherein the piston rod connecting part comprises two clamping blocks which are symmetrically arranged and integrally formed, a rod hole which is in inserting fit with the piston rod is formed between the two clamping blocks, one ends of the two clamping blocks are flexibly connected, and the other ends of the two clamping blocks are opened so as to adjust the size of the rod hole and facilitate the insertion and clamping of the piston rod;
the syringe needle connecting portion demountable installation be in the lower terminal surface of piston rod connecting portion, the lower fixed surface of syringe needle connecting portion has the direction arch, the lateral wall of syringe needle connecting portion seted up with the protruding intercommunication of direction pushes away a kind mouth.
Among the above-mentioned technical scheme, both realized the demountable installation between sample needle and the piston rod through the adaptor, after sampling each time simultaneously, need not to dismantle the sample needle, can directly push out the sample in the sample needle through pushing away the sample mouth to air feed or supply liquid in the sample needle, can realize automatic sampling and collection in succession.
Furthermore, the end of the sampling needle is fixed with a plastic clamping piece, and the sampling needle is matched with the guide bulge in a clamping manner through the plastic clamping piece.
Further, the week side cover of sample needle is equipped with the rubber ball, the joint has connecting device in the direction arch, connecting device's lower extreme radius reduces gradually, the sample needle passes through the rubber ball card is gone into in the connecting device.
Furthermore, an upper through hole is formed in the side wall of the opening end of one clamping block in a penetrating mode, an upper threaded hole coaxial with the upper through hole is formed in the other clamping block, the diameter of the upper through hole is larger than that of the upper threaded hole, an upper bolt penetrates through the upper through hole and then is in threaded connection with the upper threaded hole, and the upper bolt abuts against the side wall of the clamping block provided with the upper through hole.
Furthermore, the edge of the needle head connecting part is provided with a lower through hole in a penetrating manner, the lower end face of the piston rod connecting part is correspondingly provided with a lower threaded hole, the diameter of the lower through hole is larger than that of the lower threaded hole, a lower bolt penetrates through the lower through hole and is in threaded connection with the lower threaded hole, and the lower bolt is abutted against the lower surface provided with the needle head connecting part.
Furthermore, threads are arranged on the periphery of the sample pushing port and are used for being in threaded connection with a pipeline so as to supply air or liquid to the sampling needle, and therefore a sample in the sampling needle is pushed out.
Furthermore, the surfaces of the piston rod connecting part and the needle head connecting part are both fixed with oxidation blackening layers.
Furthermore, the end part of the piston rod, which is in plug-in fit with the rod hole, is sleeved with a raw material belt.
A device suitable for continuously obtaining micro tissue blocks on a tissue slice comprises a portal frame, a three-dimensional translation table fixed on one side of the portal frame, an air cylinder fixed with the three-dimensional translation table, a sample table movably arranged below the air cylinder, a sampling needle and a connector used for connecting an air cylinder piston rod and the sampling needle;
and an imaging device is fixed on the portal frame below the sample table and used for positioning a target area of a sample on the sample table.
A method for continuously obtaining micro-tissue blocks on a tissue slice, comprising the steps of:
s1, mounting the piston rod connecting part at the lower end of the piston rod, and screwing the upper bolt; mounting the sampling needle on the guide projection; mounting the needle head connecting part on the lower surface of the piston rod connecting part, and screwing down the lower bolt;
s2, positioning the sample through the imaging device to move the sample stage according to the position of the target area in the imaging device, so that the target area is positioned right below the sampling needle;
s3, the three-dimensional translation table drives the sampling needle to advance for sampling;
and S4, after sampling, moving the sample platform away, placing a collecting device below the sampling needle, connecting a pipeline for a sample pushing port, and supplying gas or liquid into the sampling needle so as to push the tissues in the sampling needle out of the collecting device.
Drawings
FIG. 1 is a schematic structural view of an adapter;
FIG. 2 is a schematic view of the lower end gas supply configuration of an apparatus suitable for continuous harvesting of micro-tissue pieces on tissue slices;
FIG. 3 is a schematic view of the upper end gas supply configuration of an apparatus suitable for continuous harvesting of micro-tissue pieces on a tissue slice;
FIG. 4 is a graph showing the relationship between the supplied air pressure and the number of times of debugging in the process of debugging the empirical air pressure;
FIG. 5 is a graph showing the analysis of the force applied to the lower end during the sampling process;
FIG. 6 shows the resistance F of the lower air supply pressure and the retraction of the sampling needle during the process of adjusting the empirical air pressuresA graph of relationships between;
FIG. 7 is a graph showing the analysis of the force applied when air is supplied from the upper end during sampling;
FIG. 8 shows the resistance F of the lower air supply pressure and the retraction of the sampling needle during the empirical air pressure debugging processsA graph of relationships between;
FIG. 9 is a diagram showing the result of positioning a liver tissue section by a microscope;
fig. 10 is a graph showing the results of sampling liver tissue sections by four methods.
Wherein, 1, a sample platform; 2. a sampling needle; 3. an adapter; 4. a three-dimensional translation stage; 5. a cylinder; 6. a barometer; 7. an electromagnetic valve; 8. a pressure reducing valve; 9. an imaging device; 10. a gantry; 11. a piston rod connecting portion; 12. a rod hole; 13. an upper through hole; 14. screwing a bolt; 15. a needle head connecting part; 16. a guide projection; 17. pushing a sample port; 18. a lower through hole; 19. and (4) a lower bolt.
Detailed Description
In order to make the technical problems, technical solutions and advantageous effects to be solved by the present invention more clearly apparent, the present invention is further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.
Example one
Referring to fig. 1, for the adaptor for connecting cylinder piston rod and sampling needle that this embodiment disclosed, including piston rod connecting portion 11 and syringe needle connecting portion 15, piston rod connecting portion 11 includes that two symmetries set up and integrated into one piece's clamp splice, and one side that two clamp splices are close to each other all indent into half hole, and two half hole cooperations form a pole hole 12 for with the cooperation of pegging graft of the piston rod of cylinder 5. One end of each of the two clamping blocks is flexibly connected, the other end of each of the two clamping blocks is provided with an opening, and the two clamping blocks can rotate around the flexible connection point, so that the size of the rod hole 12 is adjusted, and the piston rod can be conveniently inserted and clamped.
The rod hole 12 is arranged close to the flexible connecting point, so that the area of the opening ends of the two clamping blocks is increased, and the size of the opening is convenient to adjust; meanwhile, the flexible connection area of the two clamping blocks is reduced, and the rotation is more flexible. Two upper through holes 13 are formed in the side wall of the opening end of one clamping block in a penetrating mode, an upper threaded hole coaxial with the upper through holes 13 is correspondingly formed in the opening end of the other clamping block, and the diameter of each upper through hole 13 is larger than that of each upper threaded hole. An upper bolt 14 penetrates through the upper through hole 13, the upper bolt 14 penetrates through the upper through hole 13 and then is in threaded connection with the upper threaded hole, and the upper bolt 14 abuts against the side wall of the clamping block provided with the upper through hole 13, so that the two clamping blocks are clamped, the rod hole 12 is reduced, and the piston rod is clamped. The end part of the piston rod, which is positioned in the rod hole 12, is sleeved with a raw material belt, and the raw material belt is more closely matched with the rod hole 12 in an inserting manner.
The syringe needle connecting portion 15 is located the below of piston rod connecting portion 11, its central point department is fixed with guide bulge 16 downwards, be cavity in the guide bulge 16, syringe needle connecting portion 15's lateral wall is seted up and is pushed away appearance mouth 17 with guide bulge 16 intercommunication, form "L" type passageway after the intercommunication, the inner wall that pushes away appearance mouth 17 is provided with the screw thread, be used for the connecting tube, after the sample, for pushing away appearance mouth 17 connecting tube, to supplying air/confession liquid in the "L" type passageway, thereby release the sample in the syringe needle 2.
The end of the sampling needle 2 can be fixed with a plastic clamping piece, and the sampling needle 2 is directly clamped with the guide bulge 16 through the plastic clamping piece. The tip of sampling needle 2 can also overlap and establish the rubber ball, and the joint has connecting device in the protruding 16 inside callipers of direction, and connecting device becomes the round platform form of inversion, and its lower extreme radius reduces gradually, and the rubber ball card of 2 tip of sampling needle is gone into in connecting device, and when the connecting device joint in the protruding 16 of direction to the extrusion rubber ball enters into connecting device's lower extreme, presss from both sides the rubber ball tightly, accomplishes the protruding 16 and the installation of sampling needle 2 of direction.
The needle head connecting portion 15 is in a cuboid shape, four corners of the needle head connecting portion all penetrate through the upper through hole 13, the lower end face four corners of the piston rod connecting portion 11 are correspondingly provided with lower threaded holes, the diameter of each lower through hole 18 is larger than that of each lower threaded hole, a lower bolt 19 penetrates through each lower through hole 18 and then is in threaded connection with each lower threaded hole, and each lower bolt 19 abuts against the lower surface of the needle head connecting portion 15. Since the diameter of the lower through hole 18 is slightly larger than the lower threaded hole, when the two clamping blocks of the piston rod connecting part 11 are rotated, the lower bolt 19 can still pass through the lower through hole 18 to be aligned in the lower threaded hole, thereby mounting the needle head connecting part 15 on the lower surface of the piston rod connecting part 11.
The piston rod connecting part 11 and the needle head connecting part 15 are made of aluminum alloy, and oxidation blackening treatment is carried out on the surfaces of the piston rod connecting part and the needle head connecting part to form an oxidation blackening layer so as to avoid stray light interference generated when a microscope is used for observation or imaging in the sampling process. The oxidation blackening layer covers all the outer surfaces as well as the inner surfaces of the piston rod connection part 11 and the needle tip connection part 15.
Example two
Referring to fig. 2, the apparatus for continuously obtaining micro tissue blocks on a tissue slice disclosed in this embodiment includes a gantry 10, a sample stage 1, a sampling needle 2, a cylinder 5, and a three-dimensional translation stage 4. Three-dimensional translation platform 4 is fixed in one side of portal frame 10, and cylinder 5 is fixed at the lateral wall of three-dimensional translation platform 4, drives cylinder 5 longitudinal movement through three-dimensional translation platform 4, and the bottom joint of cylinder 5 piston rod has adaptor 3 disclosed in embodiment one, and adaptor 3 includes piston rod connecting portion 11 and syringe needle connecting portion 15, and the piston rod end portion at cylinder 5 is established to piston rod connecting portion 11 cover, and the lower extreme at syringe needle connecting portion 15 is installed to sampling needle 2.
The sampling needle 2 is driven to flexibly advance or retract by the extension and retraction of the piston rod in the cylinder 5. The sampling needle 2 is hollow, can be a stainless steel sampling needle 2 with the inner diameter range of 60-600 μm and the outer diameter range of 190-910 μm, and a cluster of cells containing a target cell is arranged in the sampling needle 2 during sampling; or the glass sampling needle 2 with the tip inner diameter of 10-40 μm is formed by drawing a silicon boric acid glass tube with the inner diameter of 0.45-0.89 mm and the outer diameter of 1.5mm, the drawing length is different according to the required inner diameter, and when the glass sampling needle 2 is used for sampling, each sampling needle 2 is internally provided with a single target cell. The side wall of the adaptor 3 is provided with a sample pushing port 17, the sample pushing port 17 is communicated with the sampling needle 2, and after sampling is finished, a sample in the sampling needle 2 is pushed out by air supply/liquid supply to the sample pushing port 17.
The sample stage 1 is a two-dimensional moving platform and can move in a transverse plane, samples are collected through a glass slide or an adhesive tape and placed on the sample stage 1, and the positions of the samples are adjusted through movement of the sample stage 1. An imaging device 9 is fixed on the portal frame 10 below the sample table 1, and the sample is observed or imaged through the imaging device 9, so that the target area of the sample is conveniently positioned. The imaging device 9 may be a microscope, and may also be a point scanning imaging optical path, a line scanning imaging optical path, a structured light imaging optical path, a light sheet imaging optical path, a two-photon imaging optical path, and the like in the prior art.
The piston rod of cylinder 5 divides the inside cavity of cylinder 5 into upper end and lower extreme, and the upper end and the lower extreme of cylinder 5 all carry out the air feed through the trachea, and barometer 6 is all installed to the trachea of upper end and lower extreme. When the lower end of the air cylinder 5 is supplied with air in the sampling process, the air pipe at the upper end is provided with the electromagnetic valve 7, the air pipe at the lower end is provided with the pressure reducing valve 8, the electromagnetic valve 7 is closed, the upper end of the air cylinder 5 stops supplying air, the pressure reducing valve 8 is adjusted, the air supply to the lower end of the air cylinder 5 is realized, and the condition of air supply pressure is observed through the barometer 6 at the lower end.
Referring to fig. 3, when air is supplied to the upper end of the air cylinder 5 in the sampling process, the pressure reducing valve 8 is installed on the air pipe at the upper end, the electromagnetic valve 7 is installed on the air pipe at the lower end, the electromagnetic valve 7 at the lower end is kept normally closed, the air supply to the lower end of the air cylinder 5 is stopped, the pressure reducing valve 8 is adjusted, the air supply to the upper end of the air cylinder 5 is realized, and the condition of the air supply pressure is. The upper end and the lower end of the air cylinder 5 are supplied with air and adjusted through the matching of the electromagnetic valve 7 and the pressure reducing valve 8, and the air pressure of the supplied air can be visually seen by the air pressure meter 6.
EXAMPLE III
In the method for continuously obtaining micro tissue blocks on a tissue section disclosed in this embodiment, the imaging device 9 is a microscope, and the sampling needle 2 is a stainless steel sampling needle, and the method includes the following steps:
s101, mounting: sleeving a raw material belt on the peripheral side of the piston rod, then inserting the raw material belt into a rod hole 12 of a piston rod connecting part 11, and screwing up a bolt 14 to enable two clamping blocks of the piston rod connecting part 11 to clamp the piston rod; the sampling needle 2 is clamped on the guide bulge 16 through a plastic clamping piece; the needle attachment portion 15 is aligned with the lower end surface of the piston rod attachment portion 11, and then the lower bolt 19 is tightened to complete the attachment of the sampling needle 2 to the piston rod.
S102, preparing: referring to fig. 2, an electromagnetic valve 7 is installed on an air pipe at the upper end of an air cylinder 5, a pressure reducing valve 8 is installed on an air pipe at the lower end of the air cylinder 5, the electromagnetic valve 7 is opened, air is supplied to the upper end of the air cylinder 5, and a piston rod is located at the lowest point in the air cylinder 5; closing the electromagnetic valve 7, adjusting the pressure reducing valve 8 and observing the piston rod of the air cylinder 5, and keeping the air supply at the lower end of the air cylinder 5 at a fixed value so as to keep the piston rod balanced and have a retraction trend, wherein the air supply pressure at the lower end is an initial value;
s103, sampling: the three-dimensional translation table 4 drives the air cylinder 5 to approach a sample at a constant speed, air supply of the air cylinder 5 is kept constant, trial sampling is carried out on a non-target area at the edge of the sample, air supply is carried out on the upper end of the air cylinder 5 for one time after sampling is finished so that the piston rod returns to the lowest point, and the non-target area after sampling testing is observed through microscope imaging so as to judge whether the trial sampling is successful;
s104, debugging the empirical air pressure: the change of the air supply along with the number of times of debugging in the whole debugging process is shown in fig. 4, starting from the initial value in S102, the air supply pressure at the lower end of the air cylinder 5 is changed from large to small in the device shown in fig. 2 by adjusting the pressure reducing valve 8, the trial sampling is repeated, the air supply pressure at the lower end is reduced by 5kPa before each sampling, the air supply pressure at the lower end is kept constant in each trial sampling process, the air supply pressure is read by the barometer 6 at the lower end, and the force generated by the air supply pressure at the lower end of the piston rod of the air cylinder 5 is Fp. Referring to fig. 5 (a), before the sampling needle 2 enters the sample, the force Fp is still balanced with the gravity G and the static friction force f of the piston rod: at this time, the sampling needle 2 advances at a constant speed and remains stationary relative to the three-dimensional translation stage 4.
Referring to fig. 5 (b), when the sampling needle 2 enters the sample, it is subjected to a resistance force Fs，FsWith the sampling needle 2 getting deeper and progressively larger in the sample, the plunger rod has a tendency to move back, i.e. Fs+ Fp ═ G + F, with static friction down and increasing gradually to a maximum until Fs+Fp≤G+fmaxThe sampling needle 2 moves forward at a constant speed all the time and keeps static relative to the three-dimensional translation stage 4.
Referring to FIG. 5 (c), when Fs+Fp≥G+fmaxThereafter, the speed of the sampling needle 2 within the sample is rapidly reduced to zero, the advance is stopped, and then the reverse acceleration is performed, and the retraction is started, as shown in fig. 5 (d), FsGradually decreases as the plunger rod is retracted until it is zero after leaving the sample.
Repeating the sampling process, observing the result after sampling by a microscope, and taking a sample under a cylinderThe size of the end air supply and the resistance F of the sampling needlesThreshold value (i.e. F at the moment of retraction)s) The relationship between them is shown in FIG. 6, and the lower the supplied air pressure at the lower end, the lower the F of the sampling needlesThe larger the threshold value of (A), FsMay fail when the threshold value of F is too smallsMay bend when the threshold value of (a) is too large. Therefore, when sampling succeeds, the supplied air pressure indicated by the reading of the lower end barometer 6 is taken as the maximum value of the empirical air pressure; after sampling is successful, the air supply pressure at the lower end is still continuously reduced until the human eye observes that the sampling needle 2 is bent or the microscope observes that the sampling needle 2 cuts open the sample, which indicates that the sampling is failed, and the air supply pressure indicated by the reading of the barometer 6 at the lower end is taken as the minimum value of the empirical air pressure.
When the air pressure of the lower air supply is reduced to zero and sampling is not successful, the lower air supply is stopped, then the air pipes at the upper end and the lower end of the air cylinder 5 are replaced, the electromagnetic valve 7 is connected to the lower end of the air cylinder 5, the pressure reducing valve 8 is replaced to the upper end of the air cylinder 5, and the replaced device is shown in fig. 3. The electromagnetic valve 7 keeps normal close, the lower end does not supply air, the pressure reducing valve 8 is adjusted to supply air to the upper end of the air cylinder 5, and the air supply pressure condition of the upper end is obtained through the reading of the air pressure meter 6 connected with the upper end. From zero, the air supply pressure at the upper end of the air cylinder 5 is gradually increased, the trial sampling is repeatedly carried out, the air supply pressure at the upper end is increased by 5kPa before each sampling, the air supply pressure at the upper end is kept constant in each sampling process, the force generated by the air supply pressure at the upper end on a piston rod of the air cylinder 5 is Fp, the static friction force F of the piston rod is upward, and the supporting force F of the piston rod on the bottom of the air cylinder 5 is FdReferring to fig. 7 (a), before the sampling needle 2 enters the sample, the piston rod is balanced: fdAt this time, the sampling needle 2 moves forward at a constant speed and remains stationary relative to the three-dimensional translation stage 4.
Referring to fig. 7 (b), when the sampling needle 2 enters the sample, it is subjected to a resistance force Fs，FsWith the sampling needle 2 getting deeper and progressively larger in the sample, the plunger rod has a tendency to move back, i.e. Fs+f+FdG + Fp, the static friction of which decreases gradually and then increases in the opposite direction to a maximum, the piston rod being subjected to a supporting force FdGradually decrease to zeroUp to Fs≤G+fmax+ Fp, the sampling needle 2 is constantly moving forward at a constant speed, and remains relatively stationary with the three-dimensional translation stage 4.
Referring to FIG. 7 (c), when Fs≥G+fmax+ Fp, the speed of the needle 2 in the sample is reduced to zero, the advance is stopped, then the speed is reversed, the retraction is started, see (d), F) in FIG. 7sGradually decreases as the plunger rod is retracted until it is zero after leaving the sample.
Repeating the sampling process, observing the result after sampling by a microscope, and measuring the air supply at the upper end of the cylinder and the resistance F of the sampling needle in the sampling processsThreshold value (i.e. F at the moment of retraction)s) The relationship between them is shown in FIG. 8, and the larger the upper end air pressure is, the larger F of the sampling needlesThe larger the threshold value of (A), FsMay fail when the threshold value of F is too smallsMay bend when the threshold value of (a) is too large. Therefore, when sampling succeeds, the air supply pressure indicated by the reading of the upper barometer 6 is taken as the minimum value of the empirical pressure; after sampling is successful, the air supply pressure at the upper end is still continuously increased until the human eye observes that the sampling needle 2 is bent or the microscope observes that the sampling needle 2 cuts open the sample, which indicates that the sampling is failed, and the air supply pressure indicated by the reading of the barometer 6 at the upper end is taken as the maximum value of the empirical air pressure.
Due to the presence of the sample stage 1, the resistance F to which the sampling needle 2 is subjectedsWhen too big, can take place to buckle, can fish tail sample section when the crooked back resumes vertical state, obtain the experience atmospheric pressure of certain limit through S104' S debugging, the air feed atmospheric pressure of selecting in the minimum and the maximum value within range of experience atmospheric pressure can make the sample succeed, can also avoid the crooked of sampling needle 2. Need not to install force transducer, in formal sampling process, control the Fs threshold value when retracting through selecting suitable Fp, when the resistance between sampling needle 2 and the sample reaches the threshold value, but cylinder 5 piston rod rapid retraction, phenomenon such as response delay fish tail sample can not appear.
S2, positioning: and S103, selecting the same type of sample used in the pilot sampling, firstly adjusting the sampling needle 2 in the center of the illumination field of the microscope, then keeping the sampling needle 2 still, moving the sample platform 1 through the observation or imaging of the microscope, and enabling the target area of the sample to be located right below the sampling needle 2 so as to perform positioning. As shown in FIG. 9, macrophages in the liver tissue section were located, and the sampling needle 2 was made of stainless steel, and had an inner diameter of 60 μm and an outer diameter of 260 μm, and the located cell mass was located in the inner diameter of the sampling needle 2.
S3, sampling: the electromagnetic valve 7 and the pressure reducing valve 8 are adjusted to enable the piston rod to be located at the lowest point in the air cylinder 5, and one empirical air pressure is selected from the empirical air pressure range obtained in S104 to supply air to the air cylinder 5 and keep constant, so that all similar samples can be sampled, and repeated sampling is achieved.
The electromagnetic valve 7 and the pressure reducing valve 8 are adjusted to enable the piston rod to be located at the lowest point in the air cylinder 5, and one empirical air pressure is selected from the empirical air pressure range obtained in S104 to supply air to the air cylinder 5 and keep constant, so that all similar samples can be sampled, and repeated sampling is achieved. Through the trial sampling debugging process of S104, the liver tissue slice is subjected to trial sampling, and the empirical air pressure range of successful sampling is as follows: the lower end is supplied with air at 7-13 kPa.
Then, a stainless steel sampling needle 2 with the inner diameter of 60 mu m and the outer diameter of 260 mu m is selected, and four methods are respectively used for sampling liver tissue slices to obtain macrophages in the liver. Referring to FIG. 10, A, the result of suction sampling using the prior art negative pressure technique of-31 kPa, the sampling result is not a standard circle due to suction caused by the negative pressure, but also carries with it surrounding tissue; fig. 10B shows the result of manually stamping with a large force, and due to the presence of the slide glass or the adhesive tape below the sliced sample, the sampling needle 2 is elastically deformed and bent when the stamping force is too large, and the sampling needle 2 cuts open the tissue during the resetting process after the sampling is finished, so that the sampling area is too large; fig. 10C, which is the result of manual poking with less force, the sampling needle 2 does not snap through the tissue, leaving only one loop of needle print on the tissue; in FIG. 10D, the tissue in the sampling needle 2 was completely removed as a result of sampling the lower end of the cylinder 5 with 10kPa supplied air in the range of 7kPa to 13kPa using the apparatus and the sampling method of the present invention.
S4, collecting: after sampling is finished, the sample stage 1 is removed, and a collecting device, which can be a perforated plate, an empty adhesive tape, an ep tube or the like, is placed below the sampling needle 2. The sampling needle 2 is supplied with air or liquid through a pipeline connected with a sample pushing port 17 on the side wall of the adapter 3, so that the tissues in the sampling needle 2 are pushed out to a collecting device below.
Aiming at the same type of samples, after a non-target area of one sample is subjected to sampling, a selection range of empirical air pressure is obtained, one empirical air pressure is selected from the selection range for sampling, the sampling success rate of the same type of samples is improved, and a sampling needle 2 is protected; after sampling, the sample platform 1 is moved away, and through the collection device and the cooperation of pushing away appearance mouth 17, the appearance is pushed away automatically, and for next sampling to make the quilt, the sample of the same kind only needs to carry out the preparation process of once trying to take a sample, can realize continuous automatic sampling, can also improve the sample success rate by a wide margin, and the interval time section between the twice sample, the sample is efficient, can satisfy the demand of sampling in batches.
The above description is only a few preferred embodiments of the present invention, and is not intended to limit the present invention, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Claims (10)
1. The adapter for connecting the piston rod of the cylinder and the sampling needle is characterized by comprising a piston rod connecting part and a needle head connecting part, wherein the piston rod connecting part comprises two symmetrically arranged clamping blocks which are integrally formed, a rod hole which is in splicing fit with the piston rod is formed between the two clamping blocks, one end of each of the two clamping blocks is flexibly connected, and the other end of each of the two clamping blocks is provided with an opening so as to adjust the size of the rod hole and facilitate the insertion and clamping of the piston rod;
the syringe needle connecting portion demountable installation be in the lower terminal surface of piston rod connecting portion, the lower fixed surface of syringe needle connecting portion has the direction arch, the lateral wall of syringe needle connecting portion seted up with the protruding intercommunication of direction pushes away a kind mouth.
2. The adapter for connecting a cylinder piston rod and a sampling needle according to claim 1, characterized in that a plastic fastener is fixed at the end of the sampling needle, and the sampling needle is in snap fit with the guide protrusion through the plastic fastener.
3. The adapter for connecting a cylinder piston rod and a sampling needle according to claim 1, characterized in that a rubber ball is sleeved on the periphery of the sampling needle, a connecting device is clamped in the guide protrusion, the radius of the lower end of the connecting device is gradually reduced, and the sampling needle is clamped in the connecting device through the rubber ball.
4. The adapter for connecting the piston rod of the cylinder and the sampling needle as claimed in claim 2 or 3, wherein one of the clamping blocks is provided with an upper through hole through the side wall of the opening end thereof, the other clamping block is provided with an upper threaded hole coaxial with the upper through hole, the diameter of the upper through hole is larger than that of the upper threaded hole, an upper bolt penetrates through the upper through hole and is in threaded connection with the upper threaded hole after penetrating through the upper through hole, and the upper bolt abuts against the side wall of the clamping block provided with the upper through hole.
5. The adapter for connecting the piston rod of the cylinder and the sampling needle as claimed in claim 2 or 3, wherein a lower through hole is formed at the edge of the needle head connecting portion in a penetrating manner, a lower threaded hole is correspondingly formed at the lower end face of the piston rod connecting portion, the diameter of the lower through hole is larger than that of the lower threaded hole, a lower bolt is arranged in the lower through hole in a penetrating manner, the lower bolt penetrates through the lower through hole and then is in threaded connection with the lower threaded hole, and the lower bolt abuts against the lower surface of the needle head connecting portion.
6. The adapter for connecting the piston rod of the cylinder and the sampling needle as claimed in claim 2 or 3, wherein the sample pushing port is provided with threads on the periphery thereof for connecting a pipeline in a threaded manner so as to supply air or liquid to the sampling needle and push out the sample in the sampling needle.
7. The adapter for connecting a cylinder piston rod and a sampling needle according to claim 2 or 3, wherein the surfaces of the piston rod connecting part and the needle head connecting part are both fixed with oxidation blackening layers.
8. The adapter for connecting the piston rod of the cylinder and the sampling needle as claimed in claim 2 or 3, wherein the end of the piston rod in the insertion fit with the rod hole is sleeved with a raw material belt.
9. A device suitable for continuously obtaining micro tissue blocks on a tissue slice is characterized by comprising a portal frame, a three-dimensional translation table fixed on one side of the portal frame, an air cylinder fixed with the three-dimensional translation table, a sample table movably arranged below the air cylinder, a sampling needle and a connector used for connecting an air cylinder piston rod and the sampling needle;
and an imaging device is fixed on the portal frame below the sample table and used for positioning a target area of a sample on the sample table.
10. A method for continuously obtaining micro-tissue blocks on a tissue slice, comprising the steps of:
s1, mounting the piston rod connecting part at the lower end of the piston rod, and screwing the upper bolt; mounting the sampling needle on the guide projection; mounting the needle head connecting part on the lower surface of the piston rod connecting part, and screwing down the lower bolt;
s2, positioning the sample through the imaging device to move the sample stage according to the position of the target area in the imaging device, so that the target area is positioned right below the sampling needle;
s3, the three-dimensional translation table drives the sampling needle to advance for sampling;
and S4, after sampling, moving the sample platform away, placing a collecting device below the sampling needle, connecting a pipeline for a sample pushing port, and supplying gas or liquid into the sampling needle so as to push the tissues in the sampling needle out of the collecting device.
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| CN201911257890.3A CN110938530B (en) | 2019-12-10 | 2019-12-10 | Method for continuously obtaining micro tissue blocks |
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| CN201911257890.3A CN110938530B (en) | 2019-12-10 | 2019-12-10 | Method for continuously obtaining micro tissue blocks |
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| CN110938530B (en) | 2021-12-21 |
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