Organoid oct embedding device
By designing an embedding device with a limiting ring and an OCT shunt, the problem of adhesion to the wall during organoid embedding was solved, achieving stable embedding of organoids and simplifying the operation.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2023-05-30
- Publication Date
- 2026-03-31
AI Technical Summary
Organoids are prone to adhesion to the wall during the embedding process, which makes the operation difficult.
An embedding device comprising an embedding cassette, a limiting ring, and an OCT shunt was designed. The limiting ring restricts the organoid to the center of the embedding cassette, and the OCT reagent flows in from multiple directions to prevent the organoid from shifting.
It effectively prevents organoids from adhering to the wall during the embedding process, simplifies the operation procedure, and improves embedding efficiency.
Smart Images
Figure CN116640661B_ABST
Abstract
Description
Technical Field
[0001] This application relates to an organoid experimental apparatus, and more particularly to an organoid OCT embedding device. Background Technology
[0002] Organoids are 3D microstructures formed by stem cells in vitro, with diameters as small as tens of micrometers. After culturing organoids in 96-well plates, they are aspirated into EP tubes using a pipette tip, and excess culture medium is removed. After washing with PBS, they are fixed with 4% paraformaldehyde; after fixation for 15-30 minutes, the paraformaldehyde is removed, and the tubes are washed three times with PBS. They are then placed in a 20-30% sucrose solution overnight for dehydration. After dehydration, the organoids with sucrose solution are transferred to an embedding cassette using a pipette tip. Next, the sucrose solution surrounding the organoids is removed, and OCT reagent is added to the embedding cassette to embed the organoids. Ideally, the organoids should be centered within the embedding cassette after embedding. However, because organoids are very small, unlike other tissues, they are easily affected by the sucrose solution aspirated by the pipette tip during sucrose removal, and their position often adheres to the edge of the embedding cassette. In addition, during the addition of OCT reagents, organoids are also prone to adhere to the edge of the embedding cassette as the OCT reagents flow. This is the so-called adhesion phenomenon, which causes difficulties for subsequent operations and processing. Summary of the Invention
[0003] In view of the above problems, this application aims to provide an organoid OCT embedding device that can effectively avoid the phenomenon of organoids sticking to the wall in the embedding box.
[0004] The organoid OCT embedding device of this application includes: an embedding box, a limiting ring, and an OCT shunt;
[0005] The embedding box is formed into a disc-shaped structure, with an upwardly extending ridge around its outer perimeter; the ridge surrounds and forms an OCT containing space;
[0006] The limiting ring is formed into a circular structure, and multiple holes are formed around the limiting ring. The diameter of the holes is smaller than the minimum diameter of the organoid.
[0007] The OCT shunt includes a funnel; the lower end of the funnel is closed; multiple pairs of shunt tubes are formed around the lower part of the funnel, each pair of shunt tubes including two shunt tubes facing each other.
[0008] The containment ring is placed at the center of the embedding cassette; the containment ring is used to accommodate the organoids transferred from the sucrose solution;
[0009] The OCT shunt is placed on the embedding cassette, and its multiple pairs of shunt tubes are supported on the embedding cassette, located in the OCT housing space, and outside the limiting ring.
[0010] Preferably, the embedding box has a V-shaped groove formed around its center.
[0011] Preferably, the groove includes a downwardly inclined outer ring slope and an upwardly inclined inner ring slope.
[0012] Preferably, the limiting ring is located on the inclined surface of the inner ring.
[0013] Preferably, the embedding box is formed horizontally at its center; the horizontal position of the center is lower than the upper edge of the outer ring slope.
[0014] Preferably, a support foot is formed on the lower side of the limiting ring; the support foot is located in the groove; supported by the support foot, a gap is formed between the limiting ring and the inner ring inclined surface.
[0015] Preferably, the lower surface of the support foot corresponds to the groove.
[0016] Preferably, the diverter tubes are formed in pairs.
[0017] The OCT embedding device of this application, through the setting of a limiting ring, confines the organoid to the center of the embedding box. When aspirating the sucrose solution, it is aspirated along the outer wall of the limiting ring. This ensures that the organoid remains within the limiting ring and prevents it from being accidentally aspirated into the pipette tip. By setting an OCT splitter, the OCT reagent is not poured in one direction, but simultaneously from multiple directions. In this way, the OCT reagent flows from the periphery to the center, preventing the organoid from sticking to the wall. Moreover, the opposing split tubes inject OCT reagent from opposite directions, which cancels out the positional effect on the organoid and is more conducive to keeping the organoid in the center. Attached Figure Description
[0018] Figure 1 This is a three-dimensional structural schematic diagram of the organoid OCT embedding device of this application;
[0019] Figure 2 This is a schematic diagram of the front view structure of the organoid OCT embedding device of this application;
[0020] Figure 3 This is a top view of the organoid OCT embedding device of this application;
[0021] Figure 4 This is a schematic cross-sectional view of the organoid OCT embedding device of this application along line AA;
[0022] Figure 5 This is a three-dimensional structural diagram of the embedding box of the organoid OCT embedding device of this application;
[0023] Figure 6for Figure 5 A schematic diagram of the cross-section of the embedding cassette;
[0024] Figure 7 This is a three-dimensional structural diagram of the limiting ring of the organoid OCT embedding device of this application;
[0025] Figure 8 for Figure 7 A schematic diagram of the main view structure of the limiting ring;
[0026] Figure 9 for Figure 7 A top view of the limiting ring structure;
[0027] Figure 10 This is a three-dimensional structural schematic diagram of the OCT shunt of the organoid OCT embedding device of this application;
[0028] Figure 11 for Figure 10 A top view of the OCT shunt. Detailed Implementation
[0029] The organoid OCT embedding device of this application will now be described in detail with reference to the accompanying drawings.
[0030] The organoid OCT embedding device of this application includes: an embedding box 10, a limiting ring 20, and an OCT shunt 30.
[0031] The embedding box 10 is formed into a disc-shaped structure, with an upwardly extending convex edge 11 around its outer periphery; the convex edge 11 surrounds and forms an OCT containing space.
[0032] The limiting ring 20 is formed as a circular ring structure 21, and multiple holes 22 are formed around the limiting ring. The diameter of the holes 22 is smaller than the minimum diameter of the organoid, so that sucrose solution and OCT reagent can pass through, but organoids cannot.
[0033] The OCT shunt 30 includes a funnel 31; the lower end 32 of the funnel 31 is closed; a plurality of pairs of shunt tubes 33 are formed around the lower part of the funnel 31, each pair of shunt tubes 33 including two shunt tubes 33 facing each other.
[0034] The limiting ring 20 is placed at the center of the embedding cassette 10; the limiting ring 20 is used to contain the organoids transferred from the sucrose solution. The main function of the limiting ring 20 is to restrict the organoids within it, preventing them from adhering to the edge of the embedding cassette.
[0035] The OCT shunt 30 is placed on the embedding cassette 10, the funnel 31 is located above the embedding cassette 10, and multiple pairs of shunt tubes 33 are supported inside the embedding cassette 10, located in the OCT containment space, and located outside the limiting ring 20.
[0036] The bottom surface of the embedding cassette 10 can be flat, like a conventional embedding cassette.
[0037] The bottom surface of the embedding box 10 can also form a groove 14.
[0038] like Figure 6 As shown, a V-shaped groove 14 is formed around the center of the embedding box 10.
[0039] The groove 14 includes a downwardly inclined outer ring slope 12 and an upwardly inclined inner ring slope 13.
[0040] The limiting ring 20 is located on the inner ring inclined surface 13.
[0041] The main function of the inner ring bevel 13 is to facilitate the removal of sucrose solution. After the organoid and sucrose solution are transferred together into the limiting ring by the nozzle, the sucrose solution easily flows along the inner ring bevel 13 into the groove 14; thus, by inserting the nozzle into the groove 14, the sucrose solution can be removed, the suction force of the nozzle on the organoid will be reduced, and the sucrose can be removed more easily or more quickly.
[0042] The outer ring slope 12 is designed primarily to allow OCT reagent to flow towards the central position 15. The lower end of the shunt tube 33 is supported on the outer ring slope 12. The OCT reagent flowing out of the shunt tube 33 flows into the groove 14 along the outer ring slope, forming a buffer for the flow of OCT reagent. Then, the OCT reagent level in the groove rises and flows evenly into the limiting ring from all directions through the gap between the coil and the inner ring slope and the hole on the limiting ring, minimizing the impact on the position of the organoids within.
[0043] The embedding box 10 is formed horizontally at its center position 15; the horizontal position of the center position 15 is lower than the upper edge of the outer ring slope 12.
[0044] In another embodiment, a support foot 23 is formed on the lower side of the limiting ring 20; the support foot 23 is located in the groove 14; supported by the support foot 23, a gap is formed between the limiting ring 20 and the inner ring inclined surface 13. The lower surface of the support foot 23 corresponds to the groove 14. The support foot 23 helps to maintain the position of the coil in the groove.
[0045] The positioning ring and support feet are preferably made of silicone, which facilitates subsequent sectioning operations. The positioning ring can also be colored, making it easier to determine the location of the organoid in the section using this color.
[0046] The shunt tubes 33 are pre-selected to be arranged in two pairs, or four in total, evenly distributed in a circumferential direction. The purpose of designing the shunt tubes 33 in pairs is to cancel out the effect of the OCT reagent flowing out of the shunt tubes in opposite directions on the position of the organoid.
[0047] In use, organoids in the sucrose solution are transferred to the containment ring using a pipette tip. The sucrose solution is then removed from the containment ring, especially from the grooves, using the pipette tip. OCT reagent is poured into a funnel and flows through multiple pairs of shunts into the embedding cassette, first filling the grooves, then spreading from the grooves into the containment ring to embed the organoids. After embedding, cryotherapy can be performed.
[0048] Unless otherwise defined, all technical and / or scientific terms used in this application have the same meaning as commonly understood by one of ordinary skill in the art to which this invention relates. The materials, methods, and embodiments mentioned in this application are illustrative only and not restrictive.
[0049] Although the present invention has been described in conjunction with specific embodiments, those skilled in the art can make appropriate substitutions, modifications and changes within the inventive spirit of this application, and such substitutions, modifications and changes still fall within the protection scope of this application.
Claims
1. An organoid OCT embedding device comprising: The embedding box, the limiting ring and the OCT shunt; The embedding box is formed as a disc structure, and a convex edge extending upward is formed around the outer periphery thereof; The convex edge forms an OCT containing space; The limiting ring is formed as a circular ring structure, and a plurality of holes are formed around the limiting ring, the diameter of the holes being smaller than the minimum diameter of the organoids; The OCT shunt comprises a funnel, the lower end of the funnel being closed, and a plurality of pairs of shunt tubes are formed around the funnel at the lower portion thereof, each pair of shunt tubes comprising two shunt tubes facing each other; The limiting ring is used to be placed at the center position of the embedding box, and the organoids transferred from the sucrose solution are accommodated in the limiting ring; The OCT shunt is placed on the embedding box, and the plurality of pairs of shunt tubes thereof are supported on the embedding box, located in the OCT containing space and outside the limiting ring.
2. The organoid OCT embedding device according to claim 1, wherein: A groove with a V-shaped cross section is formed around the center of the embedding box.
3. The organoid OCT embedding device according to claim 2, wherein: The groove comprises an outer ring inclined surface inclined downward and an inner ring inclined surface inclined upward.
4. The organoid OCT embedding device according to claim 3, wherein: The limiting ring is located on the inner ring inclined surface.
5. The organoid OCT embedding device according to claim 3, wherein: The embedding box is formed as a horizontal structure at the center position thereof, and the horizontal position of the center position is lower than the upper edge of the outer ring inclined surface.
6. The organoid OCT embedding device according to claim 4, wherein: A supporting leg is formed on the lower side of the limiting ring, the supporting leg is located in the groove, and a gap is formed between the limiting ring and the inner ring inclined surface under the support of the supporting leg.
7. The organoid OCT embedding device according to claim 6, wherein: The lower surface of the supporting leg corresponds to the groove.
8. The organoid OCT embedding device according to claim 1, wherein: The shunt tubes are formed as 2 pairs.
Citation Information
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