Search

US-12623216-B2 - Method of fabricating block-copolymer-based 3D polymorphic artificial bilayer membrane structure

US12623216B2US 12623216 B2US12623216 B2US 12623216B2US-12623216-B2

Abstract

The present invention relates to a method for manufacturing a three-dimensional structure based on a block copolymer. The method comprises the steps of injecting a block copolymer (BCP) solution into each micro-well formed on the substrate and drying it to form a block copolymer layer, and applying a buffer to the block copolymer layer to hydrate the micro-well in three dimensions Forming the structure, after the three-dimensional structure is formed, injecting and curing a hydrogel solution around the three-dimensional structure may include the step of enhancing stability. In particular, the process of hydration by applying a buffer to the micro-well is performed while an electric field is applied. By controlling the concentration of the block copolymer (BCP) and the amplitude and frequency of the electric field, a three-dimensional artificial cell membrane having a desired size and shape, such as a spherical or ciliary shape and high stability (100% survival for 50 days) is manufactured can do. The present invention can be efficiently applied to various biological fields such as artificial cells, cell-mimicking biosensors, and bioreactors.

Inventors

  • Tae Song Kim
  • Dong Hyun Kang

Assignees

  • KOREA INSTITUTE OF SCIENCE AND TECHNOLOGY

Dates

Publication Date
20260512
Application Date
20221122
Priority Date
20220225

Claims (10)

  1. 1 . A block copolymer-based three-dimensional polymorphic artificial structure manufacturing method comprising the steps of: forming a micro-well array on a substrate; forming a block copolymer layer by injecting a block copolymer (BCP) solution into each micro-well of the micro well array and drying the micro-well array; forming a three-dimensional structure from the block copolymer layer to the top of each micro-well by applying a buffer to each micro-well in which the block copolymer layer is formed, wherein at least a part of a hydration process is carried out under an electric field applied between the top and bottom of the micro-well in which the block copolymer layer is formed.
  2. 2 . The method of claim 1 , wherein the block copolymer solution comprises polybutadiene-b-polyethylene oxide (PBd-PEO).
  3. 3 . The method of claim 1 , wherein the buffer comprises a sucrose solution at a certain concentration.
  4. 4 . The method of claim 1 , wherein a size and a shape of the three-dimensional polymorphic artificial structure are determined by at least one of a concentration of the block copolymer solution, an amplitude of the electric field, and a frequency of the electric field.
  5. 5 . The method of claim 1 , further comprising enhancing a stability of the formed three-dimensional polymorphic artificial structure.
  6. 6 . The method of claim 5 , wherein the step of enhancing a stability of the three-dimensional structure comprises a process of injecting and curing a hydrogel solution around the three-dimensional polymorphic artificial structure after the three-dimensional structure is formed.
  7. 7 . The method of claim 6 , wherein the process of injecting and curing the hydrogel solution around the three-dimensional polymorphic artificial structure is carried out in a state in which the electric field is applied.
  8. 8 . The method of claim 1 , wherein a diameter (d) of the three-dimensional polymorphic artificial structure is determined by the following formula, where the concentration of the block copolymer solution is x: d= 1.14+8.7 x− 1.3 x 2 +0.1 x 3 .
  9. 9 . The method of claim 1 , wherein a frequency of the electric field is controlled to a value greater than or equal to a first frequency and less than or equal to a second frequency when a shape of the three-dimensional polymorphic artificial structure is formed in a ciliary shape, and is controlled to a value less than the first frequency or greater than the second frequency when the shape of the three-dimensional polymorphic artificial structure is formed in a spherical shape, wherein the ciliary type refers to a shape with an aspect ratio of 1.5 or more, and the first frequency is less than the second frequency.
  10. 10 . The method of claim 9 , wherein the first frequency is 1 kHz, and the second frequency is 1 MHz.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit under 35 USC 119(a) of Korean Patent Application No. 10-2022-0024927 filed on Feb. 25, 2022, in the Korean Intellectual Property Office, the entire disclosure of which is incorporated herein by reference for all purposes. TECHNICAL FIELD The present invention relates to a method of fabricating a 3D structure based on a block-copolymer, and more particularly, it relates to a method capable of manufacturing a 3D artificial structure having various sizes and shapes and high stability by controlling the concentration of the block copolymer and the amplitude and frequency of the electric field. BACKGROUND OF THE INVENTION Cells perform functions to maintain life phenomena, such as sensing and regulating extracellular changes, and communication inside and outside the cell. The core of this function consists of a double membrane with a thickness of about 5 to 10 nm and a membrane protein including ion channel, pore forming protein/peptide or other synthetic pores. Recent research has been conducted to fabricate a double membrane mimicking the function and structure of a cell, and apply it to a biosensor by combining channel membrane proteins or biological nanopores. A company called OXFORD NANOPORE TECHNOLOGIES (nanoporetech.com) in the UK has combined protein nanopores with a two-dimensional planar polymer membrane, and then they are commercializing a fluid chip that can record the single-molecule DNA sequence moving through the pore at high speed in real-time by applying an electric field between the top and bottom of the membrane. (D. Deamer et al., nature biotechnology, Vol. 34, No. 5, p. 518˜524, 2016, X. Kang et al., Vol. 29, 9145˜9153, 2019). However, an attempt to apply an artificial cell membrane based on the biological double membrane or an amphiphilic block copolymer double membrane is a two-dimensional planar structure on a specific substrate. However, the shape of the cell is usually a spherical or an irregular shape rather than a planar shape, including a curved surface. Accordingly, attempts are being made for three-dimensional manufacturing rather than flat. As an example, Korean Patent Registration No. 10-1913342 discloses a method of manufacturing a three-dimensional artificial biological membrane structure having a sufficient reaction area and high stability on a substrate using a lipid material, which is an actual cell membrane constituent material. However, although this prior art can relatively simply manufacture a three-dimensional artificial biological membrane structure using a lipid material, it is difficult to manufacture a three-dimensional structure to have various desired sizes and shapes. For example, when using a lipid material like the technique disclosed in Patent Registration No. 10-1913342, it is difficult to implement a narrow and long cilia-shaped structure. In addition, the prior art also has a problem in that it is difficult to stably maintain the manufactured three-dimensional structure for a long time. Therefore, it is possible to manufacture 3D structures of various sizes and shapes suitable for imitation targets, such as spherical shapes as well as narrow and long cilia similar to real cells, and a technology that allows the manufactured 3D structures to be maintained robustly and stably for a more extended time development is required. PRIOR ART Patent Korean Patent Registration 10-1913342 (3D lipid structure array manufacturing method and 3D lipid structure according thereto, Day of Grant: 2018 Oct. 30) CONTENT OF THE INVENTION Challenge to be Solved Accordingly, the present invention has been devised to meet the above needs, it is an object to provide a method for manufacturing a 3D polymorphic artificial structure based on a block copolymer, which can manufacture a 3D structure with high stability of a desired size and shape by controlling the concentration of the block copolymer and the amplitude and frequency of the electric field to be. Means of Solving the Problem In order to achieve the above object, the block copolymer-based three-dimensional polymorphic artificial structure manufacturing method according to the present invention comprises a step of injecting a block copolymer (BCP) solution into each micro-well of a micro-well array formed on a substrate, and drying to form a block copolymer layer; and a step of forming a three-dimensional structure on top of the micro-well from the block copolymer layer by applying a buffer (hydration buffer) to each micro-well in which the block copolymer layer is formed to hydrate. In this case, at least a part of the process of hydration by applying a buffer to each micro-well in which the block copolymer layer is formed may be performed while an electric field is applied between the upper and lower portions of the micro-well in which the block copolymer layer is formed. The block copolymer may include PBd-PEO (polybutadiene-b-polyethylene oxide).