CN114265131B

CN114265131B - Biological fluid microsphere, preparation method and optical imaging method of microsphere lens

Info

Publication number: CN114265131B
Application number: CN202111543898.3A
Authority: CN
Inventors: 杨慧; 陈希; 顾国强
Original assignee: Shenzhen Institute of Advanced Technology of CAS
Current assignee: Shenzhen Institute of Advanced Technology of CAS
Priority date: 2021-12-16
Filing date: 2021-12-16
Publication date: 2023-07-14
Anticipated expiration: 2041-12-16
Also published as: CN114265131A; WO2023109703A1

Abstract

According to the biological fluid microsphere and the preparation method thereof, the biological fluid is coated on the convex column of the PDMS, egg white on the convex column is transferred onto the liquid layer of the PDMS, the egg white is self-assembled in the PDMS to form the microsphere, the PDMS is removed to obtain the biological fluid microsphere, the biological fluid microsphere and the preparation method thereof can be used for preparing the manufacturing technology of the high-circularity microsphere with the diameter of several micrometers to hundreds of micrometers, the whole process is carried out in a stable environment, the manufacturing method is simple, and the obtained biological fluid microsphere has good biocompatibility. In addition, the application also provides an optical imaging method based on the biological fluid microsphere.

Description

Biological fluid microsphere, preparation method and optical imaging method of microsphere lens

Technical Field

The invention relates to the technical field of medical instruments, in particular to a biological fluid microsphere, a biological fluid microsphere preparation method and an optical imaging method of a microsphere lens.

Background

Optical microscopy imaging plays a very important role in observing the real world and in finding and exploring the unknown world. Limited by the diffraction limit of light, the resolution of conventional optical microscopes is not less than 200nm at maximum when the wavelength of the light source used is in the visible range. The diffraction limit exists because the evanescent wave propagates only in the vicinity of the near field, the intensity of the evanescent wave decays exponentially with increasing distance, and the evanescent wave band has a high spatial frequency signal representing fine structural information of the object.

Super-resolution imaging techniques are capable of overcoming or bypassing diffraction limited obstacles to achieve imaging of sub-diffraction limited-scale objects. Techniques for imaging include Near field super-resolution techniques (e.g., near field optical microscopy (nsar-field optical microscopy, NSOM)) and far field super-resolution imaging techniques (e.g., stimulated radiation loss (stimulated emission depletion microscopy, STED), photosensitive positioning microscopy (photoactivated localization microscopy, PALM), etc.). The observation and detection of viruses, proteins, intracellular fine structures and the like can be realized by utilizing the technologies, and the technology is an important point and a hot point of the research of the modern optical imaging technology.

However, these techniques either require expensive and complex equipment or require fluorescent labeling of the sample, which is inevitably phototoxic, and thus are greatly limited in terms of label-free and live cell imaging. In recent ten years, a novel microsphere lens-assisted super-resolution imaging technology is discovered by researchers of a plurality of subject groups at home and abroad, microspheres are combined with a conventional optical microscope, and resolution and imaging of objects with sub-diffraction limit dimensions are obtained by utilizing the super-focusing light field effect of the microspheres. The super-resolution imaging technology has the characteristics and advantages of no marking, no phototoxicity, low manufacturing cost, high imaging speed, simple device structure and the like.

The microspheres currently used for super-resolution imaging are mostly inorganic and organic compounds, such as SiO ₂ Microsphere and BaTiO ₃ Glass microspheres, polystyrene microspheres, and the like, have poor compatibility with biological samples, limiting the application of the technology in biological and medical imaging. The egg white has good biocompatibility and bioactivity, is easy to be absorbed and degraded by human bodies, has wide sources and low price, is simple and feasible in preparation process, and has not been reported similarly at present when the microsphere taking the egg white as the raw material is applied to super-resolution optical imaging.

Disclosure of Invention

In view of the above, it is necessary to provide a biological fluid microsphere for performing super-resolution imaging observation in a biological fluid microsphere array structure.

In order to solve the problems, the invention adopts the following technical scheme:

one of the purposes of the application is to provide a preparation method of biological fluid microspheres, which comprises the following steps:

coating biological liquid on the convex columns of the PDMS, wherein the biological liquid comprises egg white;

transferring the egg white on the convex column onto the liquid layer of the PDMS;

the egg white is self-assembled in the PDMS to form microspheres;

removing the PDMS to obtain the biological fluid microsphere.

In some of these embodiments, in the step of applying the egg white solution to the PDMS posts, the steps specifically include:

and transferring the egg white solution on the PDMS convex column onto the PDMS liquid layer in a light pressure mode.

In some of these embodiments, in the step of removing the PDMS to obtain the bio-fluid microspheres, specifically: PDMS was removed using ethyl acetate solvent to obtain the bio-liquid microsphere.

The second purpose of the application is to provide a biological fluid microsphere which is prepared by the preparation method of the biological fluid microsphere, wherein the diameter of the biological fluid microsphere is between 5 and 500 um.

A third object of the present application is to provide an optical imaging method of a microsphere lens, comprising the steps of:

directly peeling off the protective film on the surface of the blue-ray disc;

coating a PDMS film on the surface of the blue-ray disc;

transferring the egg white on the convex column to the PDMS film;

and imaging the grating structure of the blue-ray disc by using the biological liquid microsphere.

In some embodiments, the step of peeling the protective film from the surface of the blu-ray disc specifically includes the steps of:

the blue-ray disc was placed on a heating plate at 45-55 deg.c for 1.5-2.5 min, and then the protective film was peeled off with tweezers.

In some embodiments, in an optical imaging method of a microsphere lens, specifically: the thickness of the PDMS film is 200-1000 mu m; the egg white liquid drops can self-assemble through interfacial interaction between surface tension and PDMS to form an array distributed microsphere structure.

In some of these embodiments, the grating structure is a periodic stripe structure consisting of lines with a line width of 180nm and an arrangement of grooves with a pitch of 140 nm.

By adopting the technical scheme, the invention has the following technical effects:

according to the biological fluid microsphere and the preparation method thereof, biological fluid is coated on the convex column of PDMS, egg white on the convex column is transferred onto the liquid layer of PDMS, the egg white is self-assembled in the PDMS to form the microsphere, and the PDMS is removed to obtain the biological fluid microsphere.

In a second aspect, the present application further provides an optical imaging method of a microsphere lens, which includes peeling a protective film on a surface of the optical disc, coating a PDMS film on the surface of the blu-ray disc, transferring egg white on the convex column to the PDMS film, and imaging a grating structure of the blu-ray disc using the bio-liquid microsphere. The thickness of the PDMS film is 200-1000 mu m; the egg white liquid drops self-assemble to form an array-distributed microsphere structure through interfacial interaction between surface tension and PDMS; the grating structure is a periodic stripe structure formed by arranging lines with the line width of 180nm and grooves with the interval of 140nm, stripes of the blue-ray disc amplified by a single-layer egg white microstructure are observed through a microscope, and the imaging method can realize resolution of a sub-diffraction limit scale (less than 200 nm) sample under an optical microscope and display resolution capability exceeding diffraction limit.

Drawings

In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the following description will briefly explain the embodiments of the present invention or the drawings used in the description of the prior art, and it is obvious that the drawings described below are only some embodiments of the present invention, and other drawings can be obtained according to these drawings without inventive effort for a person skilled in the art.

FIG. 1 is a flow chart of the steps for preparing biological fluid microspheres according to example 1 of the present invention;

FIG. 2 is a schematic view of a microscope of biological fluid microspheres according to example 1 of the present invention;

FIG. 3 is a flow chart of the steps of the method for optical imaging of a microlens provided in embodiment 2 of the present application;

fig. 4 is a super-resolution imaging schematic diagram of the microsphere lens provided in embodiment 2 of the present application.

Detailed Description

Embodiments of the present invention are described in detail below, examples of which are illustrated in the accompanying drawings, wherein like or similar reference numerals refer to like or similar elements or elements having like or similar functions throughout. The embodiments described below by referring to the drawings are illustrative and intended to explain the present invention and should not be construed as limiting the invention.

In the description of the present invention, it should be understood that the directions or positional relationships indicated by the terms "upper", "lower", "horizontal", "inner", "outer", etc., are based on the directions or positional relationships shown in the drawings, are merely for convenience in describing the present invention and simplifying the description, and do not indicate or imply that the devices or elements referred to must have a specific orientation, be constructed and operated in a specific orientation, and thus should not be construed as limiting the present invention.

Furthermore, the terms "first," "second," and the like, are used for descriptive purposes only and are not to be construed as indicating or implying a relative importance or implicitly indicating the number of technical features indicated. Thus, a feature defining "a first" or "a second" may explicitly or implicitly include one or more such feature. In the description of the present invention, the meaning of "a plurality" is two or more, unless explicitly defined otherwise.

The present invention will be described in further detail with reference to the drawings and examples, in order to make the objects, technical solutions and advantages of the present invention more apparent.

Example 1

Referring to fig. 1, a flowchart of steps for preparing a biological fluid microsphere according to an embodiment of the present invention includes the following steps:

step S110: a biological fluid, including an egg white solution, was coated onto the PDMS posts.

In this embodiment, the PDMS pillars may be obtained by photolithography, machining (COC, COP, and PMMA), laser engraving, and imprinting.

Specifically, an egg white solution was applied to the PDMS cylinder head for a contact time of typically 5s, empirically.

Step S120: and transferring the egg white on the convex column onto the liquid layer of the PDMS.

In this example, the egg white on the posts was transferred to the PDMS liquid layer by light pressing.

Step S130: the egg white self-assembles in the PDMS to form microspheres.

Transferring the egg white drops on the convex columns to the PDMS film, wherein the egg white drops can self-assemble to form an array-distributed microsphere structure through surface tension and interfacial interaction between the egg white drops and PDMS;

it is understood that the egg white droplets will self-assemble into an array-distributed microsphere structure through surface tension and interfacial interactions with PDMS.

Step S140: removing the PDMS to obtain the biological fluid microsphere.

In this example, PDMS was removed using ethyl acetate solvent to obtain the bio-fluid microspheres.

Referring to fig. 2, a microscopic image of the biological fluid microsphere provided herein is shown.

The biological fluid microsphere and the preparation method thereof can be used for preparing the high-circularity microsphere with the diameter of several micrometers to hundreds of micrometers, the whole process is carried out in a stable environment, the preparation method is simple and convenient, and the prepared microsphere has good biocompatibility.

Example 2

Referring to fig. 3, a flowchart of steps of an optical imaging method of a microsphere lens is provided, which includes the following steps:

step S210: the protective film on the surface of the blue ray disc is directly peeled off.

In this example, the blu-ray disc was placed on a heating plate at 45 to 55 deg.c for a heating treatment of 1.5 to 2.5min, and then the protective film on the surface was peeled off with tweezers.

Step S220: and coating a PDMS film on the surface of the blue-ray disc, wherein the thickness of the PDMS film is 200-1000um.

In this embodiment, a PDMS film is spin-coated on the blu-ray disc.

Step S230: transferring the egg white drops on the convex columns to the PDMS film, wherein the egg white drops can self-assemble to form an array-distributed microsphere structure through surface tension and interfacial interaction between the egg white drops and PDMS;

step S240: and imaging the microstructure of the blue-ray disc by using the biological fluid microsphere.

Referring to fig. 4, the grating structure on the blue-ray disc can be observed by using the egg white array microsphere, the grating structure is a periodic stripe structure formed by lines with a line width of 180nm and groove arrangement with a pitch of 140nm, and the stripes of the blue-ray disc amplified by the single-layer egg white microstructure are observed through a microscope.

The foregoing description of the preferred embodiments of the present invention has been provided for the purpose of illustrating the general principles of the present invention and is not to be construed as limiting the scope of the invention in any way. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention, and other embodiments of the present invention as will occur to those skilled in the art without the exercise of inventive faculty, are intended to be included within the scope of the present invention.

Claims

1. An optical imaging method of a microsphere lens, comprising the steps of:

peeling off the protective film on the surface of the blue-ray disc;

coating a PDMS film on the surface of the blue-ray disc;

coating biological liquid on the PDMS convex column, wherein the biological liquid comprises egg white solution;

transferring the egg white solution on the convex column to the PDMS film, wherein the egg white solution can self-assemble to form an array-distributed microsphere structure through surface tension and interfacial interaction between the egg white solution and PDMS;

and imaging the grating structure of the blue-ray disc by using the microspheres.

2. The optical imaging method as claimed in claim 1, wherein in the step of peeling the protective film of the surface of the blu-ray disc, the steps of:

3. The optical imaging method of claim 1, wherein the PDMS film has a thickness of 200-1000 μm.

4. The optical imaging method of claim 1, wherein the microsphere has a diameter of between 5 and 500 μm.

5. The optical imaging method of claim 1, wherein the grating structure is a periodic stripe structure consisting of lines with a line width of 180nm and an arrangement of grooves with a pitch of 140 nm.