CN113447646B - By using C-SiO 2 Method for accelerating ELISA detection process by adsorption material - Google Patents
By using C-SiO 2 Method for accelerating ELISA detection process by adsorption material Download PDFInfo
- Publication number
- CN113447646B CN113447646B CN202110691502.3A CN202110691502A CN113447646B CN 113447646 B CN113447646 B CN 113447646B CN 202110691502 A CN202110691502 A CN 202110691502A CN 113447646 B CN113447646 B CN 113447646B
- Authority
- CN
- China
- Prior art keywords
- sio
- antigen
- antibody
- liquid
- adsorption material
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- 239000000463 material Substances 0.000 title claims abstract description 53
- 238000002965 ELISA Methods 0.000 title claims abstract description 36
- 238000001179 sorption measurement Methods 0.000 title claims abstract description 32
- 229910004298 SiO 2 Inorganic materials 0.000 title claims abstract description 27
- 238000000034 method Methods 0.000 title claims abstract description 25
- 238000001514 detection method Methods 0.000 title claims abstract description 14
- 239000000427 antigen Substances 0.000 claims abstract description 56
- 102000036639 antigens Human genes 0.000 claims abstract description 56
- 108091007433 antigens Proteins 0.000 claims abstract description 56
- 239000007788 liquid Substances 0.000 claims abstract description 45
- 239000011248 coating agent Substances 0.000 claims abstract description 6
- 238000000576 coating method Methods 0.000 claims abstract description 6
- 239000000243 solution Substances 0.000 claims description 33
- 239000000203 mixture Substances 0.000 claims description 24
- 238000005406 washing Methods 0.000 claims description 22
- 239000003463 adsorbent Substances 0.000 claims description 18
- 239000003575 carbonaceous material Substances 0.000 claims description 15
- 238000010790 dilution Methods 0.000 claims description 14
- 239000012895 dilution Substances 0.000 claims description 14
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 14
- VXEGSRKPIUDPQT-UHFFFAOYSA-N 4-[4-(4-methoxyphenyl)piperazin-1-yl]aniline Chemical compound C1=CC(OC)=CC=C1N1CCN(C=2C=CC(N)=CC=2)CC1 VXEGSRKPIUDPQT-UHFFFAOYSA-N 0.000 claims description 10
- 238000001035 drying Methods 0.000 claims description 10
- 239000005049 silicon tetrachloride Substances 0.000 claims description 10
- 238000002156 mixing Methods 0.000 claims description 9
- 239000007853 buffer solution Substances 0.000 claims description 8
- 239000008367 deionised water Substances 0.000 claims description 8
- 229910021641 deionized water Inorganic materials 0.000 claims description 8
- 238000003756 stirring Methods 0.000 claims description 8
- VEXZGXHMUGYJMC-UHFFFAOYSA-M Chloride anion Chemical compound [Cl-] VEXZGXHMUGYJMC-UHFFFAOYSA-M 0.000 claims description 6
- 230000003213 activating effect Effects 0.000 claims description 6
- 239000003795 chemical substances by application Substances 0.000 claims description 6
- 239000012153 distilled water Substances 0.000 claims description 6
- 238000001914 filtration Methods 0.000 claims description 6
- 238000000227 grinding Methods 0.000 claims description 6
- 238000005470 impregnation Methods 0.000 claims description 6
- 239000012089 stop solution Substances 0.000 claims description 6
- 238000001132 ultrasonic dispersion Methods 0.000 claims description 6
- 238000005303 weighing Methods 0.000 claims description 6
- 235000017060 Arachis glabrata Nutrition 0.000 claims description 4
- 241001553178 Arachis glabrata Species 0.000 claims description 4
- 235000010777 Arachis hypogaea Nutrition 0.000 claims description 4
- 235000018262 Arachis monticola Nutrition 0.000 claims description 4
- 235000020232 peanut Nutrition 0.000 claims description 4
- 238000010298 pulverizing process Methods 0.000 claims description 4
- 238000007873 sieving Methods 0.000 claims description 4
- 239000002023 wood Substances 0.000 claims description 4
- 238000007789 sealing Methods 0.000 claims description 3
- 239000000428 dust Substances 0.000 claims description 2
- 238000011534 incubation Methods 0.000 claims description 2
- 238000000967 suction filtration Methods 0.000 claims description 2
- 238000002474 experimental method Methods 0.000 abstract description 10
- 238000005516 engineering process Methods 0.000 abstract description 6
- 239000007790 solid phase Substances 0.000 abstract description 6
- 230000035484 reaction time Effects 0.000 abstract description 4
- 238000011160 research Methods 0.000 abstract description 2
- 238000012360 testing method Methods 0.000 abstract description 2
- 239000002131 composite material Substances 0.000 abstract 1
- 238000007689 inspection Methods 0.000 abstract 1
- 239000000126 substance Substances 0.000 description 5
- 230000000903 blocking effect Effects 0.000 description 4
- 238000007865 diluting Methods 0.000 description 4
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 3
- 239000011324 bead Substances 0.000 description 3
- 229910052799 carbon Inorganic materials 0.000 description 3
- 239000002994 raw material Substances 0.000 description 3
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 230000028993 immune response Effects 0.000 description 2
- 239000002699 waste material Substances 0.000 description 2
- UGFAIRIUMAVXCW-UHFFFAOYSA-N Carbon monoxide Chemical compound [O+]#[C-] UGFAIRIUMAVXCW-UHFFFAOYSA-N 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 239000002154 agricultural waste Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000027455 binding Effects 0.000 description 1
- 229910002092 carbon dioxide Inorganic materials 0.000 description 1
- 239000001569 carbon dioxide Substances 0.000 description 1
- 229910002091 carbon monoxide Inorganic materials 0.000 description 1
- 230000003197 catalytic effect Effects 0.000 description 1
- 239000003085 diluting agent Substances 0.000 description 1
- 230000008105 immune reaction Effects 0.000 description 1
- 210000000987 immune system Anatomy 0.000 description 1
- 230000003053 immunization Effects 0.000 description 1
- 238000002649 immunization Methods 0.000 description 1
- 230000007774 longterm Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- HBMJWWWQQXIZIP-UHFFFAOYSA-N silicon carbide Chemical compound [Si+]#[C-] HBMJWWWQQXIZIP-UHFFFAOYSA-N 0.000 description 1
- 229910010271 silicon carbide Inorganic materials 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 230000009870 specific binding Effects 0.000 description 1
- 230000004936 stimulating effect Effects 0.000 description 1
- 230000009466 transformation Effects 0.000 description 1
- 210000004881 tumor cell Anatomy 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/531—Production of immunochemical test materials
- G01N33/532—Production of labelled immunochemicals
- G01N33/535—Production of labelled immunochemicals with enzyme label or co-enzymes, co-factors, enzyme inhibitors or enzyme substrates
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N21/78—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/536—Immunoassay; Biospecific binding assay; Materials therefor with immune complex formed in liquid phase
- G01N33/537—Immunoassay; Biospecific binding assay; Materials therefor with immune complex formed in liquid phase with separation of immune complex from unbound antigen or antibody
- G01N33/539—Immunoassay; Biospecific binding assay; Materials therefor with immune complex formed in liquid phase with separation of immune complex from unbound antigen or antibody involving precipitating reagent, e.g. ammonium sulfate
Landscapes
- Health & Medical Sciences (AREA)
- Immunology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Chemical & Material Sciences (AREA)
- Physics & Mathematics (AREA)
- Urology & Nephrology (AREA)
- Molecular Biology (AREA)
- Hematology (AREA)
- Biomedical Technology (AREA)
- General Physics & Mathematics (AREA)
- General Health & Medical Sciences (AREA)
- Pathology (AREA)
- Biochemistry (AREA)
- Analytical Chemistry (AREA)
- Biotechnology (AREA)
- Cell Biology (AREA)
- Microbiology (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Plasma & Fusion (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Solid-Sorbent Or Filter-Aiding Compositions (AREA)
Abstract
The invention relates to the technical field of immunology and immunodetection, and discloses a method for preparing a C-SiO-based solid-liquid composite material 2 Method for accelerating ELISA detection process by using adsorption material, specifically C-SiO 2 The adsorption material has good adsorptivity and strong mechanical property, disperses and adsorbs antigen and antibody molecules, improves the adsorption rate of antigen (or antibody) and a solid phase carrier, increases the contact probability of the antigen and antibody molecules, reduces the coating time of the antigen (or antibody) and the solid phase carrier and the reaction time of the antigen and the corresponding antibody, and plays a role in accelerating ELISA technology. The method is simple to operate, does not influence the reaction result, can accelerate the detection rate of ELISA technology, and provides convenience for scientific research experiments and inspection tests.
Description
Technical Field
The invention relates to the technical field of immunology and immunodetection, in particular to a method for detecting a tumor cell by using C-SiO 2 The adsorption material has the characteristics of good adsorptivity and high mechanical strength, so as to accelerate the detection process of ELISA (enzyme-linked immunosorbent assay).
Background
In recent decades, along with the rapid development of biomedical technology, the development of immunodetection technology has been promoted. In modern immunology, "immunization" is defined as: the body recognizes itself as the same as the own component and different from the own component, and generates a response. Wherein the substance capable of stimulating the immune system of the body to elicit an immune response is an antigen. Antibodies are the products of an immune response, with antibodies raised against different antigens. Antibodies bind only to specific antigens and form a coagulum, a process called specific antigen-antibody reaction, is the basis of immunodetection techniques.
The enzyme-linked immunosorbent assay (enzyme linked immunosorbent assay, ELISA) is also called as an ELISA, and is a qualitative and quantitative detection technology for antigens or antibodies widely applied in the biomedical field at present. The catalytic efficiency of the enzyme is very high, and the reaction result can be greatly amplified, so that the measuring method has very high sensitivity. ELISA can be used for both antigen and antibody detection. ELISA detection is prepared by binding soluble antigen or antibody to a solid support, and performing immune reaction by utilizing specific binding of the antigen and antibody. However, the antigen coating and reaction time are too long, so that the overall test time is long, and scientific research and detection personnel cannot obtain identification results in time.
Disclosure of Invention
The invention provides a method for preparing a silicon carbide film by using C-SiO 2 The invention relates to a method for accelerating ELISA detection process by adsorption material, which prepares C-SiO from carbon-containing substances and silicon tetrachloride 2 An adsorbent material. By combining C-SiO 2 Mixing the adsorption material with antigen and antibody diluent, and using C-SiO 2 The adsorption material has good adsorptivity and strong mechanical property, disperses and adsorbs antigen and antibody molecules, improves the adsorption rate of antigen (or antibody) and a solid phase carrier, increases the contact probability of the antigen and antibody molecules, reduces the coating time of the antigen (or antibody) and the solid phase carrier and the reaction time of the antigen and the corresponding antibody, and plays a role in accelerating ELISA technology.
The specific technical scheme of the invention is as follows: by using C-SiO 2 A method for accelerating an ELISA detection process with an adsorbent material comprising the steps of:
step 1: pulverizing and drying carbonaceous materials by a ball mill, sieving, placing into a container, adding deionized water, performing ultrasonic dispersion, mixing uniformly, and standing at room temperature overnight to fully impregnate the carbonaceous materials.
After being crushed, dried and fully immersed, the carbon-containing substances and silicon tetrachloride can fully react, and the utilization rate of raw materials is improved.
Step 2: after full impregnation, dropwise adding silicon tetrachloride into the container, and continuously stirring at the same time; continuously stirring the obtained mixture and standing at room temperature to fully react; taking out, suction filtering for many times, and washing filter residue with distilled water until no chloride ion remains.
Step 3: activating the filter residue obtained in the step 2 in a muffle furnace at 200-300 ℃ for 0.5-3h, taking out, and fully drying to obtain C-SiO 2 An adsorbent material.
Step 4: weighing the C-SiO prepared in the step 3 2 Two parts of adsorption material are respectively placed in two containers, respectively added with buffer solutions corresponding to antigen and antibody, and oscillated to make them fully dissolved so as to obtain the invented product containing C-SiO 2 A dilution of the adsorbent material.
Step 5: adding corresponding C-SiO-containing solution obtained in step 4 into antigen and antibody solution required by ELISA 2 The dilution of the adsorbent material was shaken to dissolve it thoroughly.
Step 6: in ELISA, antigen or antibody and C-SiO are added into the ELISA plate 2 Coating the mixture of (2) at 1-5 ℃ for 1-3h; removing liquid in the holes, and washing the holes with washing liquid for a plurality of times; adding the sealing liquid, and then placing the mixture in a baking oven at the temperature of 35-40 ℃ for incubation for 0.5-1.5h; removing liquid in the holes, and washing the holes with washing liquid for a plurality of times; dripping corresponding antibody or antigen C-SiO 2 Placing the mixture in an oven at 35-40 ℃ for 20-40min; removing the liquid in the holes again, and washing the holes with washing liquid for a plurality of times; adding TMB color-developing agent into each hole for shading reaction for 5-15min, then dripping stop solution, and measuring OD value of each hole at 450nm within 10-20 min.
Preferably, in step 1: the carbonaceous material is wood dust and/or peanut shell.
Preferably, in step 1: the ball-material ratio of the ball mill is 1:12-16, the types of the three grinding balls are 2 mm, 6 mm and 10 mm, the number ratio is 6:2-3:1, and the setting frequency of the ball mill is 15-20 Hz; pulverizing, drying, and sieving with ten mesh sieve.
Preferably, in step 1: the solid-to-liquid ratio of the carbonaceous material to the deionized water is (1-2 g): (10-20 mL); the ultrasonic time is 10-30min.
Preferably, in step 2: the solid-to-liquid ratio of the carbonaceous material and the silicon tetrachloride added dropwise is (4-5 g): (3-4 mL); the feeding time is controlled within 1-2 h.
Preferably, in step 2: the room temperature standing time is 2-4h, and the suction filtration times are 3-5 times.
Preferably, in step 4: every part of C-SiO 2 The adsorption material is 0.8-1.2g, and 80-120mL of antigen or antibody corresponding buffer solution is added into each container.
Preferably, in step 5, the antigen, antibody solution and C-SiO-containing solution obtained in step 4 2 The dilution liquid mixing ratio of the adsorption material is respectively 1:1-2.5.
Compared with the prior art, the invention has the beneficial effects that:
(1) The invention utilizes C-SiO 2 The adsorption material has good adsorptivity and strong mechanical property, disperses and adsorbs antigen and antibody molecules, improves the adsorption rate of antigen (or antibody) and a solid phase carrier, increases the contact probability of the antigen and antibody molecules, reduces the coating time of the antigen (or antibody) and the solid phase carrier and the reaction time of the antigen and the corresponding antibody, and plays a role in accelerating the ELISA detection process.
(2) The invention is used for preparing C-SiO 2 The carbon-containing substances used in the adsorption material are agricultural wastes, and a large amount of polluted air such as carbon monoxide, carbon dioxide and the like can be generated by direct incineration. The reutilization of the waste can effectively utilize the waste materials.
(3) The C-SiO prepared by the invention 2 The adsorption material can be stored for a long time after being completely dried in a sealing way, thereby providing convenience for long-term experiments and detection.
Detailed Description
The invention is further described below with reference to examples.
Example 1
Step 1: 20g of carbonaceous material (wood chips) was crushed and dried by a ball mill, sieved by a ten-mesh sieve, put into a beaker and added with 200mL of deionized water for ultrasonic dispersion for 20min. After mixing well, left overnight at room temperature, allow to impregnate thoroughly. Wherein, the ball-material ratio of the ball mill is 1:14, the three grinding bead types are 2 mm, 6 mm and 10 mm, the number ratio is 6:2.5:1, and the setting frequency of the ball mill is 18 Hz.
Step 2: after full impregnation, 15mL of silicon tetrachloride was added dropwise to the beaker while stirring continuously, and the addition time was controlled at 1.5h. The product mixture was stirred and left at room temperature for 3h to allow it to react well. Taking out, suction filtering for 4 times, and washing filter residue with distilled water until no chloride ion remains.
Step 3: activating the filter residue obtained in the step 3 in a muffle furnace at 250 ℃ for 2 hours, taking out, and fully drying to obtain C-SiO 2 An adsorbent material.
Step 4: weighing the C-SiO prepared in the step 3 2 Two parts of adsorption material, each 1g, are respectively placed in 2 200mL beakers, 100mL of buffer solution corresponding to antigen and antibody is respectively added into the beakers, and the beakers are oscillated to be fully dissolved, thus obtaining the C-SiO-containing beakers 2 A dilution of the adsorbent material.
Step 5: adding the C-SiO-containing solution prepared in the step 4 into the antigen and antibody solution required by ELISA experiments 2 Diluting solution of adsorption material, antigen and antibody solution and C-SiO-containing solution prepared in step 4 2 The dilution ratio of the adsorbent material was 1:1, shaking to make it fully dissolved.
Step 6: during ELISA experiments, antigen (or antibody) C-SiO is added into the ELISA plate 2 The mixture was coated for 2h at 4 ℃. The liquid in the wells was removed and washed three times with wash liquid. After addition of the blocking solution, the mixture was placed in an oven at 37℃and incubated for 1h. The liquid in the wells was removed and washed three times with wash liquid. Dripping corresponding antibody (or antigen) C-SiO 2 The mixture was placed in an oven at 37℃for 30min. The wells were again freed from liquid and washed five times with washing liquid. After adding TMB color-developing agent into each hole for shading reaction for 10min, adding stop solution dropwise, and measuring OD value of each hole at 450nm within 15 min.
Example 2
Step 1: 20g of carbonaceous material (peanut shell) was crushed and dried by a ball mill, sieved by a ten-mesh sieve, placed in a beaker and added with 200mL of deionized water for ultrasonic dispersion for 20min. After mixing well, left overnight at room temperature, allow to impregnate thoroughly. Wherein, the ball-material ratio of the ball mill is 1:12, the three grinding bead types are 2 mm, 6 mm and 10 mm, the number ratio is 6:2:1, and the setting frequency of the ball mill is 15 Hz.
Step 2: after full impregnation, 15mL of silicon tetrachloride was added dropwise to the beaker while stirring continuously, and the addition time was controlled at 1h. The product mixture was stirred and left at room temperature for 3h to allow it to react well. Taking out, filtering for 3 times, and washing filter residue with distilled water until no chloride ion remains.
Step 3: activating the filter residue obtained in the step 3 in a muffle furnace at 200 ℃ for 2 hours, taking out, and fully drying to obtain C-SiO 2 An adsorbent material.
Step 4: weighing the C-SiO prepared in the step 3 2 Two parts of adsorption material, each 1g, are respectively placed in 2 200mL beakers, 100mL of buffer solution corresponding to antigen and antibody is respectively added into the beakers, and the beakers are oscillated to be fully dissolved, thus obtaining the C-SiO-containing beakers 2 A dilution of the adsorbent material.
Step 5: adding the C-SiO-containing solution prepared in the step 4 into the antigen and antibody solution required by ELISA experiments 2 Diluting solution of adsorption material, antigen and antibody solution and C-SiO-containing solution prepared in step 4 2 The dilution ratio of the adsorbent material was 1:1.5, shaking to make it fully dissolved.
Step 6: during ELISA experiments, antigen (or antibody) C-SiO is added into the ELISA plate 2 The mixture was coated for 2h at 4 ℃. The liquid in the wells was removed and washed three times with wash liquid. After addition of the blocking solution, the mixture was placed in an oven at 37℃and incubated for 1h. The liquid in the wells was removed and washed three times with wash liquid. Dripping corresponding antibody (or antigen) C-SiO 2 The mixture was placed in an oven at 37℃for 30min. The wells were again freed from liquid and washed five times with washing liquid. After adding TMB color-developing agent into each hole for shading reaction for 10min, adding stop solution dropwise, and measuring OD value of each hole at 450nm within 15 min.
Example 3
Step 1: 20g of carbonaceous material (wood chips) was crushed and dried by a ball mill, sieved by a ten-mesh sieve, put into a beaker and added with 200mL of deionized water for ultrasonic dispersion for 20min. After mixing well, left overnight at room temperature, allow to impregnate thoroughly. Wherein, the ball-material ratio of the ball mill is 1:16, the three grinding bead types are 2 mm, 6 mm and 10 mm, the number ratio is 6:3:1, and the setting frequency of the ball mill is 20 Hz.
Step 2: after full impregnation, 15mL of silicon tetrachloride was added dropwise to the beaker while stirring continuously, and the addition time was controlled at 2h. The product mixture was stirred and left at room temperature for 3h to allow it to react well. Taking out, suction filtering for 5 times, and washing filter residue with distilled water until no chloride ion remains.
Step 3: activating the filter residue obtained in the step 3 in a muffle furnace at 300 ℃ for 2 hours, taking out, and fully drying to obtain C-SiO 2 An adsorbent material.
Step 4: weighing the C-SiO prepared in the step 3 2 Two parts of adsorption material, each 1g, are respectively placed in 2 200mL beakers, 100mL of buffer solution corresponding to antigen and antibody is respectively added into the beakers, and the beakers are oscillated to be fully dissolved, thus obtaining the C-SiO-containing beakers 2 A dilution of the adsorbent material.
Step 5: adding the C-SiO-containing solution prepared in the step 4 into the antigen and antibody solution required by ELISA experiments 2 Diluting solution of adsorption material, antigen and antibody solution and C-SiO-containing solution prepared in step 4 2 The dilution ratio of the adsorbent material was 1:2, oscillating to make it fully dissolved.
Step 6: during ELISA experiments, antigen (or antibody) C-SiO is added into the ELISA plate 2 The mixture was coated for 2h at 4 ℃. The liquid in the wells was removed and washed three times with wash liquid. After addition of the blocking solution, the mixture was placed in an oven at 37℃and incubated for 1h. The liquid in the wells was removed and washed three times with wash liquid. Dripping corresponding antibody (or antigen) C-SiO 2 The mixture was placed in an oven at 37℃for 30min. The wells were again freed from liquid and washed five times with washing liquid. After adding TMB color-developing agent into each hole for shading reaction for 10min, adding stop solution dropwise, and measuring OD value of each hole at 450nm within 15 min.
Example 4
Step 1: 20g of carbonaceous material (peanut shell) was crushed and dried by a ball mill, sieved by a ten-mesh sieve, placed in a beaker and added with 200mL of deionized water for ultrasonic dispersion for 20min. After mixing well, left overnight at room temperature, allow to impregnate thoroughly. Wherein, the ball-material ratio of the ball mill is 1:16, the three grinding ball types are 2 mm, 6 mm and 10 mm, the number ratio is 6:3:1, and the setting frequency of the ball mill is 15 Hz.
Step 2: after full impregnation, 15mL of silicon tetrachloride was added dropwise to the beaker while stirring continuously, and the addition time was controlled at 1h. The product mixture was stirred and left at room temperature for 3h to allow it to react well. Taking out, filtering for 3 times, and washing filter residue with distilled water until no chloride ion remains.
Step 3: activating the filter residue obtained in the step 3 in a muffle furnace at 260 ℃ for 2 hours, taking out, and fully drying to obtain C-SiO 2 An adsorbent material.
Step 4: weighing the C-SiO prepared in the step 3 2 Two parts of adsorption material, each 1g, are respectively placed in 2 200mL beakers, 100mL of buffer solution corresponding to antigen and antibody is respectively added into the beakers, and the beakers are oscillated to be fully dissolved, thus obtaining the C-SiO-containing beakers 2 A dilution of the adsorbent material.
Step 5: adding the C-SiO-containing solution prepared in the step 4 into the antigen and antibody solution required by ELISA experiments 2 Diluting solution of adsorption material, antigen and antibody solution and C-SiO-containing solution prepared in step 4 2 The dilution ratio of the adsorbent material was 1:2.5, oscillating to make it fully dissolved.
Step 6: during ELISA experiments, antigen (or antibody) C-SiO is added into the ELISA plate 2 The mixture was coated for 2h at 4 ℃. The liquid in the wells was removed and washed three times with wash liquid. After addition of the blocking solution, the mixture was placed in an oven at 37℃and incubated for 1h. The liquid in the wells was removed and washed three times with wash liquid. Dripping corresponding antibody (or antigen) C-SiO 2 The mixture was placed in an oven at 37℃for 30min. The wells were again freed from liquid and washed five times with washing liquid. After adding TMB color-developing agent into each hole for shading reaction for 10min, adding stop solution dropwise, and measuring OD value of each hole at 450nm within 15 min.
The raw materials and equipment used in the invention are common raw materials and equipment in the field unless specified otherwise; the methods used in the present invention are conventional in the art unless otherwise specified.
The foregoing description is only a preferred embodiment of the present invention, and is not intended to limit the present invention, and any simple modification, variation and equivalent transformation of the above embodiment according to the technical substance of the present invention still fall within the scope of the technical solution of the present invention.
Claims (4)
1. By using C-SiO 2 The method for accelerating ELISA detection process by the adsorption material is characterized by comprising the following steps:
step 1: pulverizing and drying carbonaceous materials by a ball mill, sieving, placing the carbonaceous materials into a container, adding deionized water for ultrasonic dispersion, and standing overnight at room temperature after uniform mixing to fully impregnate the carbonaceous materials; the solid-to-liquid ratio of the carbonaceous material to the deionized water is (1-2 g): (10-20 mL); the ultrasonic time is 10-30min;
step 2: after full impregnation, dropwise adding silicon tetrachloride into the container, and continuously stirring at the same time; continuously stirring the obtained mixture and standing at room temperature to fully react; taking out, suction filtering for many times, and washing filter residues with distilled water until no chloride ion remains; the solid-to-liquid ratio of the carbonaceous material and the silicon tetrachloride added dropwise is (4-5 g): (3-4 mL); controlling the feeding time to be 1-2 h;
step 3: activating the filter residue obtained in the step 2 in a muffle furnace at 200-300 ℃ for 0.5-3h, taking out, and fully drying to obtain C-SiO 2 An adsorption material;
step 4: weighing the C-SiO prepared in the step 3 2 Two parts of adsorption material are respectively placed in two containers, respectively added with buffer solutions corresponding to antigen and antibody, and oscillated to make them fully dissolved so as to obtain the invented product containing C-SiO 2 A dilution of the adsorbent material; every part of C-SiO 2 The adsorption material is 0.8-1.2g, and 80-120mL of buffer solution corresponding to the antigen or the antibody is added into each container;
step 5: adding corresponding C-SiO-containing solution obtained in step 4 into antigen and antibody solution required by ELISA 2 Oscillating the dilution of the adsorption material to fully dissolve the adsorption material; antigen, antibody solution and C-SiO-containing solution obtained in step 4 2 The dilution liquid mixing ratio of the adsorption material is respectively 1:1-2.5;
step 6: in ELISA, antigen or antibody and C-SiO are added into the ELISA plate 2 Coating the mixture of (2) at 1-5 ℃ for 1-3h; removing liquid in the holes, and washing the holes with washing liquid for a plurality of times; adding the sealing liquid, and then placing the mixture in a baking oven at the temperature of 35-40 ℃ for incubation for 0.5-1.5h; removing liquid in the holes, and washing the holes with washing liquid for a plurality of times; dripping corresponding antibody or antigen C-SiO 2 Placing the mixture in an oven at 35-40 ℃ for 20-40min; removing the liquid in the holes again, and washing the holes with washing liquid for a plurality of times; adding TMB color-developing agent into each hole for shading reaction for 5-15min, then dripping stop solution, and measuring OD value of each hole at 450nm within 10-20 min.
2. The method according to claim 1, wherein in step 1: the carbonaceous material is wood dust and/or peanut shell.
3. The method according to claim 1, wherein in step 1: the ball-material ratio of the ball mill is 1:12-16, the types of the three grinding balls are 2 mm, 6 mm and 10 mm, the number ratio is 6:2-3:1, and the setting frequency of the ball mill is 15-20 Hz; pulverizing, drying, and sieving with ten mesh sieve.
4. The method according to claim 1, wherein in step 2: the room temperature standing time is 2-4h, and the suction filtration times are 3-5 times.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110691502.3A CN113447646B (en) | 2021-06-22 | 2021-06-22 | By using C-SiO 2 Method for accelerating ELISA detection process by adsorption material |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110691502.3A CN113447646B (en) | 2021-06-22 | 2021-06-22 | By using C-SiO 2 Method for accelerating ELISA detection process by adsorption material |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN113447646A CN113447646A (en) | 2021-09-28 |
| CN113447646B true CN113447646B (en) | 2024-03-19 |
Family
ID=77812058
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202110691502.3A Active CN113447646B (en) | 2021-06-22 | 2021-06-22 | By using C-SiO 2 Method for accelerating ELISA detection process by adsorption material |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN113447646B (en) |
Citations (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2004088315A1 (en) * | 2003-03-31 | 2004-10-14 | Council Of Scientific And Industrial Research | Rapid heat-mediated method for enzyme-linked immunosorbent assay procedure |
| RU2265611C1 (en) * | 2004-06-30 | 2005-12-10 | Федеральное государственное образовательное учреждение высшего профессионального образования Ставропольский государственный аграрный университет | Method for preparing immunosorbent |
| CN101504409A (en) * | 2008-12-31 | 2009-08-12 | 沈阳农业大学 | Fast detection ELISA reagent kit for methomyl residue |
| CN101785990A (en) * | 2010-02-09 | 2010-07-28 | 乐山师范学院 | Method for preparing activated carbon-silicon dioxide mixed adsorbing material by using silicon tetrachloride |
| CN102435729A (en) * | 2011-12-02 | 2012-05-02 | 杭州迪恩科技有限公司 | One-step enzyme-linked immunoassay method for quickly detecting melamine and kit thereof |
| CN102649047A (en) * | 2012-05-17 | 2012-08-29 | 乐山川博吸附材料有限责任公司 | Preparation method for C-SiO2 adsorbing materials |
| CN103175955A (en) * | 2011-12-26 | 2013-06-26 | 中国人民解放军军事医学科学院卫生学环境医学研究所 | Direct-coating ELISA (enzyme-linked immuno sorbent assay) method for rapid detection of chloromycetin |
| CN104888530A (en) * | 2015-05-20 | 2015-09-09 | 蚌埠德美过滤技术有限公司 | Active carbon air-conditioner filter core material and preparation method thereof |
| CN107064487A (en) * | 2017-02-14 | 2017-08-18 | 安徽雷根生物技术有限公司 | A kind of graphene accelerates elisa technique |
| CN108404879A (en) * | 2018-04-25 | 2018-08-17 | 苏州凌科特新材料有限公司 | A kind of preparation method of high-performance Dye Adsorption material |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20040191831A1 (en) * | 2003-03-25 | 2004-09-30 | Council Of Scientific And Industrial Research | Rapid heat - mediated method for enzyme - linked immunosorbent assay procedure |
| US7824867B2 (en) * | 2007-06-18 | 2010-11-02 | Genscript Holdings (Hong Kong) Limited | Rapid ELISA processes and related compositions |
-
2021
- 2021-06-22 CN CN202110691502.3A patent/CN113447646B/en active Active
Patent Citations (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2004088315A1 (en) * | 2003-03-31 | 2004-10-14 | Council Of Scientific And Industrial Research | Rapid heat-mediated method for enzyme-linked immunosorbent assay procedure |
| RU2265611C1 (en) * | 2004-06-30 | 2005-12-10 | Федеральное государственное образовательное учреждение высшего профессионального образования Ставропольский государственный аграрный университет | Method for preparing immunosorbent |
| CN101504409A (en) * | 2008-12-31 | 2009-08-12 | 沈阳农业大学 | Fast detection ELISA reagent kit for methomyl residue |
| CN101785990A (en) * | 2010-02-09 | 2010-07-28 | 乐山师范学院 | Method for preparing activated carbon-silicon dioxide mixed adsorbing material by using silicon tetrachloride |
| CN102435729A (en) * | 2011-12-02 | 2012-05-02 | 杭州迪恩科技有限公司 | One-step enzyme-linked immunoassay method for quickly detecting melamine and kit thereof |
| CN103175955A (en) * | 2011-12-26 | 2013-06-26 | 中国人民解放军军事医学科学院卫生学环境医学研究所 | Direct-coating ELISA (enzyme-linked immuno sorbent assay) method for rapid detection of chloromycetin |
| CN102649047A (en) * | 2012-05-17 | 2012-08-29 | 乐山川博吸附材料有限责任公司 | Preparation method for C-SiO2 adsorbing materials |
| CN104888530A (en) * | 2015-05-20 | 2015-09-09 | 蚌埠德美过滤技术有限公司 | Active carbon air-conditioner filter core material and preparation method thereof |
| CN107064487A (en) * | 2017-02-14 | 2017-08-18 | 安徽雷根生物技术有限公司 | A kind of graphene accelerates elisa technique |
| CN108404879A (en) * | 2018-04-25 | 2018-08-17 | 苏州凌科特新材料有限公司 | A kind of preparation method of high-performance Dye Adsorption material |
Also Published As
| Publication number | Publication date |
|---|---|
| CN113447646A (en) | 2021-09-28 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Huang et al. | Hg (II) removal from aqueous solution by bayberry tannin-immobilized collagen fiber | |
| CN103949115B (en) | A kind of method of in-situ preparation denitrating catalyst on filtrate | |
| CN109097591B (en) | Calcium alginate immobilized microbial adsorbent, preparation method thereof and application thereof in recycling platinum group metal secondary resources | |
| CN106268707A (en) | A kind of phosphoeptide based on novel magnetic porous material enrichment new method | |
| CN107537451A (en) | A kind of porous zeolite material for air purification and preparation method thereof | |
| CN106824069B (en) | For handling the preparation method of the rear-earth-doped iron Carbon Materials of arsenic-containing waste water | |
| US5470532A (en) | Composite reactive articles for the determination of cyanide | |
| CN113447646B (en) | By using C-SiO 2 Method for accelerating ELISA detection process by adsorption material | |
| CN101498719B (en) | Production method for enzyme functionalized nano immunity marker and use thereof | |
| CN110124641A (en) | A kind of radionuclide adsorbent material and its preparation method and application | |
| CN107469781A (en) | A kind of graphene oxide beta cyclodextrin aeroge sorbing materials of reduction and preparation method thereof | |
| CN112516968A (en) | Preparation method of metal organic framework material loaded modified diatomite adsorbent | |
| CN102553539A (en) | Biomass modified adsorbent capable of adsorbing cadmium ions and preparation method and application thereof | |
| CN109211651B (en) | Method for rapidly extracting arsenic and selenium in different forms in fly ash | |
| CN114849668A (en) | Preparation method and carbon neutralization application of high-hydrophobicity acid-corrosion-resistant MOFs-based composite material | |
| CN110368900A (en) | A kind of bamboo charcoal modified material and its preparation method and application | |
| CN110090627A (en) | A kind of calcium ion blotting chitosan adsorbent and the preparation method and application thereof | |
| CN110938040B (en) | Melamine hapten and artificial antibody as well as preparation method and application thereof | |
| CN115356477B (en) | Streptavidin magnetic bead and preparation method and application thereof | |
| Chen et al. | Adsorption of Pb (II) from aqueous solution by mercerized Moso bamboo chemically modified with pyromellitic dianhydride | |
| CN116375593A (en) | Metal organic frame material for iodine adsorption, ligand and application | |
| CN110465277A (en) | A kind of ammonia molecular imprinting adsorbing agent and its preparation for the separation of sludge aerobic compost mixed gas and Ammonia recovery | |
| JP2001213614A (en) | Functional carbonized material and production process thereof | |
| CN113061330A (en) | Soil heavy metal removal composite material and preparation method and application thereof | |
| CN106589241B (en) | A kind of terpolymer hydrogel and its preparation method and application |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |