CN108761060A - A kind of excellent effect mercury chelating type immune complex and preparation method thereof - Google Patents
A kind of excellent effect mercury chelating type immune complex and preparation method thereof Download PDFInfo
- Publication number
- CN108761060A CN108761060A CN201810594224.8A CN201810594224A CN108761060A CN 108761060 A CN108761060 A CN 108761060A CN 201810594224 A CN201810594224 A CN 201810594224A CN 108761060 A CN108761060 A CN 108761060A
- Authority
- CN
- China
- Prior art keywords
- mercury
- immune complex
- solution
- carrier protein
- chelating type
- 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.)
- Pending
Links
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
Landscapes
- Health & Medical Sciences (AREA)
- Immunology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Molecular Biology (AREA)
- Biomedical Technology (AREA)
- Chemical & Material Sciences (AREA)
- Hematology (AREA)
- Urology & Nephrology (AREA)
- Biotechnology (AREA)
- Microbiology (AREA)
- Cell Biology (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The invention discloses a kind of excellent effect mercury chelating type immune complex, which is following one kind：Mercury ion is incorporated into the compound of immune complex formation；Or mercury ion be incorporated into after carrier protein with and the antibody that specifically binds of the carrier protein be formed by compound；Or mercury ion be incorporated into immunoglobulin after the compound to be formed is combined with carrier protein.The invention also discloses a kind of preparation methods of excellent effect mercury chelating type immune complex, include the following steps：S1：Prepare chelating agent solution, S2：Formulation vehicle protein solution, S3：It is stirred overnight, S4：Dialysis treatment, S5：Mercury ion, S6 is added：Devil liquor recovery processing；S7：It is specifically bound.The method of the present invention scope of application is wider, can be cost-effective, and improves dialysis rate, can shorten manufacturing cycle, also have the characteristics that energy conservation and environmental protection, avoids causing chemical contamination, environment protecting good.
Description
Technical field
The present invention relates to a kind of chelate field, specially a kind of excellent effect mercury chelating type immune complex and its preparation
Method.
Background technology
Chelate Mineral Floating Process, hydrometallurgy, metallic element extraction with detach, the catalyzing and synthesizing of substance, water
Softening, electroplating technology, medical industry, dyeing course etc. in be all widely used；Existing mercury chelating type immune complex
Preparation method it is incomplete, the scope of application is ignored enough wide, it has not been convenient to the preparation process of object is completed in small-size laboratory,
And number of dialysing is excessive, can extend the manufacturing cycle of object to a certain extent, and there are no give up to the ion etc. in waste liquid
Object, which is done, clearly to be handled, and the wastes such as ion in waste liquid, which do not handle rear discharge up to standard, can cause chemical contamination.
Invention content
The purpose of the present invention is to provide a kind of excellent effect mercury chelating type immune complexs and preparation method thereof, to solve
The problems mentioned above in the background art.
To achieve the above object, the present invention provides the following technical solutions：A kind of excellent effect mercury chelating type immune complex,
The mercury chelating type immune complex is following one kind:
Mercury ion is incorporated into the compound of immune complex formation；
Or mercury ion be incorporated into after carrier protein with and the antibody that specifically binds of the carrier protein be formed by compound；
Or mercury ion be incorporated into immunoglobulin after the compound to be formed is combined with carrier protein.
Preferably, zinc fingers, sulfydryl, cysteine are at least contained in the structure of the immune complex or carrier protein
One kind in residue, mercury ion are mutually tied with immune complex or carrier protein with zinc fingers or sulfydryl or cysteine residues
It closes.
Preferably, the carrier protein is antibody protein, lipoprotein, hemoglobin, nuclear antigen, foreign sera antigen, disease
One kind among poison, bacterium, protozoon, worm or immunoglobulin.
Preferably, it is specifically bound again with energy and carrier protein after mercury ion is combined by chelating agent with carrier protein anti-
Body combines, and the chelating agent is one kind in ITCBE, EDTA, polyphosphate, amino carboxylic acid, 1,3- diketone, hydroxycarboxylic acid.
A kind of preparation method of mercury chelating type immune complex as described in claim 1, includes the following steps：
S1：First chelating agent solution is prepared and is completed, then chelating agent is dissolved in solvent, you can obtains chelating agent solution；
S2：Appropriate carrier protein is taken, carrier protein is added in borate buffer solution, you can it is molten that carrier protein is made
Liquid；
S3：Chelating agent solution obtained is added in carrier protein solution, is placed in water bath with thermostatic control and stirs evenly,
Water bath with thermostatic control 4-30h, you can obtain mixed liquor；
S4：It fetches bag filter and EDTA-NaHCO is added in bag filter3Solution boils, and bag filter is fixed on water tank
Middle part, and enough ddH are added in water tank2O, the ddH2The volume ratio of solution is 80-85 in the volume and bag filter of O:1, weight
Multiple step 2-4 times, the mixing in S3 is fitted into bag filter, bag filter is placed in constant temperature water tank, is added in constant temperature water tank
Enter enough ddH2O, the ddH2The volume ratio of solution is 80-85 in the volume and bag filter of O:1, after dialysed overnight, it will dialyse
Liquid in bag is collected.
S5：PH to 7.0 is adjusted dilute hydrochloric acid solution is added from the liquid collected in bag filter, it is molten to be slowly dropped into mercury ion
Liquid, when instillation, do not stop to shake, and instill and complete to be placed in water bath with thermostatic control and stir evenly, water bath with thermostatic control 4-30h, then repeatedly S4
Once, liquid is collected, mercury chelating type antigen is obtained.
S6：The waste liquid collected in solution and S5 in S4 outside bag filter is mixed, is added into mixed solution
Enter activated carbon to be adsorbed, and repeat to add above-mentioned absorbent charcoal material, reuses atomic fluorescence method and processed solution is carried out
Quantitative analysis, when mercury ion enrichment degree reaches 0.1mol/ml in solution, recycling mercury ion recycles.
S7：The antibody that can be specifically bound with carrier protein is added in above-mentioned mercury chelating type antigen, mercury is obtained after reaction
Chelating type immune complex.
The present invention also provides above-mentioned mercury chelating type immune complexs to prepare detection mercury chelating type CIC ELISA
Reagent or enzyme linked immunological kit in application.
The present invention also provides a kind of above-mentioned enzyme linked immunological kit of detection mercury chelating type CIC ELISA, the examinations
Agent box includes the standard items of above-mentioned mercury chelating type immune complex.
Preferably, mentioned reagent box further includes at least one following reagent：Albumen containing capture antigen antibody complex
Coating buffer, the coating buffer of antibody containing capture mercury, enzyme labelled antibody.
A method of mercury chelating type CIC ELISA is quantitatively detected, it is immune with the above-mentioned mercury chelating type of known content
Compound is detected as standard items using a pair of sample of following methods：Enzyme-linked immunization, enzyme linked immunological and Atomic absorption
Spectrum combined techniques, enzyme linked immunological and inductivity coupled plasma mass spectrometry combined techniques, purification immune complex and atomic absorption spectrum
Combined techniques, purification immune complex and inductivity coupled plasma mass spectrometry combined techniques, electrophoresis and enzyme linked immunological or atomic absorption light
Spectrum or inductivity coupled plasma mass spectrometry combined techniques.
Compared with prior art, the beneficial effects of the invention are as follows：The method of the present invention scope of application is wider, can save into
This, and increase the differential water pressures inside and outside bag filter, dialysis rate is improved, manufacturing cycle can be shortened, also to the ion in waste liquid
Equal wastes are recycled, and have the characteristics that energy conservation and environmental protection, avoid causing chemical contamination, therefore the present invention has larger city
Field competitiveness.
Description of the drawings
Fig. 1 is the flow diagram of the present invention.
Specific implementation mode
The technical scheme in the embodiments of the invention will be clearly and completely described below, it is clear that described implementation
Example is only a part of the embodiment of the present invention, instead of all the embodiments.Based on the embodiments of the present invention, this field is common
The every other embodiment that technical staff is obtained without making creative work belongs to the model that the present invention protects
It encloses.
The present invention provides a kind of technical solution：A kind of excellent effect mercury chelating type immune complex, the mercury chelating type are immune
Compound is following one kind:
Mercury ion is incorporated into the compound of immune complex formation；
Or mercury ion be incorporated into after carrier protein with and the antibody that specifically binds of the carrier protein be formed by compound；
Or mercury ion be incorporated into immunoglobulin after the compound to be formed is combined with carrier protein.
Further, it is residual that zinc fingers, sulfydryl, cysteine are at least contained in the structure of immune complex or carrier protein
One kind in base, mercury ion are combined with immune complex or carrier protein with zinc fingers or sulfydryl or cysteine residues.
Further, it is residual that zinc fingers, sulfydryl, cysteine are at least contained in the structure of immune complex or carrier protein
One kind in base, mercury ion are combined with immune complex or carrier protein with zinc fingers or sulfydryl or cysteine residues.
Further, carrier protein be antibody protein, lipoprotein, hemoglobin, nuclear antigen, foreign sera antigen, virus,
One kind among bacterium, protozoon, worm or immunoglobulin.
A kind of preparation method of mercury chelating type immune complex as claimed in claim 1, includes the following steps：
S1：First chelating agent solution is prepared and is completed, then chelating agent is dissolved in solvent, you can obtains chelating agent solution；
S2：Appropriate carrier protein is taken, carrier protein is added in borate buffer solution, you can it is molten that carrier protein is made
Liquid；
S3：Chelating agent solution obtained is added in carrier protein solution, is placed in water bath with thermostatic control and stirs evenly,
Water bath with thermostatic control is for 24 hours, you can obtains mixed liquor；
S4：It fetches bag filter and EDTA-NaHCO is added in bag filter3Solution boils, and bag filter is fixed on water tank
Middle part, and enough ddH are added in water tank2O, the ddH2The volume ratio of solution is 83 in the volume and bag filter of O:1, it repeats
Mixing in S3 is fitted into bag filter, bag filter is placed in constant temperature water tank by the step 2 time, and foot is added in constant temperature water tank
The ddH of amount2O, the ddH2The volume ratio of solution is 83 in the volume and bag filter of O:1, it, will be in bag filter after dialysed overnight
Liquid is collected.
S5：PH to 7.0 is adjusted dilute hydrochloric acid solution is added from the liquid collected in bag filter, it is molten to be slowly dropped into mercury ion
Liquid, when instillation, do not stop to shake, and instill and complete to be placed in water bath with thermostatic control and stir evenly, water bath with thermostatic control for 24 hours, then repeatedly S4 mono-
It is secondary, liquid is collected, mercury chelating type antigen is obtained.
S6：The waste liquid collected in solution and S5 in S4 outside bag filter is mixed, is added into mixed solution
Enter activated carbon to be adsorbed, and repeat to add above-mentioned absorbent charcoal material, reuses atomic fluorescence method and processed solution is carried out
Quantitative analysis, when mercury ion enrichment degree reaches 0.1mol/ml in solution, recycling mercury ion recycles.
S7：The antibody that can be specifically bound with carrier protein is added in above-mentioned mercury chelating type antigen, mercury is obtained after reaction
Chelating type immune complex.
The present invention also provides above-mentioned mercury chelating type immune complexs to prepare detection mercury chelating type CIC ELISA
Reagent or enzyme linked immunological kit in application.
The present invention also provides a kind of above-mentioned enzyme linked immunological kit of detection mercury chelating type CIC ELISA, the examinations
Agent box includes the standard items of above-mentioned mercury chelating type immune complex.
Mentioned reagent box further includes at least one following reagent：The coating of albumen containing capture antigen antibody complex
Liquid, the coating buffer containing the antibody for capturing mercury, enzyme labelled antibody.
In the present invention, it is following several to realize that the kit of the object of the invention can be listed, but it is not limited to this.
A kind of kit for detecting mercury chelating type CIC ELISA, including：It is multiple containing antigen-antibody can be captured
The coating buffer of the albumen of conjunction object, confining liquid, washing buffer, anti-mercury antibody, enzyme labelled antibody, substrate, terminate liquid, dilution buffer
Liquid, positive control, negative control.
A kind of kit for detecting mercury chelating type CIC ELISA, including：It is multiple containing antigen-antibody can be captured
The coating buffer of the albumen of conjunction object, confining liquid, washing buffer, eluent, positive control, negative control.
A kind of kit for detecting mercury chelating type CIC ELISA, including：It is multiple containing antigen-antibody can be captured
The coating buffer of the albumen of conjunction object, confining liquid, washing buffer, eluent, nitric acid, hydrogen peroxide, positive control, negative control.
A kind of kit for detecting mercury chelating type CIC ELISA, including be used to extract needed for immune complex
Solution redissolves liquid, the coating buffer containing the antibody that can capture mercury, confining liquid, washing buffer, enzyme labelled antibody, substrate, termination
Liquid, dilution buffer, standard items, negative control.
A kind of kit for detecting mercury chelating type CIC ELISA：Including being used to extract needed for immune complex
Solution redissolves liquid, positive control, negative control.
In above-mentioned several kits, coating buffer be capture CIC ELISA albumen, as C1Q, CIF albumen,
Anti-C_3 antibody；Confining liquid is the bovine serum albumin(BSA) or skimmed milk power that mass concentration is 2%-4%；Eluent includes but not limited to
The Tris-HCl buffer solutions of papain；It is to include but not limited to redissolve liquid；Glue bed medium includes but not limited to agarose
Gel, polyacrylamide gel；Include but not limited to PEG solution, boric acid salt buffer for extracting solution needed for immune complex
Liquid；Substrate includes but not limited to TMB solution, ABTS solution；Sample-loading buffer includes but not limited to the Tris- containing bromophenol blue
HCl buffer solutions；Enzyme labelled antibody is the antibody marked containing enzymes such as horseradish peroxidase, alkaline phosphatases, as HRP enzyme marks are anti-
Body；Standard items include but not limited to the present invention mercury chelating type CIC ELISA, other be chelated with the immune complex of mercury；
Positive control include but not limited to mercury chelating type CIC ELISA of the present invention, other be chelated with the immune complex of mercury, the moon
Property control be dilution buffer.
Another object of the present invention is to provide a kind of method quantitatively detecting mercury chelating type CIC ELISA, with
Know that the above-mentioned mercury chelating type immune complex of content as standard items, is detected using a pair of sample of following methods：It is enzyme-linked
Immunization, enzyme linked immunological and atomic absorption spectrum combined techniques, enzyme linked immunological and inductivity coupled plasma mass spectrometry combined techniques, purification
Immune complex and atomic absorption spectrum combined techniques, purification immune complex and inductivity coupled plasma mass spectrometry combined techniques, electricity
Swimming and enzyme linked immunological or atomic absorption spectrum or inductivity coupled plasma mass spectrometry combined techniques.
In the present invention, the having of can listing of method with detection mercury chelating type immune complex is following several, but not
It is limited to following several.
Unless otherwise indicated, the laboratory operating procedures being related to are the step of this field routine to the present invention, and reagent, material are such as
It is following cited, do not enumerate in the present invention come be commonly used in the art or can be obtained by mode purchased in market：
Dilution buffer is the 0.05M carbonate buffer solutions of pH9.6, preparation method example：Take the Na of 1.5g2CO3With
2.93g NaHCO3Dissolving plus ddH2O is settled to 1000mL；
Washing buffer is the 0.15MPBS buffer solutions of pH7.4, preparation method example：Take the KH of 0.2g2PO4, 2.90g
Na2HPO4·12H2O, KCl, 0.5mLTween-20 of NaCl, 0.2g of 8.0g, dissolving plus ddH2O is settled to 1000mL；
Confining liquid is bovine serum albumin solution, preparation method example：0.1g bovine serum albumin(BSA)s are taken, washing buffer is added
Liquid dilution is settled to 100mL；
Terminate liquid is 2MH2SO4, preparation method example：Take the ddH of 178.3mL2O, to ddH2It is added dropwise along wall in O dense
H2SO4, it is stirring while adding, it is settled to 200mL；
The pH of substrate buffer solution is 5.0, Na2HPO4Molar concentration be 0.2M, the molar concentration of citric acid is 0.1M, often
The preparation method of the substrate buffer solution of 50mL is as follows：Take 1.42gNa2HPO4, 0.96g citric acids, ddH is then added2O to 50mL,
To obtain the final product；
Substrate is methyl biphenyl amine (TMB) solution, and methyl biphenyl amine (TMB) solution is by the group distribution according to following ratio
It makes：TMB：Substrate buffer solution：0.75%H2O2=0.5mL：10mL：32 μ L, wherein TMB are the methyl biphenyl amine second of 2g/L
Alcoholic solution；
The albumen that immune complex can be captured is the albumen that can be specifically bound with immune complex, including but not limited to
Such as C1Q, CIF albumen, anti-C_3 antibody；In following embodiment, can capture immune complex albumen it is specifically used be
C1qRecombinantProtein, article No. are " NOVUSH00000712-p01 "；
The substance of capture mercury is that purchase is anti-from the mouse that the bio tech ltd Ba Ao get article No. is " bar proud AP7014 "
HgmAb；Wherein, substance, the anti-mercury antibody of the present invention with mercury specific binding is the substance for capturing mercury；
The antibody that can be specifically bound with people's carrier protein is rabbit-anti people's carrier protein antibodies, is commercially available.
Dilution multiple proportions is w/v below.
Method one：ELISA method detection mercury chelating type immune complex, is as follows：
1) albumen that can capture immune complex is coated on solid phase carrier：C1Q is diluted with dilution buffer
Albumen is added to 2600-20000 times in elisa plate micropore, and 4 DEG C of 16-18 hour overnight or 37 DEG C of water-baths 1-3 hours store
Refrigerator；
2) it closes：Dilution buffer is removed, washing buffer is used in combination to be washed, is waited after the completion of washing, adds confining liquid, 37
It DEG C places 1 hour, removes confining liquid, washing buffer is used in combination to be washed, after the completion of washing, elisa plate is placed 1 small in 37 DEG C
When；
3) add sample to be tested, and incubate：It is sampled from the circulatory system, makees sample to be tested；With the mercury chelating type of known content
Immune complex makees standard items；10-40 times is diluted with dilution buffer, is added in micropore, 37 DEG C act on 1-2 hours；
4) substance of capture mercury is added, and incubates：Sample to be tested is removed, washing buffer is used in combination to be washed, it is to be washed
After the completion of washing, addition is diluted with dilution buffer to be had the substance of affinity with mercury or can react to form antigen antibody complex with mercury
Anti- mercury antibody be diluted to 31000-250000 times, 37 DEG C act on 1-2 hours, it is made to be reacted with the mercury on immune complex；
5) enzyme conjugates incubates：Anti- mercury antibody is removed, washing buffer is used in combination to be washed, is waited after the completion of washing, is added
With the diluted HRP enzyme labelled antibodies of dilution buffer, 37 DEG C act on 1-2 hours, make itself and anti-mercury antibody response；
6) substrate incubates：Enzyme labelled antibody is removed, washing buffer is used in combination to be washed, is waited after the completion of washing, substrate is added,
37 DEG C are protected from light effect 30 minutes；
7) reaction is terminated：Terminate liquid is added dropwise to each micropore；
8) it is 405nm to take wavelength, and elisa plate is placed in microplate reader to the OD for reading sample to be tested group and standard items respectively
Value draws standard curve, acquires the content of sample to be tested.
In this method, when step 8) detects, microplate reader can not be used, qualitative inspection is directly carried out by the situation that develops the color yet.
This method utilizes enzyme linked immunosorbent assay (ELISA) (ELISA) principle, can be compound by the nospecific immunity in serum
Object extracts, and weight mercury is partly chelated on the immune complex extracted, and the mercury on this partial immunity compound can be with
It is captured by the substance for having affinity with mercury or the specific antibody that the anti-mercury to form antigen antibody complex can be reacted with mercury, it
Can (antibody nonrecognition coating egg be captured by the antibody that the enzymes such as horseradish peroxidase, alkaline phosphatase mark again afterwards
In vain), the antibody in capture reads OD values under the action of color developing agent and terminate liquid under instrument, and exempts from without containing chelating mercury
Epidemic disease compound will not then be captured by the specific antibody of anti-mercury, will not be with the enzymes such as horseradish peroxidase, alkaline phosphatase
The antibody of label is captured, and mercury (negative control group result is feminine gender) is not contained in agents useful for same yet, thus when read
When OD value results are shown as the positive, you can prove the mercury for detecting to chelate on CIC ELISA.
Method two：ELISA method+AAS methods detect mercury chelating type immune complex, are as follows：
1) albumen that can capture immune complex is coated on solid phase carrier：Coating protein is diluted with dilution buffer
It to 5500-15000 times, is added in elisa plate micropore, 4 DEG C of 16-18 hour overnight or 37 DEG C of water-baths 1-3 hours, storage refrigerator；
2) it closes：Dilution buffer is removed, washing buffer is used in combination to be washed, is waited after the completion of washing, adds confining liquid, 37
It DEG C places 1 hour, removes confining liquid, washing buffer is used in combination to be washed, after the completion of washing, elisa plate is placed 1 small in 37 DEG C
When；
3) add sample to be tested, and incubate：It is sampled from the circulatory system, makees sample to be tested；With the mercury chelating type of known content
Immune complex makees standard items；10-40 times is diluted with dilution buffer, is added in micropore, 37 DEG C act on 1-2 hours；
4) it elutes：Sample to be tested is removed, washing buffer is used in combination to be washed, is waited after the completion of washing, eluent is added, in
It is acted on 1-2 hours at 37 DEG C；
5) it detects：It samples from ELISA micropores, is chelated in the mercury on immune complex in Atomic Absorption Spectrometer detection,
Read respective value.
This method further combines atomic absorption spectrum (AAS) principle on the basis of enzyme linked immunological principle, utilizes original
Sub- absorption spectrometer detection chelating is in the mercury on CIC ELISA, due to only containing immune complex in solution, and it is used
(negative control group result is feminine gender) not mercurous in reagent, will not shine at interference, thus when read result is shown result
For the positive when, you can prove the mercury for detecting to chelate on CIC ELISA.
The method of the present invention scope of application is wider, can be cost-effective, and increases the differential water pressures inside and outside bag filter, improves
It dialyses rate, can shorten manufacturing cycle, also the wastes such as ion in waste liquid are recycled, there is energy-saving and environment-friendly spy
Point avoids causing chemical contamination, therefore the present invention has the larger market competitiveness.
It although an embodiment of the present invention has been shown and described, for the ordinary skill in the art, can be with
Understanding without departing from the principles and spirit of the present invention can carry out these embodiments a variety of variations, modification, replace
And modification, the scope of the present invention is defined by the appended.
Claims (9)
1. a kind of excellent effect mercury chelating type immune complex, which is characterized in that the mercury chelating type immune complex is with next
Kind:
Mercury ion is incorporated into the compound of immune complex formation；
Or mercury ion be incorporated into after carrier protein with and the antibody that specifically binds of the carrier protein be formed by compound；
Or mercury ion be incorporated into immunoglobulin after the compound to be formed is combined with carrier protein.
2. mercury chelating type immune complex according to claim 1, it is characterised in that:The immune complex or carrier egg
At least containing one kind in zinc fingers, sulfydryl, cysteine residues, mercury ion and immune complex or carrier in white structure
Albumen is combined with zinc fingers or sulfydryl or cysteine residues.
3. mercury chelating type immune complex according to claim 2, it is characterised in that:The carrier protein is antibody egg
In vain, among lipoprotein, hemoglobin, nuclear antigen, foreign sera antigen, virus, bacterium, protozoon, worm or immunoglobulin
It is a kind of.
4. mercury chelating type immune complex according to claim 1, it is characterised in that:The mercury ion passes through chelating agent and load
Body protein is combined with the antibody of energy and carrier protein specific binding again after combining, and the chelating agent is ITCBE, EDTA, more phosphorus
One kind in hydrochlorate, amino carboxylic acid, 1,3- diketone, hydroxycarboxylic acid.
5. a kind of preparation method of mercury chelating type immune complex as described in claim 1, which is characterized in that including following step
Suddenly：
S1：First chelating agent solution is prepared and is completed, then chelating agent is dissolved in solvent, you can obtains chelating agent solution；
S2：Appropriate carrier protein is taken, carrier protein is added in borate buffer solution, you can carrier protein solution is made；
S3：Chelating agent solution obtained is added in carrier protein solution, is placed in water bath with thermostatic control and stirs evenly, constant temperature
Water-bath 4-30h, you can obtain mixed liquor；
S4：It fetches bag filter and EDTA-NaHCO is added in bag filter3Solution boils, and bag filter is fixed on to the middle part of water tank,
And enough ddH are added in water tank2O, the ddH2The volume ratio of solution is 80-85 in the volume and bag filter of O:1, repeating should
Mixing in S3 is fitted into bag filter by step 2-4 times, and bag filter is placed in constant temperature water tank, and foot is added in constant temperature water tank
The ddH of amount2O, the ddH2The volume ratio of solution is 80-85 in the volume and bag filter of O:1, it, will be in bag filter after dialysed overnight
Liquid collect.
S5：PH to 7.0 is adjusted dilute hydrochloric acid solution is added from the liquid collected in bag filter, is slowly dropped into mercury ion solution,
Do not stop to shake when instillation, instills and complete to be placed in water bath with thermostatic control and stir evenly, water bath with thermostatic control 4-30h, then repeatedly S4 mono-
It is secondary, liquid is collected, mercury chelating type antigen is obtained.
S6：The waste liquid collected in solution and S5 in S4 outside bag filter is mixed, is added and lives into mixed solution
Property charcoal adsorbed, and repeat add above-mentioned absorbent charcoal material, reuse atomic fluorescence method and processed solution quantified
Analysis, when mercury ion enrichment degree reaches 0.1mol/ml in solution, recycling mercury ion recycles.
S7：The antibody that can be specifically bound with carrier protein is added in above-mentioned mercury chelating type antigen, mercury chelating is obtained after reaction
Type immune complex.
6. mercury chelating type immune complex as described in claim 1 chelates the reagent of CIC ELISA preparing detection mercury
Or the application in enzyme linked immunological kit.
7. a kind of enzyme linked immunological kit of detection mercury chelating CIC ELISA, it is characterised in that：The kit includes such as
The standard items of mercury chelating type immune complex described in claim 1.
8. enzyme linked immunological kit as claimed in claim 6, which is characterized in that the kit further includes that at least one or less is tried
Agent：The coating buffer or the coating buffer containing the antibody for capturing mercury, enzyme labelled antibody of albumen containing capture antigen antibody complex.
9. a kind of method of qualitative detection mercury chelating type CIC ELISA, which is characterized in that wanted with the right of known content
It asks the mercury chelating type immune complex described in 1 as standard items, is detected using a pair of sample of following methods：Enzyme linked immunological
Method, enzyme linked immunological and atomic absorption spectrum combined techniques, enzyme linked immunological and inductivity coupled plasma mass spectrometry combined techniques, purification are immune
Compound and atomic absorption spectrum combined techniques, purification immune complex and inductivity coupled plasma mass spectrometry combined techniques, electrophoresis and
Enzyme linked immunological or atomic absorption spectrum or inductivity coupled plasma mass spectrometry combined techniques.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810594224.8A CN108761060A (en) | 2018-06-11 | 2018-06-11 | A kind of excellent effect mercury chelating type immune complex and preparation method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810594224.8A CN108761060A (en) | 2018-06-11 | 2018-06-11 | A kind of excellent effect mercury chelating type immune complex and preparation method thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN108761060A true CN108761060A (en) | 2018-11-06 |
Family
ID=64020849
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201810594224.8A Pending CN108761060A (en) | 2018-06-11 | 2018-06-11 | A kind of excellent effect mercury chelating type immune complex and preparation method thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN108761060A (en) |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105137059A (en) * | 2015-07-14 | 2015-12-09 | 上海拜豪生物科技有限公司 | Mercury chelating immune complex and preparation method and application thereof |
-
2018
- 2018-06-11 CN CN201810594224.8A patent/CN108761060A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105137059A (en) * | 2015-07-14 | 2015-12-09 | 上海拜豪生物科技有限公司 | Mercury chelating immune complex and preparation method and application thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN110862881B (en) | Special cleaning solution or diluent for full-automatic chemiluminescence determinator and preparation method thereof | |
| CN102426239A (en) | Colloidal gold chromatographic test strip for rapidly testing lead ion | |
| CN108593619A (en) | The detection kit of heavy metal cadmium ion and its application | |
| CN108508213A (en) | The detection kit of heavy metal lead ion and its application | |
| CN108761060A (en) | A kind of excellent effect mercury chelating type immune complex and preparation method thereof | |
| CN104965076B (en) | Cadmium chelate immune complex and preparation method and application thereof | |
| CN108828207A (en) | A kind of excellent effect cadmium chelating type immune complex and preparation method thereof | |
| CN108663528A (en) | A kind of excellent effect lead chelating type immune complex and preparation method thereof | |
| CN105137059B (en) | Mercury chelating immune complex and preparation method and application thereof | |
| CN108802362A (en) | A kind of excellent effect arsenic chelating type immune complex and preparation method thereof | |
| CN105044369B (en) | Lead-chelated immune complex and preparation method and application thereof | |
| CN101885775B (en) | Preparation and enzyme-linked immunosorbent assay method for heavy metal mercury polyclonal antibody | |
| CN102253213A (en) | Colloidal gold chromatography test strip for quickly detecting lead ions | |
| CN105044324A (en) | Arsenic-chelated immune complex and preparation method and application thereof | |
| CN109001448A (en) | A kind of novel purification arsenic chelating type immune complex method | |
| CN105021548B (en) | Arsenic-fibrinogen chelate as well as preparation method and application thereof | |
| CN105115914B (en) | Cadmium-low density lipoprotein chelate as well as preparation method and application thereof | |
| CN105044008B (en) | Cadmium-high density lipoprotein chelate as well as preparation method and application thereof | |
| CN104961824A (en) | Lead and high-density lipoprotein chelate as well as preparation method and application thereof | |
| CN105044006B (en) | Mercury-IgG chelate and preparation method and application thereof | |
| CN105017430B (en) | Nickel chelate immune complex and preparation method and application thereof | |
| CN105021554B (en) | arsenic-IgM chelate as well as preparation method and application thereof | |
| CN105044004B (en) | Mercury-IgM chelate as well as preparation method and application thereof | |
| CN105021550B (en) | Mercury-high density lipoprotein chelate and preparation method and application thereof | |
| CN105021552B (en) | Nickel-high density lipoprotein chelate as well as preparation method and application thereof |
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 | ||
| WD01 | Invention patent application deemed withdrawn after publication | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20181106 |