CN101576548A - Method for detecting Al<3+> in drinking water or cells based on functionalized gold nanoprobe colour comparison - Google Patents
Method for detecting Al<3+> in drinking water or cells based on functionalized gold nanoprobe colour comparison Download PDFInfo
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Abstract
The invention relates to a method for detecting Al<3+> in drinking water or cells based on functionalized gold nanoprobe colour comparison. The method uses functionalized gold nano-particles modified by polypeptide as a detection probe and detects Al<3+> in drinking water or cells with high sensitivity and selectivity. The detection limit of the drinking water is 0.1mumol/l and the linear range for detecting Al<3+> in cells is (1-10) mumol/l. The detection method is simple and quick, high in sensitivity and high in selectivity and can realize open hole detection.
Description
Technical field
The present invention relates to based on Al in the functionalization golden nanometer particle colorimetric detection potable water or in the cell
3+Method.
Technical background
Optical characteristics that distance that metal nanoparticle showed relies on and big characteristics such as specific surface area make it can be used as important colorimetric material and use.The molar extinction coefficient that their surface plasma resonance absorbs is more than 100 times of organic dyestuff.The optical property of utilizing their size and dimension to rely on provides sensitivity and selectivity (chemistry summary, Chem.Rev., 2005,105,1547-1562 that can be comparable with fluorescent method based on the molecular recognition method of nano particle; The chemistry summary, Chem.Rev., 2004,104,293-346).At present, the sensor based on metal nanoparticle has been applied to many aspects (JACS, J.Am.Chem.Soc., 2003,125,6642-6643 such as DNA, protein, Ag-Ab immune response and metal ion detection; The nanometer communication, Nano.Letters, 2008,8,529-533).
Development metal ion detection device is the focus of association area research always.Metallic ion all plays crucial effects in environment and biosome.Although it is the content in human body is few, closely bound up with people's existence and health.Their excess intake, deficiency or shortage all can cause the unusual of Human Physiology to some extent or disease takes place.Therefore it all is very important and necessary detecting metallic ion in environment or biosome.The metal ion detection device also from the past traditional detection method such as atomic absorption, chromatogram and induced plasma mass spectrum etc. change into need simple, fast, can detection by quantitative and the detecting device highly sensitive and that selectivity is good of monitoring in real time detect various useful or harmful metallic ions.Developed a lot of ion detectors at present, but the rarer Al that is applied to
3+Detecting device.
Aluminium has the important physical effect in vivo.On the one hand, a large amount of external (in vitro) experimental results show that aluminium can suppress the activity of the enzyme in many basic metabolism such as tricarboxylic acid cycle and the glycolysis; On the other hand, the aluminium that enters in the human body by many approach (food, medicine and potable water) can cause a series of the nervous system diseases, as (science such as senile dementia, Alzheimer's disease, parkinsonism, dialysis encephalopathy, osteomalacia and anemias, Science, 1980,208,297-299; Science, Science, 1973,180,511-513).The method of traditional detection aluminium generally is atomic absorption and emission spectrum etc., but only can detect total aluminium content.At present also there is not golden nanometer particle to detect Al in water or in the cell based on functionalization
3+Report.
Summary of the invention
In order to solve the problem that prior art exists, the purpose of this invention is to provide based on the golden nanometer particle detection potable water of functionalization or the Al in the cell
3+Method.
The present invention utilizes amino acid sequence the nm of gold surface to be modified a kind of detector probe that obtains for halfcystine-alanine-leucine-asparagine-asparagine (CALNN).CALNN and gold surface have very strong compatibility, can form fine and close protective seam, make the water-soluble solution of golden nanometer particle, and can keep long-term stability.The mercaptan key and the gold surface of CALNN peptide chain N end halfcystine can form firm covalent bond, and hydrophobic alanine and leucine can promote the self assembly of polypeptide, and two Ns holding at C make polypeptide hydrophilic.Therefore the nano Au particle of CALNN modification has good water-solubility.And the C of CALNN end has carboxyl, can and Al
3+Specific complexing can the good detection Al of highly sensitive selectivity
3+And not influenced by other metallic ions.And have negative charge by the nano Au particle of CALNN functionalization, can not have an effect with cell.Therefore, the present invention can utilize the detection Al of this system highly sensitive and high selectivity in potable water or cell
3+
The concrete technical scheme that realizes method of the present invention is as follows:
1. synthesize golden nanometer particle as the functionalization of probe:
At first, according to the Frens-Turkevich method, utilize sodium citrate reduction gold chloride to prepare particle diameter and be the 13nm gold particle: with concentration is that to join particle diameter be in the 13nm gold particle aqueous solution for the pentapeptide CALNN of 1mg/ml, make pentapeptide CALNN in above-mentioned mixed solution ultimate density be 0.1mg/ml, leave standstill reaction 1 hour under the room temperature, abandon supernatant behind the centrifugal 13000rpm; Centrifugal precipitation of getting off is added deionized water, abandon supernatant behind the centrifugal again 13000rpm; At last with centrifugal get off sediment be dissolved in the deionized water, obtain the golden nanometer particle probe solution of functionalization;
2. in deionized water, detect Al
3+
At first, with 20 μ l, the golden nanometer particle probe solution (p-GNPs) of the functionalization of 12nmol/l joins 16 metal ion species of 180 μ l: Na respectively
+, K
+, Ca
2+, Mg
2+, Ba
2+, Zn
2+, Fe
2+, Cd
2+, Cu
2+, Ni
2+, Co
2+, Mn
2+, Al
3+, Fe
3+, Pb
2+And Hg
2+Aqueous solution in, make the ultimate density of every metal ion species solution be respectively 0.1-10 μ mol/l; After 5 minutes, observe the solution colour variation and measure corresponding ultraviolet-visible absorption spectroscopy;
When every metal ion species concentration is 1.5 μ mol/l, only contain Al
3+Solution colour change: golden nanometer particle probe solution (p-GNPs) color of functionalization become lilac by redness, and contain other ion solution color no changes; Al in solution
3+When concentration increased to 6 μ mol/l from 1.5 μ mol/l, the variation of solution colour was followed successively by grey violet, purple, bluish violet and blueness; Work as Al
3+Concentration is during greater than 6 μ mol/l, and the color of solution no longer continues to change; During less than 10 μ mol/l, significant change does not all take place to other every metal ion species in solution colour in concentration;
Measure the ultraviolet-visible absorption spectroscopy of above-mentioned solution, Al in solution
3+When concentration is 1.5 μ mol/l, ultraviolet-visible absorption spectroscopy generation minor shifts: be offset to 524nm from 522nm; Al in solution
3+When concentration was 6 μ mol/l, ultraviolet-visible absorption spectroscopy was offset to 580nm; Continue to increase Al
3+Concentration, ultraviolet-visible absorption spectroscopy no longer continue to be offset; When other every metal ion species concentration was less than or equal to 10 μ mol/l, ultraviolet-visible absorption spectroscopy did not have obvious skew.Because of international standard regulation potable water Al content is up to 0.2mg/l, i.e. 7.4 μ mol/l, so this probe can only not change by solution colour by means of any instrument: by redness to blue gradual change, the Al in just can the open hole detection aqueous systems
3+
3. in potable water, detect Al
3+
Be acidified to pH 3 with drinking the HCl of water sample, make the Al in the potable water be converted into Al with 6mol/l
3+Form; NaOH with 6mol/l transfers to neutrality with pH again, and with 20 μ L, the golden nanometer particle probe solution (p-GNPs) of the functionalization of 12nmol/l joins drinking in the water sample and mixing of the above-mentioned processing of 180 μ l, after 5 minutes, observes the solution colour situation of change.If drink Al in the water sample
3+Concentration less than 1.5 μ mol/l, the color of solution does not have significant change, the redness of golden nanometer particle probe solution (p-GNPs) color of functionalization; If drink the Al in the water sample
3+Greater than 1.5 μ mol/l, then the color of solution from the redness of the golden nanometer particle probe solution of functionalization successively to lilac, purple, bluish violet, blue direction changes; Al in drinking water sample
3+Concentration is during greater than 6 μ mol/l, and the color of solution no longer continues to change, and still be blueness;
4. in cell, detect Al
3+
At first cervix cancer (Hela) cell being put in the cell culture medium (DMEM), is 5%CO in volume ratio
2With hatched 24 hours in the cell culture incubator of 95% air, incubation temperature is 37 ℃; After cell is hatched, get Al
3+Concentration be respectively each 200 μ l of above-mentioned solution of 1,5,8,10,50 and 100 μ mol/l, join respectively in the above-mentioned cell culture medium (DMEM), be 5%CO in volume ratio
2In the cell culture incubator of 95% air, cultivated altogether 2 hours under 37 ℃, be 7.2 with pH more afterwards, the buffer solution that composition is made up of the NaCl of the 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES) of 20mmol/l and 0.15mol/l is given a baby a bath on the third day after its birth time; With 200 μ l, the golden nanometer particle probe solution (p-GNPs) of the functionalization of 2.4nmol/l is 5%CO in volume ratio again
2In the cell culture incubator of 95% air, cultivated altogether 45 minutes under 37 ℃, after giving a baby a bath on the third day after its birth time with above-mentioned buffer solution, observe at optics light field microscopically; The Al of variable concentrations
3+Cause golden nanometer particle in various degree gathering and make cell present from shallow to deep redness: Al
3+When concentration was 1 μ mol/l, cell only can be seen a spot of very shallow redness; Al
3+When concentration was 5 μ mol/l, the redness on the cell deepened; Al
3+When concentration was 10 μ mol/l, the redness on the cell was quite obvious; Al
3+When concentration increases to 100 μ mol/l, still be bright redness on the cell; No Al
3+When cultivating altogether, do not observe golden nanometer particle on the cell and assemble, color is colourless.Therefore, the golden nanometer particle probe solution of functionalization can be used for detecting the Al in the cell
3+Content.
5. the selectivity experiment that detects in the cell
Experimental technique is identical with 4.At first cervix cancer (Hela) cell being put in the cell culture medium (DMEM), is 5%CO in volume ratio
2With hatched 24 hours in the cell culture incubator of 95% air, incubation temperature is 37 ℃; After cell is hatched, with K
+, Mg
2+, Ca
2+, Ba
2+, Zn
2+, Cu
2+, Fe
2+, Co
2+, Ni
2+, Mn
2+, Cd
2+, Fe
3+, Pb
2+And Hg
2+Join in the cell culture fluid (DMEM) Deng 15 metal ion species storing solutions, be mixed with the metal ion solution that concentration is 10 μ mol/l; Get 200 μ l respectively, the various metal ion solutions of 10 μ mol/l join in the above-mentioned cell culture medium (DMEM), are 5%CO in volume ratio
2In the cell culture incubator of 95% air, cultivated altogether 2 hours under 37 ℃, be 7.2 with pH more afterwards, the buffer solution that composition is made up of the NaCl of the HEPES of 20mmol/l and 0.15mol/l is given a baby a bath on the third day after its birth time; With 200 μ l, the golden nanometer particle probe solution (p-GNPs) of the functionalization of 2.4nmol/l is 5%CO in volume ratio again
2In the cell culture incubator of 95% air, cultivated altogether 45 minutes under 37 ℃, after giving a baby a bath on the third day after its birth time with above-mentioned buffer solution, observe at optics light field microscopically; Under the situation that above-mentioned 15 kinds of ions exist, do not observe the gathering of nano Au particle on the cell, color is colourless.Therefore, the golden nanometer particle probe solution of functionalization can not detect and remove Al in the cell
3+Other metallic ion in addition is so have good selectivity.
Beneficial effect: the peptide C ALNN of (1) application function of the present invention carries out finishing to nm of gold.Polypeptide plays two kinds of effects: a kind of is carboxyl and the Al that utilizes the C end
3+Optionally identification and complexing realize the detection of highly sensitive high selectivity; Another kind is as stabilizing agent, can stablize the parcel nano Au particle, also can make system have negative charge and not with cytosis.
Utilize peptide C ALNN to stablize golden nanometer particle, the halfcystine of CALNN peptide chain N end can form firm covalent bond with mercaptan key and gold surface, and alanine and leucine can promote the self assembly of polypeptide, and two Ns holding at C make polypeptide hydrophilic.Polypeptide and nm of gold reaction are at room temperature carried out, and mild condition need not the auxiliary of specific installation.This synthetic method system is stable, and is simple to operate, and required time is short.
(2) utilize the nano Au particle of functionalization in cell, to detect Al first
3+All the time, Al
3+The important physical effect is arranged in vivo.On the one hand, a large amount of external (in vitro) experimental results show that aluminium can suppress the activity of the enzyme in many basic metabolism such as tricarboxylic acid cycle and the glycolysis; On the other hand, the aluminium that enters in the human body by many approach (food, medicine and potable water) can cause a series of the nervous system diseases, as senile dementia, Alzheimer's disease, parkinsonism, dialysis encephalopathy, osteomalacia and anemia etc.Therefore, no matter be in potable water or in cell, to detect Al
3+, all have very important significance.
(3) first the nano Au particle of functionalization is applied to metal ion detection in the cell.
Although the metal ion detection device has a lot, can be applied to the also few of living things system detection.Because this needs the detecting device good water solubility, and is highly sensitive, not with the living things system effect of complexity, or the like.At present, being applied to detect in the cell metallic ion only has the several method of reporting in the document, and all is based on the fluoroscopic examination principle.Also there is not the nano Au particle of bibliographical information application functionization in cell, to detect the method for metallic ion.The present invention is applied to nano Au particle detect in the cell Al first
3+, and obtained good result.
(4) detect simple, intuitive.The detection means of the cell detection of comparing---electron microscope, fluorescent microscope etc., this testing result can be by common light field observation by light microscope.Sample does not need other pre-service, and detection time is short, and each hour can be detected up to a hundred samples.The colour rendering of golden nanometer particle is stable, can quenching property be preserved by test sample for a long time.The detection method simple, intuitive.
Description of drawings
Fig. 1 (a) is the ultraviolet-visible absorption spectroscopy of the 13nm gold particle of CALNN coating; (b) be to add 10 μ mol/l Al
3+Ultraviolet-visible absorption spectroscopy.Wherein, do not add Al
3+The time golden nanometer particle be discrete state, the ultraviolet absorption maximum is λ=522nm ((a)); Add Al
3+Back golden nanometer particle is state of aggregation, and the ultraviolet absorption maximum is λ ≈ 580nm ((b)).
Fig. 2 absorbs ratio (ε with the mole delustring
580/ ε
522) expression in water, detect Al
3+Range of linearity ultraviolet spectrogram.
Fig. 3 is with molar extinction coefficient ratio (ε
580/ ε
522) expression in water, detect Al
3+Optionally ultraviolet spectrogram.Wherein: 1, the golden nanometer particle probe solution; 2, Na
+3, K
+4, Mg
2+5, Ca
2+6, Ba
2+7, Zn
2+8, Mn
2+9, Cu
2+10, Co
2+11, Ni
2+12, Cd
2+13, Fe
2+14, Al
3+15, Fe
3+16, Hg
2+17, Pb
2+Various ion concentrations are 10 μ mol/l among the figure.
Fig. 4 (a) is the ultraviolet-visible absorption spectroscopy of the golden nanometer particle of CALNN coating; (b) detect Al in the potable water
3+Ultraviolet-visible absorption spectroscopy, record Al in the potable water
3+Concentration is greater than 6 μ mol/l.
Fig. 5 is the Al that detects variable concentrations in cell
3+Optics light field microphotograph.(a) 5 μ mol/l Al
3+(b) 10 μ mol/l Al
3+(c) no Al
3+Wherein the cell color has less redness in (a); (b) cell presents vivid redness in; (c) cell is colourless in.
Fig. 6 is the Al with elder generation and variable concentrations
3+Cultivate altogether again and be total to the postdigestive ultraviolet-visible absorption spectroscopy of cultured cells with the golden nanometer particle probe solution.(a)1μmol/l?Al
3+；(b)5μmol/l?Al
3+；(c)8μmol/l?Al
3+；(d)10μmol/l?Al
3+；(e)50μmol/l?Al
3+；(f)100μmol/l?Al
3+。The peak position of the absorbance of surveying is changed to λ=530nm in the illustration.
Fig. 7 is when concentration is 10 μ mol/l, detects to comprise Al in cell
3+Optics light field microphotograph at 15 interior metal ion species.
Embodiment
Embodiment 1 provides a kind of and has detected Al based on the functionalized nano particle in aqueous systems
3+Method.At first, according to the Frens-Turkevich method, utilize sodium citrate reduction gold chloride to prepare particle diameter and be the 13nm gold particle: with concentration is that to join particle diameter be in the 13nm gold particle aqueous solution for the pentapeptide CALNN of 1mg/ml, make pentapeptide CALNN in above-mentioned mixed solution ultimate density be 0.1mg/ml, leave standstill reaction 1 hour under the room temperature, abandon supernatant behind the centrifugal 13000rpm; Centrifugal precipitation of getting off is added deionized water, abandon supernatant behind the centrifugal again 13000rpm; At last with centrifugal get off sediment be dissolved in the deionized water, obtain the golden nanometer particle probe solution of functionalization;
With 20 μ l, the golden nanometer particle probe solution of the functionalization of 12nmol/l joins 16 metal ion species of 180 μ l variable concentrations: Na respectively
+, K
+, Ca
2+, Mg
2+, Ba
2+, Zn
2+, Fe
2+, Cd
2+, Cu
2+, Ni
2+, Co
2+, Mn
2+, Al
3+, Fe
3+, Pb
2+And Hg
2+Aqueous solution in, make the ultimate density of various metal ion solutions be respectively 0.1-10 μ mol/l; After 5 minutes, observe the solution colour variation and measure corresponding ultraviolet-visible absorption spectroscopy.
When concentration of metal ions is 1.5 μ mol/l, only contain Al
3+Solution colour change: become lilac from the redness of the golden nanometer particle probe solution of functionalization, and contain other ion solution no changes; Al in solution
3+When concentration increased to 6 μ mol/l from 1.5umol/l, the variation of solution colour was followed successively by grey violet, purple, bluish violet and blueness; Work as Al
3+Concentration is during greater than 6 μ mol/l, and the color of solution no longer changes; During less than 10 μ mol/l, significant change does not all take place to other ion in solution colour in concentration;
Measure the ultraviolet-visible absorption spectroscopy of above-mentioned solution, Al in solution
3+When concentration is 1.5 μ mol/l, ultraviolet-visible absorption spectroscopy generation minor shifts: be offset to 524nm from 522nm; Al in solution
3+When concentration was 6 μ mol/l, ultraviolet-visible absorption spectroscopy was offset to 580nm; Continue to increase Al
3+Concentration, ultraviolet-visible absorption spectroscopy no longer continue to be offset; When other concentration of metal ions was less than or equal to 10 μ mol/l, ultraviolet-visible absorption spectroscopy did not have obvious skew.
The synthetic method of embodiment 2p-GNPs is identical with embodiment 1.At first, according to the Frens-Turkevich method, utilize sodium citrate reduction gold chloride to prepare particle diameter and be the 13nm gold particle: with concentration is that to join particle diameter be in the 13nm gold particle aqueous solution for the pentapeptide CALNN of 1mg/ml, make pentapeptide CALNN in above-mentioned mixed solution ultimate density be 0.1mg/ml, leave standstill reaction 1 hour under the room temperature, abandon supernatant behind the centrifugal 13000rpm; Centrifugal precipitation of getting off is added deionized water, abandon supernatant behind the centrifugal again 13000rpm; At last with centrifugal get off sediment be dissolved in the deionized water, obtain the golden nanometer particle probe solution of functionalization;
Certain potable water sample is acidified to pH 3 with the HCl of 6mol/l, makes the Al in the potable water be converted into Al
3+Form; NaOH with 6mol/l transfers to neutrality with pH again.With 20 μ l, the golden nanometer particle probe of 12nmol/l joins drinking in the water sample and mixing of the above-mentioned processing of 180 μ l.After 5 minutes, the color that bore hole promptly can be observed solution becomes blueness by the redness of golden nanometer particle probe solution.
At first, according to the Frens-Turkevich method, utilize sodium citrate reduction gold chloride to prepare particle diameter and be the 13nm gold particle: with concentration is that to join particle diameter be in the 13nm gold particle aqueous solution for the pentapeptide CALNN of 1mg/ml, make pentapeptide CALNN in above-mentioned mixed solution ultimate density be 0.1mg/ml, leave standstill reaction 1 hour under the room temperature, abandon supernatant behind the centrifugal 13000rpm; Centrifugal precipitation of getting off is added deionized water, abandon supernatant behind the centrifugal again 13000rpm; At last with centrifugal get off sediment be dissolved in the deionized water, obtain the golden nanometer particle probe solution of functionalization;
At first cervix cancer (Hela) cell being put in the cell culture medium (DMEM), is 5%CO in volume ratio
2With hatched 24 hours in the cell culture incubator of 95% air, incubation temperature is 37 ℃; After cell is hatched, get Al
3+Concentration be respectively each 200 μ l of above-mentioned solution of 1,5,8,10,50 and 100 μ mol/l, join respectively in the above-mentioned cell culture medium (DMEM), be 5%CO in volume ratio
2In the cell culture incubator of 95% air, cultivated altogether 2 hours under 37 ℃, be 7.2 with pH more afterwards, the buffer solution that composition is made up of the NaCl of the 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES) of 20mmol/l and 0.15mol/l is given a baby a bath on the third day after its birth time; With 200 μ l, the golden nanometer particle probe solution (p-GNPs) of the functionalization of 2.4nmol/l is 5%CO in volume ratio again
2In the cell culture incubator of 95% air, cultivated altogether 45 minutes under 37 ℃, after giving a baby a bath on the third day after its birth time with above-mentioned buffer solution, observe at optics light field microscopically; The Al of variable concentrations
3+Cause golden nanometer particle in various degree gathering and make cell present from shallow to deep redness: Al
3+When concentration was 1 μ mol/l, cell only can be seen a spot of very shallow redness; Al
3+When concentration was 5 μ mol/l, the redness on the cell deepened; Al
3+When concentration was 10 μ mol/l, the redness on the cell was quite obvious; Al
3+When concentration increases to 100 μ mol/l, still be bright redness on the cell; No Al
3+When cultivating altogether, do not observe golden nanometer particle on the cell and assemble, color is colourless.Therefore, the golden nanometer particle probe solution of functionalization can be used for detecting the Al in the cell
3+Content.
Embodiment 4
Embodiment 4 difference from Example 3 are that the concentration difference of golden nanometer particle probe solution of checking functionization is to detecting the influence of effect.
The synthetic method of nano Au particle probe is identical with embodiment 1.At first, according to the Frens-Turkevich method, utilize sodium citrate reduction gold chloride to prepare particle diameter and be the 13nm gold particle: with concentration is that to join particle diameter be in the 13nm gold particle aqueous solution for the pentapeptide CALNN of 1mg/ml, make pentapeptide CALNN in above-mentioned mixed solution ultimate density be 0.1mg/ml, leave standstill reaction 1 hour under the room temperature, abandon supernatant behind the centrifugal 13000rpm; Centrifugal precipitation of getting off is added deionized water, abandon supernatant behind the centrifugal again 13000rpm; At last with centrifugal get off sediment be dissolved in the deionized water, obtain the golden nanometer particle probe solution (p-GNPs) of functionalization;
At first cervix cancer (Hela) cell being put in the cell culture medium (DMEM), is 5%CO in volume ratio
2With hatched 24 hours in the cell culture incubator of 95% air, incubation temperature is 37 ℃; After cell is hatched, get Al
3+Concentration be respectively each 200 μ l of above-mentioned solution of 1,5,8,10,50 and 100 μ mol/l, join respectively in the above-mentioned cell culture medium (DMEM), be 5%CO in volume ratio
2In the cell culture incubator of 95% air, cultivated altogether 2 hours under 37 ℃, be 7.2 with pH more afterwards, the buffer solution that composition is made up of the NaCl of the 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES) of 20mmol/l and 0.15mol/l is given a baby a bath on the third day after its birth time; With concentration be respectively afterwards 1.2,3.6 or each 200 μ L of the golden nanometer particle probe solution of 4.8nmol/l join in the above-mentioned cell, be 5%CO in volume ratio
2In the cell culture incubator of 95% air, cultivate 45 minutes altogether under 37 ℃ after, give a baby a bath on the third day after its birth time with above-mentioned buffer solution again.By the microscopic examination of optics light field, although the concentration difference of detector probe p-GNPs, experimental result is identical with the experimental result of embodiment 3: the Al of variable concentrations
3+Cause golden nanometer particle in various degree gathering and make cell present from shallow to deep redness: the Al that cultivates altogether
3+When concentration was 1 μ mol/l, cell only can be seen a spot of very shallow redness; Be total to the Al that cultivates
3+When concentration was 5 μ mol/l, the redness on the cell deepened; As the Al that cultivates altogether
3+When concentration was 10 μ mol/l, the redness on the cell was quite obvious; As the Al that cultivates altogether
3+When concentration increases to 100 μ mol/l, still be bright redness on the cell; No Al
3+When cultivating altogether, do not observe golden nanometer particle on the cell and assemble, color is colourless.Therefore, the concentration of the golden nanometer particle probe solution of functionalization does not have influence to testing result.
Claims (1)
1, the gold nanoprobe colour comparison based on functionalization detects Al in potable water or the cell
3+Method, it is characterized in that step and condition are as follows:
1) golden nanometer particle of synthetic functionalization as probe:
At first, according to the Frens-Turkevich method, utilize sodium citrate reduction gold chloride to prepare particle diameter and be the 13nm gold particle: with concentration is that to join particle diameter be in the 13nm gold particle aqueous solution for the pentapeptide CALNN of 1mg/ml, make pentapeptide CALNN in above-mentioned mixed solution ultimate density be 0.1mg/ml, leave standstill reaction 1 hour under the room temperature, abandon supernatant behind the centrifugal 13000rpm; Centrifugal precipitation of getting off is added deionized water, abandon supernatant behind the centrifugal again 13000rpm; At last with centrifugal get off sediment be dissolved in the deionized water, obtain the golden nanometer particle probe solution of functionalization;
2) in deionized water, detect Al
3+
At first, with 20 μ l, the golden nanometer particle probe solution of the functionalization of 12nmol/l joins 16 metal ion species of 180 μ l: Na respectively
+, K
+, Ca
2+, Mg
2+, Ba
2+, Zn
2+, Fe
2+, Cd
2+, Cu
2+, Ni
2+, Co
2+, Mn
2+, Al
3+, Fe
3+, Pb
2+And Hg
2+Aqueous solution in, make the ultimate density of every metal ion species solution be respectively 0.1-10 μ mol/l; After 5 minutes, observe solution colour and change;
When every metal ion species concentration is 1.5 μ mol/l, only contain Al
3+Solution colour change: the golden nanometer particle probe solution color of functionalization become lilac by redness, and contain other ion solution color no changes; Al in solution
3+When concentration increased to 6 μ mol/l from 1.5 μ mol/l, the variation of solution colour was followed successively by grey violet, purple, bluish violet and blueness; Work as Al
3+Concentration is during greater than 6 μ mol/l, and the color of solution no longer continues to change; During less than 10 μ mol/l, significant change does not all take place to other every metal ion species in solution colour in concentration;
3) in potable water, detect Al
3+
Be acidified to pH 3 with drinking the HCl of water sample, make the Al in the potable water be converted into Al with 6mol/l
3+Form; NaOH with 6mol/l transfers to neutrality with pH again, and with 20 μ L, the golden nanometer particle probe solution of the functionalization of 12nmol/l joins drinking in the water sample and mixing of the above-mentioned processing of 180 μ l, after 5 minutes, observes the solution colour situation of change; If drink Al in the water sample
3+Concentration less than 1.5 μ mol/l, the color of solution does not have significant change, the redness of the golden nanometer particle probe solution color of functionalization; If drink the Al in the water sample
3+Greater than 1.5 μ mol/l, then the color of solution from the redness of the golden nanometer particle probe solution of functionalization successively to lilac, purple, bluish violet, blue direction changes; Al in drinking water sample
3+Concentration is during greater than 6 μ mol/l, and the color of solution no longer continues to change, and still be blueness;
4) in cell, detect Al
3+
At first cervix cancer Hela cell being put in the cell culture medium, is 5%CO in volume ratio
2With hatched 24 hours in the cell culture incubator of 95% air, incubation temperature is 37 ℃; After cell is hatched, get Al
3+Concentration be respectively each 200 μ l of above-mentioned solution of 1,5,8,10,50 and 100 μ mol/l, join respectively in the above-mentioned cell culture medium (DMEM), be 5%CO in volume ratio
2In the cell culture incubator of 95% air, cultivated altogether 2 hours under 37 ℃, be 7.2 with pH more afterwards, the buffer solution that composition is made up of the NaCl of the 4-hydroxyethyl piperazine ethanesulfonic acid of 20mmol/l and 0.15mol/l is given a baby a bath on the third day after its birth time; With 200 μ l, the golden nanometer particle probe solution of the functionalization of 2.4nmol/l is 5%CO in volume ratio again
2In the cell culture incubator of 95% air, cultivated altogether 45 minutes under 37 ℃, after giving a baby a bath on the third day after its birth time with above-mentioned buffer solution, observe at optics light field microscopically; The Al of variable concentrations
3+Cause golden nanometer particle in various degree gathering and make cell present from shallow to deep redness: Al
3+When concentration was 1 μ mol/l, cell only can be seen a spot of very shallow redness; Al
3+When concentration was 5 μ mol/l, the redness on the cell deepened; Al
3+When concentration was 10 μ mol/l, the redness on the cell was quite obvious; Al
3+When concentration increases to 100 μ mol/l, still be bright redness on the cell; No Al
3+When cultivating altogether, do not observe golden nanometer particle on the cell and assemble, color is colourless.
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