CN103290098B - A kind of method detecting lithium ion and test kit - Google Patents
A kind of method detecting lithium ion and test kit Download PDFInfo
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- CN103290098B CN103290098B CN201310229489.5A CN201310229489A CN103290098B CN 103290098 B CN103290098 B CN 103290098B CN 201310229489 A CN201310229489 A CN 201310229489A CN 103290098 B CN103290098 B CN 103290098B
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Abstract
The present invention relates to a kind of method detecting lithium ion and test kit.Specifically, the invention discloses a kind of method detecting lithium ion, described method is based on the catalytic action to 3,5 adenosine diphosphate (ADP)s of 3, the 5 nucleoside diphosphate acid enzymes and utilizes the anti-correlation of its catalytic efficiency and amount of lithium ions to measure the amount of lithium ion in sample.Described method specificity is high, good stability, measures accurately, and conventional method has good dependency, fully meets clinical needs, can promote the use of.
Description
Technical field
The invention belongs to field of biological detection.Specifically, the invention provides a kind of method detecting lithium concentration and test kit.
Background technology
Within 1949, Australian psychiatrist Cade have unexpectedly discovered that the effect of lithium carbonate treatment bipolar affective disorder, creates the miracle in psychiatric treatment history.Lithium salts was approved as treatment manicdepressive disease drug in 1970 by FDA.Lithium carbonate is often used to treatment bipolar disorder (bipolardisorder), it is by stablizing calcium and serum usually stabilizing the emotions, because of lithium salts discord protein bound, remove by way of single (passing through renal excretion), blood lithium concentration scope little (being about 0.4～1.5mmol/L), there is untoward reaction when blood lithium concentration is 2.0mmol/L or higher in nearly all case.Having bibliographical information lithium salts to treat minimum poisoning concentration is 1.6mmol/L, and therefore patient needs to carry out therapeutic drug monitoring in the process using this medicine.
At present, what hospital's use was more is flame spectrometry and the measurement of ion selective electrode measurement method, the former cost is low, but error is bigger, it is impossible to promptly and accurately detect result, the latter be at present in hospital use more method, but the out of order probability of electrolyte analyser used is higher, and particularly the maintenance to suction needle, adds testing cost, and repeatability is poor, in addition it is also necessary to buy this analyser specially and train relevant operator.
Therefore, at present research and development a kind of easy and simple to handle, cost is low, the method detecting serum lithium concentration that repeated, is significant.
Summary of the invention
The method that it is an object of the invention to provide low, the reproducible detection lithium ion of a kind of easy and simple to handle, quick, cost.
It is a further object of the present invention to provide a kind of test kit for above-mentioned detection method.
In a first aspect of the present invention, it is provided that a kind of measure the method for lithium ion in sample, including step:
(1) providing a liquid mixed system, described system contains sample to be tested, 3,5-adenosine diphosphate (ADP)s and 3,5-nucleoside diphosphate acid enzymes；
(2) under the catalysis of 3,5-nucleoside diphosphate acid enzymes, described mixed system carries out reaction as follows:
3,5-adenosine diphosphate (ADP) → AMP
(3) measure in described system AMP presence or absence and/or quantity and/or production rate, and convert and draw the measurement result of lithium ion in sample.
In another preference, described sample to be tested is liquid；It is preferably containing water sample；It is more preferably serum or blood sample.
In another preference, described sample to be tested without or be substantially free of AMP.
In another preference, described sample to be tested is treated or undressed sample.
In another preference, in step (3), the further catalytic reaction of described AMP obtains detectable colored substance plastoquinone.
In another preference, step (3) further comprises the steps of: compares the presence or absence of AMP and/or production rate and/or quantity and standard value or standard curve, thus drawing the measurement result of lithium ion in sample.
In another preference, in step (3), the method measuring AMP quantity and/or production rate, including step:
(3-1) 5-nucleotidase, ADA Adenosine deaminase, polynucleotide phosphorylase, xanthine oxidase exist under, by the reaction system of step (2) or the AMP in system is carried out as follows reaction:
AMP → hypoxanthine → hydrogen peroxide
(3-2) production rate of hydrogen peroxide and/or quantity in the reaction system of measuring process (3-1), and draw the measurement result of AMP.
In another preference, in step (3-2), the method measuring hydrogen peroxide quantity includes step:
Peroxidase, 4-AA, 4-chlorophenol existence under, hydrogen peroxide in the reaction system of step (3-2) is carried out chromogenic reaction, the absorbance of assaying reaction system or absorbance rate of change, thus drawing production rate and/or the quantity of hydrogen peroxide.
In another preference, in step (1), possibly together with 5-nucleotidase, ADA Adenosine deaminase, polynucleotide phosphorylase, xanthine oxidase, peroxidase, 4-AA, 4-chlorophenol in described mixed system；And step (2) and step (3) are replaced by following steps (ii) and step (iii):
(ii) 3,5-nucleoside diphosphate acid enzymes, 5-nucleotidase, ADA Adenosine deaminase, polynucleotide phosphorylase, xanthine oxidase catalysis under, carry out in described mixed system as follows reaction:
3,5-adenosine diphosphate (ADP) → AMP → hypoxanthine → hydrogen peroxide；With
Peroxidase, 4-AA, 4-chlorophenol existence under, hydrogen peroxide carries out chromogenic reaction；With
(iii) measure absorbance or the absorbance rate of change of described system, and draw the measurement result of lithium ion in sample.
In another preference, step (iii) further comprises the steps of: compares absorbance or absorbance rate of change and the 3rd standard value or the 3rd standard curve, thus drawing the measurement result of lithium ion in sample.
In another preference, 3,5-described adenosine diphosphate (ADP) concentration are 20～60mmol/L.
In another preference, 3,5-described nucleoside diphosphate acid enzyme concentrations are 1～6ku/L.
In another preference, described 5-nucleotidase concentration is 1～3ku/L.
In another preference, described ADA Adenosine deaminase concentration is 1～3ku/L.
In another preference, described polynucleotide phosphorylase concentration is 1～3ku/L
In another preference, described xanthine oxidase concentration is 1～3ku/L.
In another preference, described peroxidase concn is 0.5～1.5ku/L.
In another preference, described 4-AA concentration is 0.1～0.5g/L.
In another preference, described 4-chlorophenol concentration is 0.1～0.8g/L.
In another preference, described mixed system is possibly together with buffer, surfactant, stabilizer, activator, chelating agen, protective agent.
In another preference, described buffer is pH is the buffer of 5.5～7.5.
In another preference, described buffer is two (2-ethoxy) imino group Pehanorm buffer.
In another preference, described buffer be 50～100mmol/L, pH be 6.50 two (2-ethoxy) imino group Pehanorm buffer.
In another preference, described surfactant is nonionic surfactant.
In another preference, described surfactant includes: alkyl polyglycoside type, polyoxyethylene-type, polyol type, alkylol amide type, block polyether type.
In another preference, described surfactant concentration is 0.1 ‰～1 ‰ (percentage by weights).
In another preference, described stabilizer includes bovine serum albumin, β-dredge base ethanol, Cys or its combination；And/or
Described activator includes anhydrous calcium chloride, Magnesium sulfate heptahydrate, potassium ferrocyanide, zinc sulfate or its combination；And/or
Described chelating agen includes EDTA and sodium salt, polymethylacrylic acid, organic multicomponent phosphonic acids, hydroxyethylethylene diamine tri-acetic acid (HEDTA), bicine N-or its combination；And/or
Described protective agent includes sucrose, glucose, fucose, maltose, lactose or its combination.
In another preference, described stabilizer concentration is 0.1～0.5g/L.
In another preference, described activator concentration is 1～3mmol/L.
In another preference, described chelating agent concentrations is 0.1～0.2g/L.
In another preference, described protective agent concentration is 1～5g/L.
In second aspect present invention, it is provided that a kind of test kit, described test kit includes: the description of 3,5-nucleoside diphosphate acid enzymes, 3,5 adenosine diphosphate (ADP)s and description using method.
In another preference, 3,5-described nucleoside diphosphate acid enzymes or 3,5 adenosine diphosphate (ADP)s are arranged in same container or lay respectively at different containers.
In another preference, described test kit also includes: 5-nucleotidase, ADA Adenosine deaminase, polynucleotide phosphorylase, xanthine oxidase.
In another preference, described test kit also includes: peroxidase, 4-AA, 4-chlorophenol.
In third aspect present invention, it is provided that the purposes of a kind of test kit as described in respect of the second aspect of the invention, it is used for detecting lithium ion in sample.
Should be understood that within the scope of the present invention, above-mentioned each technical characteristic of the present invention and can combining mutually between specifically described each technical characteristic in below (eg embodiment), thus constituting new or preferred technical scheme.As space is limited, tired no longer one by one state at this.
Accompanying drawing explanation
Fig. 1 is the equation of linear regression schematic diagram of theoretical concentration and test concentrations.
Fig. 2 shows the method for the present invention and the dependency of ISE method test sera sample results.
Fig. 3 is the standard curve that embodiment 1 records.
Fig. 4 is lithium concentration is the response curve of the sample of 1.55mmol/L.
Detailed description of the invention
The present inventor is through extensive and deep research, it has unexpectedly been found that lithium ion is very sensitive to 3,5-nucleoside diphosphate acid enzymes, and in sample to be tested, the concentration of lithium ion or content and 3,5-nucleoside diphosphate acid enzymes are linear to the catalysis activity of 3,5-diphosphonic acid.Present invention desmoenzyme catalytic reaction first and colorimetry are for detecting quantity or the presence or absence of lithium ion in sample, and the method is easy, quick and reproducible.On this basis, inventor completes the present invention.
In detection method, 3,5 nucleoside diphosphate acid enzymes the reaction of catalysis 3,5 adenosine diphosphate (ADP) can generate AMP.Concrete course of reaction is as follows: under the catalytic action of 3,5 nucleoside diphosphate acid enzymes, and 3,5 adenosine diphosphate (ADP)s generate AMP；Under the catalytic action of 5-nucleotidase and ADA Adenosine deaminase and polynucleotide phosphorylase, AMP generates hypoxanthine；Under the effect of xanthine oxidase, hypoxanthine generates uric acid and hydrogen peroxide；Hydrogen peroxide and 4-AA under the effect of peroxidase, 4-chlorophenol generation chromogenic reaction, there is maximum absorption band at 546nm place.
Further, 3,5 nucleoside diphosphate acid enzymes are very sensitive to lithium ion, IC50Being about 0.1mmol/L, therefore, lithium ion can reduce the catalytic efficiency of 3,5 nucleoside diphosphate acid enzymes, and namely the catalytic efficiency of 3,5 nucleoside diphosphate acid enzymes and lithium concentration are inverse correlation.
The invention provides a kind of method one preferably detecting lithium ion, described method one includes step:
(1) providing a liquid mixed system, described system contains sample to be tested, 3,5-adenosine diphosphate (ADP)s and 3,5-nucleoside diphosphate acid enzymes；
(2) under the catalysis of 3,5-nucleoside diphosphate acid enzymes, described mixed system carries out reaction as follows:
3,5-adenosine diphosphate (ADP) → AMP
(3) measure in described system AMP presence or absence and/or quantity and/or production rate, and convert and draw the measurement result of lithium ion in sample.
In step (3), AMP quantity and/or production rate can measure according to known method, it would however also be possible to employ following method:
Method 1. includes step:
The quantity of AMP and/or production rate and the first standard value or the first standard curve are compared, thus drawing the measurement result of lithium ion in sample.
The method for making of described first standard value or the first standard curve includes step:
A () provides multiple liquid mixed system, described system contains master sample, 3,5-adenosine diphosphate (ADP)s and 3,5 nucleoside diphosphate acid enzymes, is the sample containing normal concentration lithium ion in described master sample；
B (), under the catalysis of 3,5-nucleoside diphosphate acid enzymes, carries out reaction as follows in described mixed system:
3,5-adenosine diphosphate (ADP) → AMP
C () measures production rate and/or the quantity of AMP in every individual system, thus obtaining the first standard value or the first standard curve.
Method 2. includes step:
(3-1) 5-nucleotidase, ADA Adenosine deaminase, polynucleotide phosphorylase, xanthine oxidase exist under, by the reaction system of step (2) or the AMP in system is carried out as follows reaction:
AMP → hypoxanthine → hydrogen peroxide
(3-2) production rate of hydrogen peroxide and/or quantity in the reaction system of measuring process (3-1), and draw the measurement result of AMP.
Wherein, in step (3-2), can measuring according to known method of hydrogen peroxide quantity, can also adopt with the following method, including step: peroxidase, 4-AA, 4-chlorophenol existence under, hydrogen peroxide in the reaction system of step (3-2) is carried out chromogenic reaction, generate coloured quinone, this quinones substance has absworption peak at 546nm place, by the absorbance A 1 of biochemistry analyzer METHOD FOR CONTINUOUS DETERMINATION reaction system or absorbance rate of change Δ A1, thus drawing the quantity of coloured quinone, thus drawing quantity and/or the production rate of peroxide.
It is highly preferred that further comprise the steps of:, absorbance A 1 or absorbance rate of change Δ A1 and the second standard value or the second standard curve are compared, thus drawing the quantity of hydrogen peroxide.
The method for making of described second standard value and the second standard curve includes step: peroxidase, 4-AA, 4-chlorophenol existence under, the hydrogen peroxide of multiple normal concentrations is carried out chromogenic reaction, measure absorbance A corresponding to each normal concentration hydrogen peroxide or absorbance rate of change Δ A, thus obtaining the second standard value or the second standard curve.
The invention provides the another kind of method two preferably detecting lithium ion, described method two includes step:
I () provides a liquid mixed system, described system contains sample to be tested, 3,5-adenosine diphosphate (ADP)s and 3,5-nucleoside diphosphate acid enzymes；Possibly together with 5-nucleotidase, ADA Adenosine deaminase, polynucleotide phosphorylase, xanthine oxidase, peroxidase, 4-AA, 4-chlorophenol in described mixed system；
(ii) 3,5-nucleoside diphosphate acid enzymes, 5-nucleotidase, ADA Adenosine deaminase, polynucleotide phosphorylase, xanthine oxidase catalysis under, carry out in described mixed system as follows reaction:
3,5-adenosine diphosphate (ADP) → AMP → hypoxanthine → hydrogen peroxide；
And peroxidase, 4-AA, 4-chlorophenol existence under, hydrogen peroxide carries out chromogenic reaction；With
(iii) measure absorbance A 2 or the absorbance rate of change Δ A2 of described system, and draw the measurement result of lithium ion in sample.
In step (iii), further comprise the steps of: and absorbance A 2 or absorbance rate of change Δ A2 and the three standard value or the 3rd standard curve are compared, thus drawing the measurement result of lithium ion in sample (concentration).
The method for making of described 3rd standard value and the 3rd standard curve includes step:
(i-1) multiple liquid mixed system is provided, described system contain master sample, 3,5 adenosine diphosphate (ADP)s and 3,5 nucleoside diphosphate acid enzymes, 5-nucleotidase, ADA Adenosine deaminase, polynucleotide phosphorylase, xanthine oxidase, peroxidase, 4-AA, 4-chlorophenol, be the sample containing normal concentration lithium ion in described master sample；
(ii-1) 3,5-nucleoside diphosphate acid enzymes, 5-nucleotidase, ADA Adenosine deaminase, polynucleotide phosphorylase, xanthine oxidase catalysis under, carry out in described mixed system as follows reaction:
3,5-adenosine diphosphate (ADP) → AMP → hypoxanthine → hydrogen peroxide；
And peroxidase, 4-AA, 4-chlorophenol existence under, hydrogen peroxide carries out chromogenic reaction；With
(iii-1) absorbance A 3 or the absorbance rate of change Δ A3 of every individual system are measured, thus obtaining the 3rd standard value or the 3rd standard curve.
In above-mentioned detection method, 3,5 described adenosine diphosphate (ADP) concentration are preferably 20～60mmol/L；3,5 described nucleoside diphosphate acid enzyme concentrations are preferably 1～6ku/L；Described 5-nucleotidase concentration is preferably 1～3ku/L；Described ADA Adenosine deaminase concentration is preferably 1～3ku/L；Described polynucleotide phosphorylase concentration is preferably 1～3ku/L；Described xanthine oxidase concentration is preferably 1～3ku/L；Described peroxidase concn is preferably 0.5～1.5KU/L；Described 4-AA concentration is preferably 0.1～0.5g/L；Described 4-chlorophenol concentration is preferably 0.1～0.8g/L.
Mixed system of the present invention can also contain other additive, and described additive comprises buffer, surfactant, stabilizer, activator, chelating agen, protective agent.
Wherein, described buffer is preferably the buffer that pH is 5.5～7.5；It is preferred that described buffer is two (2-ethoxy) imino group Pehanorm buffer；More preferably, described buffer be 50～100mmol/L, pH be 6.50 two (2-ethoxy) imino group Pehanorm buffer.
Described surfactant is preferably nonionic surfactant；It is preferred that described surfactant includes, but is not limited to: TritonX, tween, span, AEO-9 etc.；Described surfactant concentration is 0.1 ‰～1 ‰ (percentage by weights).
Described stabilizer includes, but is not limited to: bovine serum albumin, β-dredge base ethanol, Cys；It is preferred that described stabilizer concentration is 0.1～0.5g/L.
Described activator includes, but is not limited to: anhydrous calcium chloride, Magnesium sulfate heptahydrate, potassium ferrocyanide, zinc sulfate；It is preferred that described activator concentration is 1～3mmol/L.
Described chelating agen includes, but is not limited to: EDTA and sodium salt, polymethylacrylic acid, organic multicomponent phosphonic acids, hydroxyethylethylene diamine tri-acetic acid (HEDTA), bicine N-；It is preferred that described chelating agent concentrations is 0.1～0.2g/L.
Described protective agent includes, but is not limited to: sucrose, glucose, fucose, maltose, lactose；It is preferred that described protective agent concentration is 1～5g/L.
Described additive can provide the reaction environment of necessity for enzymatic reaction；The homogeneity that can make reagent is better, test result is reproducible；Various enzyme can be protected, improve enzyme stability in preservation process；Reagent reacting sensitivity can be improved；The interference of other ions can be got rid of；Enzymatic activity can be maintained.
Described additive also comprises preservative, and described preservative includes, but is not limited to Hydrazoic acid,sodium salt；Gentamycin sulfate；Proclin300；Germall 115；MIT etc..The concentration of described preservative is preferably 0.1g/L～0.5g/L.
The present invention has a major advantage in that:
1. providing a kind of method detecting lithium ion, described method passes through enzymic colorimetric, can detect or monitor the lithium ion in patient blood in common biochemical instruments, and described method has easy and simple to handle, quick, reproducible advantage.
2. additionally provide a kind of test kit for above-mentioned detection method.
Below in conjunction with being embodied as, the present invention is expanded on further.Should be understood that these embodiments are merely to illustrate the present invention rather than restriction the scope of the present invention.The experimental technique of unreceipted actual conditions in the following example, generally conventionally condition, or according to manufacturer it is proposed that condition.Unless otherwise indicated, otherwise percentage ratio and number are calculated by weight.
Detection method
Hitachi 7170 biochemical instruments detects the absorption intensity of target solution；Adopt speed A method, read point 27-34, ten minutes response time, dominant wavelength: 546nm, commplementary wave length: 700nm.
Sample application amount is 5 μ l, and the application of sample amount of test agent () is 200 μ l, and the application of sample amount of test agent (two) is 100 μ l.
Embodiment 1 makes standard curve
1.1 preparation reaction reagents
1.1.1 test agent () (or being called R1 reagent) is prepared by table 1:
Table 1
1.1.2 test agent (two) (or being called R2 reagent) is prepared by table 2:
Table 2
1.2 make standard curve
Take R1 reagent 200 μ l, the R2 reagent 100 μ l of above-mentioned preparation, mix with the serum sample of following lithium concentration respectively: 0.00mmol/L(deionized water), 1.55mmol/L, each 5 μ l.Adopting the mode of two-point calibration, calibrating mode is linear gauging.
1.2.1 detection method
According to the parameter in detection method, in reaction cup, add 5 μ l serum samples, be subsequently adding the R1 reagent of 200 μ l, after hatching 5 minutes, add the R2 reagent of 100 μ l.After three minutes, start to use 546nm wavelength detecting continuously.
1.2.2 testing result
(1) testing result is as shown in table 3, and the calibration curve of formation is as it is shown on figure 3, Fig. 4 is lithium concentration is the response curve of the sample of 1.55mmol/L.
Table 3
| Sample | 1 | 2 |
| Concentration (mmol/L) | 0.00 | 1.55 |
| ΔA546nm(1*10-4Abs) | 6 | 763 |
(2) with 1.00mmol/L and 2.50mmol/L for two Quality Control solution.After calibration, the error of two Quality Controls of detection, result is as shown in table 4.
Table 4
Result shows: the accuracy of Quality Control detection, repeatability all meets the prescription of external diagnosis reagent.The detection method of the present invention meets the prescription of external diagnosis reagent.
Embodiment 2
By 3.00mmol/L serum sample with normal saline by the dilution proportion of 1/15,1/5,2/5,3/5,4/5,5/5, obtain diluted sample 4.1,4.2,4.3,4.4,4.5 (theoretical concentration refers to table 5).
According to the calibration results (standard curve as shown in Figure 3 namely obtained according to embodiment 1) of embodiment 1, obtain the test concentrations that each sample is corresponding.
Table 5
| Sample | Theoretical concentration | Test concentrations | Predicted concentration | Deviation |
| 4.1 | 0.20 | 0.21 | 0.195 | -7.14% |
| 4.2 | 0.60 | 0.59 | 0.605 | 2.54% |
| 4.3 | 1.20 | 1.23 | 1.22 | -0.81% |
| 4.4 | 1.80 | 1.82 | 1.835 | 0.82% |
| 4.5 | 2.40 | 2.44 | 2.45 | 0.41% |
| 4 | 3.00 | 3.08 | 3.065 | -0.49% |
Obtain linear regression graph as shown in Figure 1 according to theoretical concentration and test concentrations, and the linear regression graph according to theoretical concentration and Fig. 1 obtains predicted concentration.
Result shows: in the concentration range of linear high level 3.00mmol/L, and curvilinear correlation is fine, and R2 0.99 meets clinical needs.And the predicted concentration that obtains according to the linear regression graph of Fig. 1 is only small with the deviation of test concentrations, meets clinical needs.
Visible, the standard curve error that method of testing of the present invention obtains is little, entirely appropriate clinical practice.
Embodiment 3
According to the calibration results of embodiment 1, (standard curve namely obtained according to embodiment 1 as shown in Figure 3), tests 40 examples to the serum of the patient that its blood lithium concentration is monitored or blood sample.
Meanwhile, adopt ISE method, on U.S.'s Easylyte electrolyte analyser, namely measure these 40 described example samples (its supporting calibration solution, flushing liquor, purchased from American MEDICA company).
Test result is as in figure 2 it is shown, result shows: the detection method of the present invention is good with ISE method dependency, and slope is 1.0098, R2 is 0.9975.(abscissa and vertical coordinate unit: mmol/L)
Embodiment 4
Test agent corkage after embodiment 1 being calibrated is placed in Hitachi 7170 biochemical instruments agent bin, every 5 days, with Quality Control solution (Quality Control 1 target value: 1.00mmol/L；Quality Control 2 target value: 2.50mmol/L) stability of test agent behind detection Kaifeng.Experimental result is as shown in table 6.
Table 6
| Quality Control 1 | Quality Control 2 | |
| The detection date | Measured value | Measured value |
| January 5 | 1.03 | 2.48 |
| January 10 | 1.01 | 2.43 |
| January 15 | 1.03 | 2.46 |
| January 20 | 0.98 | 2.51 |
| January 25 | 0.99 | 2.43 |
| January 30 | 1.02 | 2.55 |
| February 4 | 0.99 | 2.51 |
Result shows: the test agent of the present invention, and Kaifeng rear stability is good, and the test result of Quality Control solution all fluctuates near target value, and need not repeat calibration in one month, fully meets the use of clinic.
In sum, the detection method adopting the present invention can detect lithium ion content or concentration in sample quickly, effectively, easily, and detectable provided by the invention is all highly stable, properties all meets the requirement of Clinical Laboratory, can be promoted use in clinic completely.
The all documents mentioned in the present invention are incorporated as reference all in this application, are individually recited as reference such just as each section of document.In addition, it is to be understood that after the above-mentioned teachings having read the present invention, the present invention can be made various changes or modifications by those skilled in the art, these equivalent form of values fall within the application appended claims limited range equally.
Claims (6)
1. a nondiagnostic measures the method for lithium ion in sample, it is characterised in that include step:
(1) providing a liquid mixed system, described system contains sample to be tested, 3,5-adenosine diphosphate (ADP)s and 3,5-nucleoside diphosphate acid enzymes；
(2) under the catalysis of 3,5-nucleoside diphosphate acid enzymes, described mixed system carries out reaction as follows:
3,5-adenosine diphosphate (ADP) → AMP
(3) measure in described system AMP presence or absence and/or quantity and/or production rate, and convert and draw the measurement result of lithium ion in sample；
Wherein, the method for described mensuration AMP presence or absence and/or quantity and/or production rate, including step:
(3-1) 5-nucleotidase, ADA Adenosine deaminase, polynucleotide phosphorylase, xanthine oxidase exist under, by the reaction system of step (2) or the AMP in system is carried out as follows reaction:
AMP → hypoxanthine → hydrogen peroxide
(3-2) production rate of hydrogen peroxide and/or quantity in the reaction system of measuring process (3-1), and draw the measurement result of AMP；
Wherein, in step (3-2), the method measuring hydrogen peroxide quantity includes step:
Peroxidase, 4-AA, 4-chlorophenol existence under, hydrogen peroxide in the reaction system of step (3-2) is carried out chromogenic reaction, the absorbance of assaying reaction system or absorbance rate of change, thus drawing production rate and/or the quantity of hydrogen peroxide.
2. the method for claim 1, it is characterised in that
In step (1), possibly together with 5-nucleotidase, ADA Adenosine deaminase, polynucleotide phosphorylase, xanthine oxidase, peroxidase, 4-AA, 4-chlorophenol in described mixed system；And
Step (2) and step (3) are replaced by following steps (ii) and step (iii):
(ii) 3,5-nucleoside diphosphate acid enzymes, 5-nucleotidase, ADA Adenosine deaminase, polynucleotide phosphorylase, xanthine oxidase catalysis under, carry out in described mixed system as follows reaction:
3,5-adenosine diphosphate (ADP) → AMP → hypoxanthine → hydrogen peroxide；With
Peroxidase, 4-AA, 4-chlorophenol existence under, hydrogen peroxide carries out chromogenic reaction；With
(iii) measure absorbance or the absorbance rate of change of described system, and draw the measurement result of lithium ion in sample.
3. the method for claim 1, it is characterised in that described mixed system is possibly together with buffer, surfactant, stabilizer, activator, chelating agen, protective agent.
4. method as claimed in claim 3, it is characterised in that described buffer is pH is the buffer of 5.5～7.5.
5. method as claimed in claim 3, it is characterised in that described surfactant is nonionic surfactant.
6. method as claimed in claim 3, it is characterised in that
Described stabilizer includes bovine serum albumin, β-dredge base ethanol, Cys or its combination；And/or
Described activator includes anhydrous calcium chloride, Magnesium sulfate heptahydrate, potassium ferrocyanide, zinc sulfate or its combination；And/or
Described chelating agen includes EDTA and sodium salt, polymethylacrylic acid, organic multicomponent phosphonic acids, hydroxyethylethylene diamine tri-acetic acid (HEDTA), bicine N-or its combination；And/or
Described protective agent includes sucrose, glucose, fucose, maltose, lactose or its combination.
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