CN1137381C - Turbidity measuring ultramicro-detecting technology and method using multifunctional biochemical enzyme labelling analyzer - Google Patents
Turbidity measuring ultramicro-detecting technology and method using multifunctional biochemical enzyme labelling analyzer Download PDFInfo
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- CN1137381C CN1137381C CNB001343912A CN00134391A CN1137381C CN 1137381 C CN1137381 C CN 1137381C CN B001343912 A CNB001343912 A CN B001343912A CN 00134391 A CN00134391 A CN 00134391A CN 1137381 C CN1137381 C CN 1137381C
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Abstract
The present invention provides a method for measuring turbidity ultramicro detecting technology by using a multifunctional biochemical enzyme labeling analyzer, which utilizes the existing micropore plate to replace 96 cuvettes and directly carry out a turbidity test in a pore, and the vein hemospasia can be omitted by jointly using a trace quantity blood sampling plastic pipe, a quantitative adding sample and a liquid transmitting device. A measured sample is measured by a standard substance, the unit content number can be obtained by adopting linear regression according to the quantitative value of the sample after the absorbance is measured by adopting a standard solution, a standard curve and a standard pipe are replaced, so the present invention can greatly improve the detecting speed in a turbidity detecting item, the corresponding assorted multifunctional biochemical enzyme labeling analyzer is combined to test, and a rapid detection system of the turbidity is improved.
Description
The invention belongs to field of medical examination.Be a kind of method of testing the ultramicron Fast Detection Technique of turbidity project with multifunctional biochemical enzyme label analyzer specifically.
The turbidity check is one of important component part of medical test.Nephelometric analysis towards trace, fast, accurately, easily direction to develop be the inexorable trend of current domestic and international medical test.
In the turbidity Interventions Requested, be material with blood, patient be can't do without vein haemospasia, just necessarily requires the sample of every mensuration the least possible for fear of it, but the accuracy that does not influence the result is the difficult point that current domestic and international turbidity check exists.
Check the method for turbidity at present, what the laboratory was the most frequently used is to compare with artificial standard turbidity pipe ready visual contrast, or the transmittance turbidimetry, with tintmeter and turbidity instrumentation absorbance, be abscissa again with concentration, each concentration absorbance is that ordinate preparation standard curve checks in content.These method good stabilities, but need drawing standard curve or calculating, needing single sample colorimetric operation, work efficiency is not high.And, can show the project content of being measured immediately with laser immunoturbidimetry instrument colorimetric, and promptly convenient, quick again, must the single sample colorimetric estimation but also exist, and when detecting next sample, the inconvenience of test again behind the mixing again again.If can realize adopting trace, RAPID COLORIMETRIC ASSAY FOR, can test the equipment and the technical method of 90 parts of samples behind mixing of sample, the practical significance that then has bigger technical superiority and use.
The objective of the invention is to avoid above-mentioned the deficiencies in the prior art part and adopt existing microwell plate to replace cuvette, use the tip trace blood, determine after the absorbance again quantitative values with the standard solution of variable concentrations according to sample, adopt linear regression to obtain the unit content number, make it to reach a kind of method that improves detection speed with multifunctional biochemical enzyme label analyzer test turbidity ultramicron detection technique.
Purpose of the present invention can reach by following measure:
A kind of method with multifunctional biochemical enzyme label analyzer test turbidity detection technique is characterized in that:
A. press tip routine blood test acquisition method: with trace blood plastic tube upper port contact drop of blood, lower in the lower end, the limit is gently squeezed the limit and is entered blood scorification lower end to the requirement, clutch sealing or open-ended with tweezers, the trace blood plastic tube is converted in the syringe needle plastic jacket of " V " type insertion disposable syringe, stick the adhesive sticker sequence number on syringe needle plastic casing or blood sampling plastic tube, centrifuging serum is standby;
B. get one of clean 8 * 12 standards, 96 microwell plate;
C. adopt the micro quantitative determination sample injector, get standard items and sample, quantitatively add in each hole of microwell plate;
D. use micropipettor,, quantitatively add in each hole of above-mentioned gauge orifice and sample by required reagent dosage;
E. microwell plate is put in the multifunctional biochemical enzyme label analyzer measuring chamber, in 90s, printed 1～90 part of result's report simultaneously.
A kind of method with multifunctional biochemical enzyme label analyzer test turbidity detection technique is characterized in that:
A. press tip routine blood test acquisition method: with trace blood plastic tube upper port contact drop of blood, lower in the lower end, the limit is gently squeezed the limit and is entered blood scorification lower end to the requirement, clutch sealing or open-ended with tweezers, the trace blood plastic tube is converted in the syringe needle plastic jacket of " V " type insertion disposable syringe, stick the adhesive sticker sequence number on syringe needle plastic casing or blood sampling plastic tube, centrifuging serum is standby;
B. get one of clean 8 * 12 standards, 96 microwell plate;
C. adopt the micro quantitative determination sample injector, get sample, quantitatively add in each hole of microwell plate;
D. use micropipettor,, quantitatively add in each hole of above-mentioned sample by required reagent dosage;
E. prepare the standard solution of four kinds of concentration;
F. quantitatively drawing standard solution with adjustable pipette adds respectively in each two hole of microwell plate;
G. after the absorbance that determines with four kinds of standard solution again according to the quantitative values of sample, adopt linear regression to obtain the unit content number;
H. microwell plate is put in the multifunctional biochemical enzyme label analyzer measuring chamber, setting unit content number prints 1～90 part of result's report simultaneously in 90s.
Purpose of the present invention can also reach by following measure:
The present invention adopts 1 μ l, 2 μ l, 3 μ l, 5 μ l, 10 μ l, 20 μ l, 25 μ l micro quantitative determination sample injectors, gets standard items and sample, quantitatively adds in each hole of microwell plate.
The present invention adopts the standard solution of 5,10,15,20 unit concentration.
The present invention adopts 100-500 μ l trace adjustable pipette quantitatively to draw respectively in each two hole of standard solution 300 μ l adding microwell plate.
The present invention adopts 100-500 μ l trace adjustable pipette quantitatively to draw in each hole of the required reagent adding of 300 μ l microwell plate.
The present invention adopts the vinyl tube of internal diameter 2-3mm to make the trace blood plastic tube of 5-7cm specification.
The present invention's method compared to existing technology has following advantage:
1. in the turbidity Interventions Requested, be material how with blood, patient be can't do without vein haemospasia, and the method for the present invention adopts a kind of ultramicron detection technique with multifunctional biochemical enzyme label analyzer test turbidity project has been avoided the venous patient blood drawing.
2. some test item, as: thymol turbidity, serum potassium etc., in full-automatic, semi-automatic biochemical analyzer, can not detect, and the method for the present invention adopts a kind of ultramicron detection technique with multifunctional biochemical enzyme label analyzer test turbidity project has remedied above-mentioned deficiency.
3. China's most laboratory is to turbidity test at present, and the most frequently used is to compare with artificial standard turbidity pipe ready visual contrast, or with tintmeter, turbidity instrumentation absorbance, is abscissa again with concentration, and each concentration absorbance is that ordinate preparation standard curve checks in content.Detection speed is slow, work efficiency is not high.And a kind of method of testing the ultramicron detection technique of turbidity project with multifunctional biochemical enzyme label analyzer that the present invention adopts, realized need not the drawing standard curve, need not artificial colorimetric and calculating, in sample in enormous quantities detects, analysis speed is fast, and 90 duplicate samples only can test printing go out result's report with 90s.
4. the method for a kind of ultramicron detection technique with multifunctional biochemical enzyme label analyzer test turbidity project of adopting of the present invention, after only need utilizing the titer of variable concentrations to determine absorbance, again according to the quantitative values of sample, draw the unit content number with linear regression, in surveying, this visual inspection only in batch reagent, replaced standard items, typical curve and standard pipe later on this unit content number, simplified the detecting operation step, improved work efficiency and saved reagent.
5. the result is accurate, good reproducibility.The method of a kind of ultramicron detection technique with multifunctional biochemical enzyme label analyzer test turbidity project that the present invention adopts, its precision, linearity, related coefficient, standard deviation, the coefficient of variation etc. all are better than other color comparator.
Embodiment 1
Thymol turbidity test (TTT)
1. principle:
When liver had solid lesion, the quality and quantity of serum proteins all changed.Do the time spent when the barbitol buffer solution that serum and thymol are saturated, thymol can lower the dispersancy of serum lipoids; Under the situation of albumen (as albumin) attenuating that plays stabilization or mass change, globulin solubleness in low ionic strength buffer liquid lowers and precipitates.Sediment is the complex of globulin, lipoids and thymol.Its turbidity is compared with labor standard turbidity pipe, draws thymol turbidity unit.
2. reagent:
Thymol-barbitol buffer solution:
Take by weighing barbital sodium 2.06g, barbital 2.76g, adding distil water 1L heats to boiling.Be chilled to about 37 ℃, add 100g/L thymol alcohol solution 10ml, firmly jolting is placed room temperature (20-25 ℃) and is spent the night.Wherein thymol concentration is 1g/L.
3. the preparation of standard opacity tube:
Barium chloride solution (0.0481mol/L): accurately take by weighing barium chloride (BaCI
22H
2O) 1.175g, or anhydrous barium chloride 1g place in the 100ml volumetric flask, and the adding distil water dissolving also is diluted to 100ml, are stored in behind the mixing in the close plug bottle, can forever preserve.
Barium sulphate standard stoste:
In the 100ml volumetric flask, add 0.1mol/L sulfuric acid 70ml, slowly drip 0.0481mol/L barium chloride solution 3ml, with adding, add 0.1mol/L sulfuric acid at last to scale with shaking.This suspension is equivalent to 22.2 Maxwell units.
The preparation steps of thymol standard turbidity pipe
Maxwell unit 05 10 15 20
Barium sulphate stoste (ml) 0 1.35 2.70 4.05 5.40
0.1mol/L sulfuric acid (ml) 6 4.65 3.30 1.95 0.60
Abundant mixing, press following operation:
4. thymol standard turbidimetric analysis turbidimetry:
(1) gets one of clean 8 * 12 standards, 96 microwell plate.
(2) add respectively in each two hole of microwell plate with the quantitative standard solution 300 μ l that draw above-mentioned 5,10,15,20 variable concentrations units of 100-500 μ l trace adjustable pipette.
(3) microwell plate is put in the multifunctional biochemical enzyme label analyzer measuring chamber, used 630nm, single wavelength is established blank well and each gauge orifice position, starts to measure turbidity content and print the result simultaneously and the absorbance report.
(4) result: thymol standard turbidimetric analysis turbidimetry turbidimetry reagent: press reagent compound method preparation lot number: 002022 titer (product): autogamy before using: 5,10,15,20 Maxwell unit's wavelength modes: single wavelength measurement wavelength: 630nm report manner: 1. normal concentration test result 2. absorbance test result standard solution are provided with: 1=0000,2=0005,3=0010,4=0015;
Two kinds of report manners of 5=0020, measurement result are as follows: 1, capable 01,02 hole of normal concentration test result: A is a blank well, and 03,04 hole is " 0 "
The point hole, 05,06 hole is standard turbidity 5 units, 07,08
The hole is standard turbidity 10 units, and 09,10 holes are the standard turbidity
15 units, 11,12 holes are standard turbidity 20 unit testings knots
Really.2, absorbance test result: with corresponding each the hole absorbance of standard turbidimetric analysis turbidimetry result.
Normal concentration test result plate number: 06 date: 00-02-22 time: 08:28 blank well: mode: single wavelength identification code: 01 wavelength: 630 humidity: temperature: 01 02 03 04 05 06 07 08 09 10 11 12A+000.1+000.0-000.0-000.0+005.0+005.1+010.0+009.9+014.9+015.0+019.5+019.7
Absorbance test result plate number: 06 date: 00-02-22 time: 08:28 blank well: mode: single wavelength identification code: 01 wavelength: 630 humidity: temperature: blank well value :+0.035 01 02 03 04 05 06 07 08 09 10 11 12A+0.001-0.001-0.002-0.003+0.106+0.109+0.214+0.214+0.321+0.323+0.421+0.425
5. operation:
(1) tip blood is gathered:
A. the blood sampling site adult is advisable with the left hand third finger, and infant below half years old is usually with thumb or heel blood sampling.
B. massage blood sampling site gently, make it congested naturally, with 75% ethanol cotton balls sterilize local skin, wait to do.
C. tightly pinch thorn blood position, get blood with the puncture of sterile blood sampling pin.Action should be rapid, and the about 2-3mm of the degree of depth can flow out with squeezes blood a little and to be advisable.With trace blood plastic tube upper port contact drop of blood, lower in the lower end, the limit is gently squeezed the limit and is entered blood scorification lower end to the requirement, clutch sealing or open-ended with tweezers, the trace blood plastic tube is converted in the syringe needle plastic jacket of " V " type insertion disposable syringe, stick the adhesive sticker sequence number on syringe needle plastic casing or blood sampling plastic tube, centrifuging serum is standby;
(2) get one of clean 8 * 12 standards, 96 microwell plate;
(3) getting tested serum 5l with the micro quantitative determination sample injector adds in the above-mentioned micropore plate hole;
(4) quantitatively drawing reagent 300l with the 100-500l adjustable pipette adds in each hole of microwell plate;
(5) after the absorbance that determines with above-mentioned four kinds of standard solution again according to the quantitative values of sample, adopt linear regression to obtain the unit content number to be: 046.2.
(6) test enzyme is marked orifice plate and put in the multifunctional biochemical enzyme label analyzer measuring chamber, use 630nm, single wavelength is established the blank well position, confirms the Au*C formula, setting unit content number, and C=046.2 measures simultaneously in 90s and prints 1～95 part of turbidity test result report.
(7) result: thymol test: turbidimetry reagent: press reagent compound method preparation lot number: 002022 wavelength mode: single wavelength measurement wavelength: 630nm report manner: turbidity test result unit content number is provided with: the C=046.2 measurement result is as follows: turbidity test result: A capable 01,02 hole is former blank well, 03,04 hole is former " 0 "
The point hole, 05,06 hole is primary standard turbidity 5 units, 07,08
The hole is primary standard turbidity 10 units, and 09,10 holes are primary standard
Turbidity 15 units, 11,12 holes are primary standard turbidity 20 units
Test result.Errorless with primary standard concentration determination result after tested
Difference.All the other holes are the sample result.
Turbidimetric analysis turbidimetry is plate number as a result: 02 date: 00-02-22 time: 11:55 blank well: mode: single wavelength identification code: 01 wavelength: 630 humidity: temperature:
Negative control hole value (N):
Formula: Au*C
01 02 03 04 05 06 07 08 09 10 11 12A +000.1 +000.0 -000.0 -000.0 +005.0 +005.1 +010.0 +009.9 +014.9 +015.0 +019.5 +019.7B +015.4 +015.6 +015.3 +007.2 +012.1 +014.1 +008.6 +015.4 +009.8 +008.9 +015.2 +010.2C +014.3 +019.6 +009.6 +011.1 +011.4 +005.4 +012.3 +011.4 +021.0 +011.9 +020.3 +016.4D +011.7 +014.2 +005.7 +010.0 +011.0 +016.2 +011.8 +006.1 +008.9 +010.9 +014.0 +012.6E +009.6 +009.3 +014.6 +012.9 +007.8 +015.1 +012.0 +009.8 +007.3 +008.3 +010.3 +011.5F +009.1 +011.8 +009.1 +015.6 +014.7 +010.9 +013.2 +014.4 +012.9 +009.7 +008.4 +012.6G +010.1 +006.8 +013.8 +012.3 +006.8 +014.6 +009.2 +016.1 +007.2 +007.6 +011.7 +009.6H +009.9 +014.6 +011.4 +016.1 +021.7 +009.3 +010.4 +006.9 +010.3 +007.4 +014.0 +007.8
Embodiment 2
Serum potassium is measured-the new direct turbidimetry of tetraphenylboron-sodium
This law is different from the tetraphenylboron-sodium turbidimetry that is eliminated.This reagent is to add NaOH to improve pH value and increased sensitivity and made the result stable, and adds EDTA-Na2 and eliminate and disturb the basis upward to develop.This law and potassium sodium analyser comparative determination r=0.96, CV=3.98%, the recovery 95～103%.
1. principle:
Potassium ion and tetraphenylboron-sodium effect in the serum form water-fast tetraphenylboron potassium precipitation because reagent pH, has improved the sensitivity of test more than 12, and the turbidity of generation is directly proportional with the concentration of potassium ion within the specific limits.Can record the content of potassium in the serum according to turbidity.
2. reagent:
A, tetraphenylboron-sodium reagent: in the 100ml volumetric flask, add the about 70ml of physiological saline, 1mol/L NaOH 1ml, EDTA-Na2 75mg, tetraphenylboron-sodium 2.5g adds physiological saline to 100ml after the dissolving.
B, potassium titer 5mmol/L.
3. collection of specimens: press tip routine blood test acquisition method, with trace blood plastic tube upper port contact drop of blood, lower in the lower end, the limit is gently squeezed the limit and is entered blood scorification lower end to the requirement, clutch sealing or open-ended with tweezers, the trace blood plastic tube is converted in the syringe needle plastic jacket of " V " type insertion disposable syringe, sticks the adhesive sticker sequence number on syringe needle plastic casing or blood sampling plastic tube, centrifuging serum is standby;
4. application of sample: get according to the form below application of sample of clean 96 microwell plates:
Admixture (μ l) blank well gauge orifice is measured the hole
Distilled water 5
Titer (0.5mmol/L) 5
Serum (slurry) 5
Tetraphenylboron-sodium reagent 300 300 300
Mixing is put room temperature after 5 minutes, again mixing.
5. test: microwell plate is put in the multifunctional biochemical enzyme label analyzer measuring chamber, with the single wavelength of 630nm, establish blank well and gauge orifice position, confirmed the Au/As*C formula, the product unit content that sets up standard C=OO5.O carries out each hole serum potassium content of colorimetric measurement.
6. result: blood potassium is measured: new tetraphenylboron sodium turbidimetry reagent: press reagent compound method autogamy lot number: 000808 wavelength mode: single wavelength measurement wavelength: 630nm titer (product): 5mmol/L uses formula: the setting of Au/As*C standard items unit content: C=OO5.O 1, blood potassium measurement result: A capable 01,02 hole is blank " O " hole of transferring, 03,04 hole is the standard items measurement result, and all the other measure the hole result for sample.2, absorbance test result: with corresponding each the hole absorbance of blood potassium measurement result.
Blood potassium measurement result plate number: 01 date: 00-06-08 time: 10:27 blank well: mode: single wavelength identification code: 01 wavelength: 630 humidity: temperature:
Negative control hole value (N) :+0.265
Formula: Au/As*C
01 02 03 04 05 06 07 08 09 10 11 12A +000.1 -000.1 +004.9 +005.0 +004.0 +003.5 +004.3 +004.6 +003.8 +004.5 +003.5 +004.6B +004.3 +005.0 +005.4 +004.7 +005.1 +007.3 +004.7 +004.7 +004.2 +007.5 +004.8 +005.2C +002.4 +004.2 +002.5 +005.2 +005.4 +003.0 +003.1 +002.9 +002.3 +002.4 +002.1 +002.0
Absorbance test result plate number: 01 date: 00-08-08 time: 10:28 blank well: mode: single wavelength identification code: 01 wavelength: 630 humidity: temperature:
Blank well value :+0.045
01 02 03 04 05 06 07 08 09 10 11 12A +0.008 -0.009 +0.264 +0.265 +0.213 +0.187 +0.231 +0.246 +0.202 +0.240 +0.189 +0.249B +0.232 +0.267 +0.287 +0.252 +0.274 +0.389 +0.254 +0.249 +0.227 +0.102 +0.256 +0.275C +0.127 +0.227 +0.135 +0.273 +0.287 +0.161 +0.164 +0.156 +0.125 +0.125 +0.113 +0.109
Claims (6)
1. method with multifunctional biochemical enzyme label analyzer test turbidity is characterized in that:
A. press tip routine blood test acquisition method: with trace blood plastic tube upper port contact drop of blood, lower in the lower end, the limit is gently squeezed the limit and is entered blood scorification lower end to the requirement, clutch sealing or open-ended with tweezers, the trace blood plastic tube is converted in the syringe needle plastic jacket of " V " type insertion disposable syringe, stick the adhesive sticker sequence number on syringe needle plastic casing or blood sampling plastic tube, centrifuging serum is standby;
B. get one of clean 8 * 12 standards, 96 microwell plate;
C. adopt the micro quantitative determination sample injector, get sample, quantitatively add in each hole of microwell plate;
D. use micropipettor,, quantitatively add in each hole of above-mentioned sample by required reagent dosage;
E. prepare the standard solution of four kinds of concentration;
F. quantitatively drawing standard solution with adjustable pipette adds respectively in each two hole of microwell plate;
G. after the absorbance that determines with four kinds of standard solution again according to the quantitative values of sample, adopt linear regression to obtain the unit content number;
H. microwell plate is put in the multifunctional biochemical enzyme label analyzer measuring chamber, setting unit content number prints 1～90 part of result's report simultaneously in 90s.
2. the method with multifunctional biochemical enzyme label analyzer test turbidity according to claim 1, it is characterized in that: adopt 1 μ l, 2 μ l, 3 μ l, 5 μ l, 10 μ l, 20 μ l, 25 μ l micro quantitative determination sample injectors; get standard items and sample, quantitatively add in each hole of microwell plate.
3. the method with multifunctional biochemical enzyme label analyzer test turbidity according to claim 1 is characterized in that: the standard solution that adopts 5,10,15,20 unit concentration.
4. the method with multifunctional biochemical enzyme label analyzer test turbidity according to claim 1 is characterized in that: adopt 100-500 μ l trace adjustable pipette quantitatively to draw standard solution 300 μ l respectively and add in each two hole of microwell plate.
5. the method with multifunctional biochemical enzyme label analyzer test turbidity according to claim 1 is characterized in that: adopt 100-500 μ l trace adjustable pipette quantitatively to draw the required reagent of 300 μ l and add in each hole of microwell plate.
6. the method with multifunctional biochemical enzyme label analyzer test turbidity according to claim 1 is characterized in that: the vinyl tube of employing internal diameter 2-3mm is made the trace blood plastic tube of 5-7cm specification.
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| Application Number | Priority Date | Filing Date | Title |
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| CNB001343912A CN1137381C (en) | 2000-12-12 | 2000-12-12 | Turbidity measuring ultramicro-detecting technology and method using multifunctional biochemical enzyme labelling analyzer |
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| CN1137381C true CN1137381C (en) | 2004-02-04 |
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