CN108761070B - Urine transferrin detect reagent box of wide detection range - Google Patents
Urine transferrin detect reagent box of wide detection range Download PDFInfo
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- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
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Abstract
The invention discloses a wide detection range urine transferrin detection kit, which comprises a first reagent and a second reagent, wherein the first reagent comprises a metal ion additive, the metal ion additive is zinc salt, and the concentration of the zinc salt is 6-19 mM; the second reagent is latex particle solution marked with UTRF antibody, and the particle size range of the latex particles is 172-200 nm. The upper limit of the linear range of the kit in the invention exceeds 80mg/L, more than 90% of patients can be covered, the repeatability is good near a reference value of 2mg/L, and the judgment of a gray area sample is facilitated; in addition, the kit of the invention can replace imported reagents, thus saving the expense of patients.
Description
Technical Field
The invention relates to the technical field of biology, in particular to a urine transferrin detection kit with a wide detection range.
Background
Transferrin (UTRF) is a small molecule circulating protein with negative charges, and both transferrin and albumin are difficult to pass through glomerular filters under physiological conditions, but when the kidney is diseased, transferrin and albumin can leak out of the glomeruli, so urinary transferrin is a sensitive indication of glomerular filter damage. The uniqueness of transferrin not only makes it a feature for detecting normal renal function, but also enables its broader use in the detection of earlier, more mild renal injuries.
The transferrin content in normal adult urine is lower, usually less than 2mg/L, but the transferrin content in normal adult blood is up to 2200-4000mg/L, when the glomerulus is slightly damaged, the transferrin in blood leaks into the urine, so that the UTRF in the urine is obviously increased; in the middle and later stage nephropathy patients, UTRF leaks more, so that the concentration of UTRF in urine is as high as several hundred or even 1000 mg/L. At present, immunoturbidimetry reagents for detecting blood transferrin are commonly used in clinic, and urine transferrin cannot be detected due to the sensitivity problem. At present, a plurality of companies have reagents for determining the UTRF of urine, except for an imported urine transferrin detection reagent of Siemens and Beckmann Coulter; the Meikangbi, Beijing Baiotaikang and the company adopt a latex immunoturbidimetry method, wherein the detection lower limit of the Meikangbi, Beijing Baiotaikang and the company reagent can reach below 0.6mg/L, but the linear range of the Meikangbi, Beijing Baiotaikang and the company reagent is only 30mg/L with the highest linear range, and most manufacturers do not mark an antigen excess range.
In clinical application, the existing latex immunoturbidimetric reagent has the problem of too narrow linear range, and researches show that the UTRF in urine of more than 50% of early glomerulonephritis patients exceeds 30mg/L, and the UTRF in urine of less than 10% of early glomerulonephritis patients exceeds 80 mg/L. Therefore, the existing kit for detecting UTRF in urine by using a latex immunoturbidimetry reagent cannot meet the detection of the UTRF content in urine of most early glomerulonephritis patients at all, and a large number of samples are diluted and rechecked by using manpower or instruments, so that the labor intensity of hospital staff is increased, and the reagent loss cost is increased.
At present, the main reagents for detecting the UTRF content in urine are imported UTRF reagents in a Siemens BNII (2.2-35mg/L) system and UTRF reagents in a Beckmann Coulter (BC) IM800(2.0-40mg/L) system, both of which adopt a scattering immune turbidimetry method, although the detection range is relatively wide and can reach 35-40mg/L, and cover 60% of patients, the detection limit can only reach 2.2 and 2.0 mg/L. As the sample smaller than the minimum detection limit can not report the result, and the reference value of the item is 2mg/L, the clinical application is relatively embarrassed, the result can not be detected by the grey area sample, and the measurement repeatability of the weak positive sample (the UTRF content in urine is 2.0-5mg/L) is poor. In addition, the imported reagent is expensive, and the existing kit for detecting UTRF in urine has extremely slow detection speed which is less than 250t/h, and cannot meet the requirements of a large number of patients in a large hospital and massive samples for physical examination.
Based on the existing UTRF latex turbidimetric reagent, the detection range is expanded by an instrument predilution mode, the detection speed of a biochemical analyzer is reduced by 50% -70%, compared with the existing Siemens and BC reagents, the method has the advantages that the detection speed is not obviously improved, the detection range expanding capacity is limited, only 50-60mg/L can be achieved, and the requirement that the upper limit of a linear range exceeds 80mg/L cannot be met. In addition, the dosage of the antibody is greatly increased, although the dosage can be used for widening the linear range of the reagent, the dosage of the antibody is simply increased, so that the excessive range of the antigen is adversely affected, the detection cost is greatly increased, and the method for widening the linear range of the reagent by greatly increasing the dosage of the antibody under the background of national strict medical expenses obviously cannot be popularized and used.
Therefore, the technical personnel in the field need to solve the problem of providing a domestic urine transferrin detection kit which has a linear range upper limit exceeding 80mg/L, can cover more than 90% of patients, has good repeatability near a reference value of 2mg/L and is beneficial to judging gray zone samples.
Disclosure of Invention
In view of the above, the invention provides a urine transferrin detection kit with a wide detection range, wherein the upper limit of the linear range exceeds 80mg/L, more than 90% of patients can be covered, the repeatability around a reference value of 2mg/L is good, and the judgment of a gray area sample is facilitated; in addition, the kit of the invention can replace imported reagents, thus saving the expense of patients.
In order to achieve the purpose, the invention adopts the following technical scheme:
the urine transferrin detection kit with the wide detection range is characterized by comprising a first reagent and a second reagent, wherein the first reagent comprises a metal ion additive.
According to the urine transferrin detection kit disclosed by the invention, under the condition that the antigen surplus range meets 1000mg/L, the upper limit of the linear range exceeds 80mg/L, the gray area sample meets the detection of 2mg/L concentration, the precision CV is less than 2%, and the detection of more than 90% of patients can be met; in addition, the kit of the invention can completely replace imported reagents, thereby greatly saving the expense of patients.
Further, the metal ions in the metal ion additive include iron ions, copper ions, or zinc ions.
Iron ions, copper ions or zinc ions are used as metal ions in the metal ion additive, so that the low-end sensitivity of the transferrin test can be obviously improved. The saturation degree of the transferrin binding ions can influence the protein conformation, namely the capability of forming antigen-antibody complexes in immunoturbidimetric assay, and the higher the saturation degree is, the stronger the turbidity forming capability is. A certain amount of metal ions are added into the reagent I, the transferrin content of a low-value specimen is low, the saturation of combined ions is high, and turbidity signal values are high, on the contrary, the transferrin content of a high-concentration specimen is high, and the relative ion combined saturation is low, so that the turbidity forming capacity is not obviously changed.
Further, the metal ion additive is capable of binding to transferrin and is readily soluble in water; the metal ion additive comprises zinc salt, the concentration of the zinc salt is 6-19mM, and the concentration of the zinc salt is preferably 10-15 mM.
Further, the zinc salt is one or more of zinc sulfate, zinc chloride or zinc acetate
Wherein the metal ion additive also comprises iron salt and copper salt, and the iron salt is ferrous sulfate; the copper salt is copper sulfate.
Although iron salt, copper salt or zinc salt can be used as a metal ion additive in the first reagent, ferrous sulfate and copper sulfate have great influence on the color of the first reagent solution, and the relative cost is high; the zinc salt is dissolved, and the color of the reagent solution is not greatly influenced, and the zinc salt has more chemical applications and lower cost, so the zinc salt is preferably adopted as the metal ion additive.
Further, the reagent II is a latex particle solution marked with UTRF antibody; the particle size range of the latex particles is 172-200 nm; the preferred particle size range for the latex particles is 185-200 nm.
The use of latex with smaller particle size can reduce the sensitivity of the reagent as a whole, and can expand the linear range by using the theoretical basis of contradiction between sensitivity and linearity and the antigen surplus range, but the direct use of latex with smaller particle size can cause low-end sensitivity and cause the problem that gray zone samples cannot be distinguished. Even if a sensitizer is added to improve the sensitivity of the reagent as a whole, increasing the sensitivity of the low end results in an excessively high signal value of high-value sensitivity, and decreases the linear range and the antigen excess range.
Therefore, in the detection of urine UTRF, when the low-value sensitivity loss caused by using latex with smaller particle size and the improvement of low-value sensitivity brought by adding a proper amount of metal ions into the reagent are mutually offset, and the improvement slightly affects the high-value sensitivity, the linear range of the urine UTRF kit can be expanded, and the low-value sensitivity can be maintained, so that the accuracy of measuring the grey zone sample is ensured.
Further, the reagent I also comprises sodium chloride or potassium chloride with the concentration of 400 mM/L; a sensitizer at a concentration of not more than 20g/L and a dispersant at a concentration of 5 ml/L. Among them, potassium chloride acts in the same manner as sodium chloride in the present kit, but potassium chloride is expensive in cost as compared with sodium chloride, and sodium chloride is preferable in the present invention.
Preferably, the first reagent further comprises a preservative, a buffer and the like; wherein the preservative comprises sodium azide or Proclin series; the buffer has a pH of 5.5-8.0 and comprises citric acid, Good's, phosphate, glycine or Tris buffer. According to the kit, a proper amount of preservative and buffer can be added according to actual detection conditions, wherein UTRF is insensitive to pH value change, so that the addition of the preservative, the buffer and other reagents does not influence the detection performance of the kit.
Further, the sensitizer is one or more of polyethylene glycol PEG4000, polyethylene glycol PEG6000, polyethylene glycol PEG8000, polyethylene glycol PEG12000 or polyethylene glycol PEG 20000.
The invention also utilizes a secondary convergence polymerization latex modification technology disclosed in the patent No. 201710986970.7 with the patent name of 'a method for improving the antigen surplus and linear range of latex immunoturbidimetry', and realizes the improvement of the linear range of the urine UTRF detection kit under the conditions that the antibody feeding volume is not increased, namely the cost is not changed, the detection speed is not reduced by using pre-dilution of an instrument, and the antigen surplus range meets 1000 mg/L. The invention patent with patent number 201710986970.7 (a method for improving antigen surplus and linear range of latex immunoturbidimetry) utilizes a secondary convergence polymerization latex modification technology, and can expand the linear range of the UTRF detection reagent to 0.2-40mg/L under the conditions that the antibody feeding volume is not increased, instruments are not used for pre-dilution, and the antigen surplus range meets 1000mg/L, thereby reaching the linear range of the imported reagent, and the low-value sensitivity is superior to that of the imported reagent, but can not completely meet the clinical requirements. Compared with the invention patent with patent No. 201710986970.7 (a method for improving antigen surplus and linear range of latex immunoturbidimetry), the kit of the invention can realize the detection that the upper limit of the linear range of the kit exceeds 80mg/L and the gray area sample meets the concentration of 2mg/L under the condition that the antigen surplus range meets 1000mg/L, and the precision CV of the kit is less than 2 percent and can meet the detection of more than 90 percent of patients.
According to the technical scheme, compared with the prior art, the urine transferrin detection kit with a wide detection range has the following technical advantages:
(1) according to the kit, the metal ion additive is added into the reagent I, the proper particle size range of latex particles is selected, the loss of low-value sensitivity caused by using latex with smaller particle size and the improvement of low-value sensitivity caused by metal ions are mutually offset, and the improvement slightly affects high-value sensitivity, so that the linear range of the kit can be expanded, the low-value sensitivity can be maintained, and the accuracy of measuring the gray zone sample is ensured.
(2) Under the condition that the antigen surplus range meets 1000mg/L, the upper limit of the linear range exceeds 80mg/L, the gray area sample meets the detection of 2mg/L concentration, the precision CV is less than 2%, and the detection of more than 90% of patients can be met;
(3) the kit provided by the invention can replace imported reagents for detection, so that the expense of patients is saved.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, it is obvious that the drawings in the following description are only embodiments of the present invention, and for those skilled in the art, other drawings can be obtained according to the provided drawings without creative efforts.
FIG. 1 is a graph showing the effect of different concentrations of zinc acetate on the sensitivity of the assay for different concentrations of UTRF provided by the present invention.
FIG. 2 is a graph showing the effect of different concentrations of ferrous sulfate on the sensitivity of the determination of different concentrations of UTRF provided by the present invention.
FIG. 3 is a graph showing the effect of different concentrations of copper sulfate on the sensitivity of the assay for different concentrations of UTRF provided by the present invention.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Example 1
The linear range of the first generation UTRF reagent of this company was expanded by the method described in patent No. 201710986970.7, i.e., the secondary convergent polymerization latex modification technique, to obtain reagent a, and the performance of the first generation UTRF reagent and reagent a was verified.
First generation UTRF reagent: 20162400121 is registered by Shanghai dynasty, the linear range is 0.2-24mg/L, and the antigen surplus range is 1000 mg/L; the main components are as follows:
a first reagent: sodium chloride with concentration of 400 mM/L; polyethylene glycol PEG6000 with the concentration of 1 g/L; tween-20 with concentration of 5 ml/L;
and a second reagent: latex particles, 3g/L, latex particle size 297 nm; UTRF antibody (cat # Q0327), 10ml/L, brand Dako.
The method comprises the following specific steps of carrying out secondary convergence polymerization latex modification on a first generation UTRF reagent:
(1) 500ml of a latex solution of Polymicrospheres, having the commercial designation CB0297C, containing 10% by mass of latex having a particle size of 297nm, was used in the same manner as the first generation UTRF reagent, and subjected to cross-flow filtration by adding distilled water through a 50KD cut-off pore size ultrafiltration membrane of Millipore, to thereby complete 8-fold liquid exchange.
(2) Adding the latex solution after replacing distilled water into a four-port reaction kettle with the capacity of 1000ml, adding 1.5g of lithium chloride and 0.4g of sodium dodecyl benzene sulfonate, placing the reaction kettle in a constant-temperature water bath kettle, keeping the constant temperature of 50 ℃, and mechanically stirring at 260rpm for 10 min. Then argon is introduced to remove oxygen for 5min, 0.15g of potassium persulfate is added to initiate the convergent polymerization reaction in the microspheres for the second time, and the reaction is continued for 10h at a constant temperature of 50 ℃ under the mechanical stirring of 260 rpm.
(3) The latex solution after completion of the convergent polymerization reaction was added to a MES-NaOH solution having a concentration of 50mM/L and a pH of 5.0, and subjected to cross-flow filtration through a 50KD cut-off pore size ultrafiltration membrane cartridge manufactured by Millipore corporation to complete 16-fold liquid exchange.
Particle size D of the latex in the latex solution before the Convergence polymerization1297nm, and the particle diameter of the latex in the latex solution is D after the latex is reformed by secondary convergence polymerization2＝0.78D1＝231nm。
The reagent A mainly comprises the following components:
a first reagent: sodium chloride with concentration of 400 mM/L; polyethylene glycol PEG6000 with the concentration of 1 g/L; tween-20 with concentration of 5 ml/L; preservative PC-300 with the concentration of 0.35 ml/L;
and a second reagent: the concentration of latex particles after secondary convergence polymerization is 3g/L, and the particle diameter of the latex is D2(D2＝0.78D1)231 nm; UTRF antibody (cat # Q0327), 10ml/L, brand Dako, reagent two was prepared using conventional chemical coupling.
Meanwhile, 2ul of sample, 105ul of reagent I and 35ul of reagent II are used for the first generation UTRF reagent and the reagent A, and the performance is verified on Hitachi7180 under the conditions of 570nm wavelength and 18-34 read points, and the specific data are shown in tables 1-3.
TABLE 1 first generation UTRF reagent calibration data
| mg/ |
0 | 1.5 | 3 | 6 | 12 | 24 |
| ΔABS | 6 | 433 | 912 | 1990 | 4161 | 9374 |
Table 2A reagent calibration data
| mg/ |
0 | 2 | 10 | 20 | 40 |
| ΔABS | 2 | 275 | 1533 | 3471 | 7592 |
TABLE 3 comparison of the first generation UTRF reagents and reagent A Linear Range and antigen excess Range
As can be seen from tables 1-3, the upper limit of the linear range of the reagent A can be expanded to be more than 40mg/L and the antigen surplus range can reach 1000mg/L compared with the first generation UTRF reagent.
Example 2
Verification of the Effect of the Metal additives
Using reagent A, different concentrations of zinc acetate, copper sulfate and ferrous sulfate were added to reagent A, reagent one, respectively, and Δ ABS was measured using 2ul of sample, 105ul of reagent one, 35ul of reagent two, using the same lot of UTRF standards of different concentrations at 570nm wavelength, at 18-34 read points on Hitachi7180, with the results shown in tables 4-6 and FIGS. 1-3.
TABLE 4 Effect of different concentrations of Zinc acetate on the sensitivity of determination of different concentrations of UTRF
TABLE 5 influence of ferrous sulfate of different concentrations on the sensitivity of determination of UTRF of different concentrations
TABLE 6 influence of different concentrations of copper sulfate on the sensitivity of determination of different concentrations of UTRF
As can be seen from tables 4-6 and FIGS. 1-3, when zinc acetate, copper sulfate or ferrous sulfate is added to the first reagent of reagent A, a small amount of metal ions can significantly improve the sensitivity of the low concentration UTRF assay (about 120% improvement), and as the concentration of metal ions increases, the sensitivity improvement of the low concentration (2mg/L) UTRF tends to be stable (2mg/L concentration sensitivity change in the third example of Table 6), and the sensitivity of the medium and high concentration (e.g., 10mg/L) UTRF assay starts to gradually increase as the addition amount of metal ions increases, but when the concentration of metal additives is too high, e.g., 24mM/L, the linear range is 40mg/L, the ABS is out of bounds, and the requirement that the upper limit of the linear range exceeds 40mg/L cannot be met. In addition, in all cases, the linear range of the metal additive simply added to the first reagent of the reagent A cannot reach 80mg/L, and ABS is out of limits.
Wherein, the reagent I added with copper sulfate is light blue, the reagent I added with ferrous sulfate is light green, which does not meet the requirement that most of the reagents I in the field are colorless or light yellow clear solution, and the cost is slightly higher, which is not preferable, and the zinc salt with colorless solution and low industrialization price is used in the subsequent embodiment.
Example 3
The conventional thought in the field is used, the latex particle size is reduced in a certain range, and the linear range of the reagent is expanded.
D was obtained by replacing the latex having a particle diameter of 297nm of Polymicrospheres company, reagent A, of example 1 with a latex having a particle diameter of 200nm of Bangslab, according to the technique for modifying a latex for secondary convergence polymerization described in patent No. 201710986970.73Reagent B was obtained from a latex with 0.78 × 200nm to 156nm, the main components of reagent B being:
a first reagent: sodium chloride with concentration of 400 mM/L; polyethylene glycol PEG6000 with the concentration of 1 g/L; tween-20 with concentration of 5 ml/L; preservative PC-300 with the concentration of 0.35 ml/L;
and a second reagent: the concentration of the latex particles after secondary convergence polymerization is 3g/L, and the particle diameter D of the latex3Is 156 nm; UTRF antibody (cat # Q0327), at a concentration of 10ml/L, brand Dako, reagent two was prepared using conventional chemical coupling.
In reagent B, zinc acetate was added at different concentrations to reagent A, reagent B was added to reagent B, 2ul of the sample, 105ul of reagent A, and 35ul of reagent B were used, and in Hitachi7180, the same lot of UTRF standards at different concentrations were used at a wavelength of 570nm and at a reading point of 18 to 34, to measure Δ ABS, and the results are shown in tables 7 to 9.
TABLE 7 Effect of different concentrations of Zinc acetate on Δ ABS in different concentrations of UTRF
TABLE 8 influence of different concentrations of Zinc acetate on the precision of the determination of the concentration of the reference values
| Concentration of Zinc acetate | mean(mg/L) | SD | CV |
| 0mM | 2.03 | 0.173 | 8.52％ |
| 6mM | 2.02 | 0.032 | 1.58％ |
| 12mM | 2.03 | 0.033 | 1.63％ |
| 19mM | 2.01 | 0.03 | 1.49％ |
| 24mM | 2.06 | 0.031 | 1.50％ |
TABLE 9 Effect of different concentrations of Zinc acetate on the antigen excess range and Linear range
This example reduces the latex particle size to 200nm by using the double convergent polymerization latex modification technique described in patent No. 201710986970.7 and adding 6-19mM zinc acetate to reagent one of reagent B. As can be seen from Table 8, the reagents of this example achieve a precision CV of < 2% at 2 mg/L; as is clear from tables 7 and 9, the reagent of the present example has a wide linear range of UTRF reagent, in which the upper limit of the linear range is more than 80mg/L and the antigen excess range satisfies 1000 mg/L. Meanwhile, with the increase of the concentration of the zinc acetate, the sensitivity of the reagent is gradually improved when the reagent is used for measuring a high value, but the antigen surplus range is gradually reduced; when the concentration of the zinc acetate is 6-12mM, the antigen surplus range is as high as 1200mg/L or more; however, when the concentration of zinc acetate is 19mM, the theoretical antigen surplus range is 1000mg/L, but the linear range of the concentration measurement is only 80.16mg/L, and the antigen surplus range is already at the edge of the 80mg/L linear range; if the concentration of zinc acetate is continuously increased to 24mM, the ABS in the linear range of 80mg/L is out of limit, and the requirement that the linear range reaches 80mg/L cannot be met.
Example 4
Verification of lower limit of latex particle diameter
The latex in reagent B in example 3 was replaced with a latex with a particle size of 172nm (product No. CB0172E from Polymicrospheres), and since the overall sensitivity decreased significantly and the low-level sensitivity could not be met by zinc acetate alone, the sensitizer was adjusted to polyethylene glycol PEG8000 at a concentration of 20g/L, and D was obtained by the secondary convergent polymerization latex modification technique described in patent No. 201710986970.74Further, a reagent C was obtained from the latex with 0.78 × 172nm — 134nm, and the main components of the reagent C were:
a first reagent: sodium chloride with concentration of 400 mM/L; polyethylene glycol PEG8000, the concentration is 20 g/L; tween-20 with concentration of 5 ml/L; preservative PC-300 with the concentration of 0.35 ml/L;
and a second reagent: the concentration of the latex particles after secondary convergence polymerization is 3g/L, and the particle diameter D of the latex4Is 134 nm; UTRF antibody (cat # Q0327), at a concentration of 10ml/L, brand Dako, reagent two was prepared using conventional chemical coupling.
In reagent C, zinc acetate was added at different concentrations to reagent C, and in combination with reagent C, 2ul of the sample, 105ul of reagent I, and 35ul of reagent II were used, and in Hitachi7180, the same lot of UTRF standards at different concentrations at a wavelength of 570nm and at a reading point of 18 to 34 was used to measure Δ ABS, precision around the reference value, linear range, and antigen excess range, and the results are shown in tables 10 to 12.
TABLE 10 Effect of different concentrations of Zinc acetate on Δ ABS in different concentrations of UTRF
TABLE 11 influence of different concentrations of Zinc acetate on the precision of the determination of the concentration of the reference values
| Concentration of Zinc acetate | mean(mg/L) | SD | CV |
| 0mM | 1.92 | 0.221 | 11.51％ |
| 6mM | 2.01 | 0.039 | 1.94％ |
| 12mM | 1.99 | 0.038 | 1.91％ |
| 19mM | 2.02 | 0.04 | 1.98％ |
| 24mM | 1.99 | 0.039 | 1.96％ |
TABLE 12 Effect of different concentrations of Zinc acetate on the antigen excess range and Linear Range
As is clear from tables 10 to 12, in the present example, when the secondary convergence polymerization method described in patent No. 201710986970.7 was used to reduce the latex particle size to 172nm and zinc acetate was added to the reagent 1 in an amount of 6 to 19mM, the precision CV of 2mg/L was less than 2%, the linear range was more than 80mg/L, and the antigen excess range satisfied the wide linear range UTRF reagent required at 1000 mg/L. However, it can be seen that when the latex particle size is reduced to 172nm, polyethylene glycol PEG8000 as a conventional sensitizer has been added to a higher concentration, namely 20g/L, which is close to 25g/L, which is likely to cause non-specific reaction of PEG precipitated protein, and then the latex particle size is reduced, so that precision CV is difficult to ensure. In this example, the applicable range of latex particle size is 172-200nm at zinc acetate concentration of 6-19 mM.
In addition, as can be seen from the comparison of examples 3 and 4, the latex particle size increases and the linear range and the antigen excess range are greatly reduced. The latex particle size of 200nm in example 3 is already at the edge of the antigen surplus range at 19mM zinc acetate concentration and 1g/L PEG 6000.
The invention discloses a urine transferrin detection kit with wide detection range, which adds a metal ion additive which can be combined with transferrin and is easy to dissolve in water into a kit reagent I, and selects a proper latex particle size range, so that the improvement of low value sensitivity brought by metal ions and the loss of low value sensitivity caused by using latex with smaller particle size are mutually offset, the improvement of low value sensitivity has little influence on high value sensitivity, under the condition that the antigen surplus range meets 1000mg/L, the upper line of the linear range of the kit is expanded to be more than 80mg/L, and the low value sensitivity can be maintained, thereby ensuring that an ash area sample meets the detection of 2mg/L concentration, and the precision CV is less than 2%, and the kit can meet the detection of more than 90% patients, and the kit of the invention is used for replacing an imported reagent, can greatly save the expense of patients.
The embodiments in the present description are described in a progressive manner, each embodiment focuses on differences from other embodiments, and the same and similar parts among the embodiments are referred to each other. The device disclosed by the embodiment corresponds to the method disclosed by the embodiment, so that the description is simple, and the relevant points can be referred to the method part for description.
The previous description of the disclosed embodiments is provided to enable any person skilled in the art to make or use the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the invention. Thus, the present invention is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.
Claims (3)
1. The urine transferrin detection kit with wide detection range is characterized by comprising a first reagent and a second reagent, wherein the first reagent comprises a metal ion additive; the metal ion additive can be combined with transferrin and is easy to dissolve in water; the metal ion additive comprises zinc salt, and the concentration of the zinc salt is 6-19 mM; the zinc salt is zinc acetate;
the reagent I also comprises 20g/L of polyethylene glycol PEG 8000;
the reagent II is a latex particle solution marked with transferrin antibody; the particle size range of the latex particles is 172-200 nm.
2. The urine transferrin assay kit as claimed in claim 1, wherein the reagent one further comprises sodium chloride or potassium chloride at a concentration of 400 mM/L; and a dispersant at a concentration of 5 ml/L.
3. The urine transferrin assay kit as claimed in claim 2, wherein the dispersant is tween-20.
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