CN113671192A - CRP latex enhanced kit and preparation method and application thereof - Google Patents
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- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
- G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
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Abstract
The invention discloses a CRP latex enhanced kit and a preparation method and application thereof, belonging to the technical field of medical products and technology, the CRP latex enhanced kit comprises a reagent R1 and a reagent R2, wherein the reagent R2 comprises the following components: buffer solution 1, buffer solution 2, latex microspheres 1, latex microspheres 2, EDC, C-reactive protein antibody 1, C-reactive protein antibody 2, stabilizing agent and blocking agent. The uniformly dispersed reagent R2 is obtained by controlling the feeding amount and simplifying the preparation process, the sensitivity of the obtained kit can reach 0.15mg/L, the CRP content under a high concentration level can be accurately measured, and the kit can be used for diagnosing various infectious and inflammatory diseases and the like.
Description
Technical Field
The invention belongs to the field of medical products and technologies, and particularly relates to a CRP latex enhanced kit and a preparation method and application thereof.
Background
C-reactive protein (CRP) is synthesized by hepatocytes and produced in fetal stage, non-maternal placental transfer. When the body is infected or the tissues are damaged, macrophages and other white blood cells are activated, cytokines such as interleukin-6, interleukin-1, tumor necrosis factor (TNF-a) and other mediators are produced, and the cytokines and the mediators reach the liver and stimulate the liver cells and the epithelial cells to synthesize CRP.
The C-reactive protein can be divided into common CRP, hypersensitive CRP and full-scale CRP due to different detection sensitivity performance. The common CRP is mainly used for evaluating infection, tissue injury and inflammatory diseases clinically, provides data support for diagnosis of the inflammatory diseases and provides diagnosis and treatment information for treatment and monitoring of the diseases. The hypersensitive CRP is a sensitive index for diagnosing a low-level inflammatory state, and the level of the hypersensitive CRP in serum is closely related to the occurrence, the severity and the prognosis of cardiovascular and cerebrovascular diseases such as atherosclerosis and Acute Cerebral Infarction (ACI). The full-scale CRP can be used for evaluating disease activity and curative effect monitoring, predicting the probability of cardiovascular diseases of healthy people in the future and evaluating the degree of disease infection.
At the present stage, in the preparation process of R2 in the CRP reagent, the time consumption is long, the efficiency is low, the raw material loss is high, the product is not beneficial to the expanded production, and the purpose of mass production cannot be achieved, free EDC, antibodies and the like need to be removed centrifugally in the manufacturing process, and the latex particles need to be dispersed and resuspended in an ultrasonic mode so as to improve the performance of the R2 reagent; otherwise, if the free EDC and the antibody cannot be removed, the performance of the reagent R2 is directly affected, the sensitivity and accuracy are reduced, and the market demand cannot be met.
Therefore, research on a kit for enhancing the CRP latex by a one-step method, which has a simple and efficient process, and a preparation method and application thereof are needed.
Disclosure of Invention
In order to solve the technical problems, the invention provides a CRP latex enhanced kit and a preparation method and application thereof, in the preparation process of R2 in a CRP reagent, the uniformly dispersed reagent R2 is obtained by controlling the dosage and simplifying the preparation process, the obtained kit can accurately measure the CRP content of a low concentration level below 0.5mg/L, the sensitivity can reach 0.15mg/L, and the CRP content of a high concentration level can also be accurately measured, so that the kit can be used for diagnosing various infectious and inflammatory diseases and the like.
In order to achieve the purpose, the invention provides the following technical scheme:
a CRP latex enhanced kit comprising reagent R1 and reagent R2, said reagent R2 comprising the following concentrations of ingredients: 5-50mmol/L buffer solution 1, 5-50mmol/L buffer solution 2, 0.5-2g/L latex microsphere 1, 0.5-2g/L latex microsphere 2, 0.08-0.25g/L EDC, 0.03-0.08g/L C reactive protein antibody 1, 0.03-0.05g/L C reactive protein antibody 2, 15-40g/L stabilizer, 1.5-5.5g/L blocking agent. EDC is 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride.
Further, the reagent R2 comprises the following components in concentration: 50mmol/L buffer solution 1, 50mmol/L buffer solution 2, 2g/L latex microsphere 1, 1g/L latex microsphere 2, 0.15g/L EDC, 0.08g/L C reactive protein antibody 1, 0.05g/L C reactive protein antibody 2, 40g/L stabilizer, 2.5g/L blocking agent.
Further, the buffer solution 1 is any one of Tris-HCl buffer solution, glycine buffer solution, PBS buffer solution and Heppes buffer solution;
the buffer solution 2 is any one of 2- (N-morpholine) ethanesulfonic acid buffer solution, glycine buffer solution and Heppes buffer solution.
Further, the stabilizer is trehalose or sucrose or glucose; the sealant is bovine serum albumin or casein.
Further, the particle size of the latex microsphere 1 is 80nm, and the solid content is 10%; the latex microspheres 2 have a particle size of 240nm and a solid content of 10%.
Further, the reagent R1 comprises the following components in concentration: 15.0g/L-20.0g/L of Na2HPO4·12H2O, 1.2g-4.8g/L NaH2PO4·2H2O, 5.0-20.0 g/L NaCl, 5.0-15.0 g/L polyethylene glycol 6000, 1-5 g/L Tween 20 and 1ml Proclin 300.
The invention provides a preparation method of the CRP latex reinforced kit, the reagent R1 is prepared according to a conventional method, and the preparation method of the reagent R2 comprises the following steps of:
(1) and (3) activation: taking two containers A and B, then respectively adding 500ml of buffer solution 2, adding latex microspheres 1 and a stabilizer into the container A, adding latex microspheres 2 and a stabilizer into the container B, and stirring for 12-15min in a shaking table at 37 ℃ to mix uniformly; respectively adding EDC, stirring for 30-35min, and mixing to obtain activated latex microspheres;
(2) marking: adding the C reactive protein antibody 1 into the activated latex microspheres in the container A while stirring, adding the C reactive protein antibody 2 into the activated latex microspheres in the container B while stirring, and uniformly mixing in a shaking table at 37 ℃ for 60-120min to obtain marked latex microspheres;
(3) and (3) sealing: respectively adding a sealing agent into the marked latex microspheres in the two containers, fully dissolving and mixing for 40-60min, and sealing to obtain sealed latex microspheres;
(4) and respectively adding 500ml of buffer solution 1 into the closed latex microspheres in the two containers, fully and uniformly mixing, and then mixing the substances in the container A and the container B to obtain the reagent R2.
Further, the rotation speed of the stirring is 180-200 rpm.
The invention provides application of the CRP latex enhanced kit, and the kit is used for detecting CRP content.
Further, the CRP detection concentration range of the kit is 0.15mg/L-320 mg/L.
The invention has the following beneficial effects:
1. in the preparation process of the reagent R2 of the CRP latex enhanced kit, the feeding amount is controlled, the preparation process is simplified, FIG. 1 is a flow chart of the preparation process of the reagent R2, as shown in FIG. 1, in the invention, a coupling agent EDC is added into latex microspheres for activation, the latex microspheres become unstable intermediates, at the moment, the activated latex microspheres are added with an antibody label, and the antibody is immobilized on the latex microspheres to finally form a reagent R2; the preparation process has short time consumption and low raw material loss, and improves the production efficiency.
2. The CRP latex enhanced kit has high sensitivity and accuracy, can accurately measure the CRP content of a low concentration level below 0.5mg/L, meets the standard of a full-range CRP linear range of 0.5-80mg/L in YY/T1513-2017C reactive protein determination kit, has the sensitivity of 0.15mg/L, can also accurately measure the CRP content of a high concentration level of 320mg/L, improves the detection range and sensitivity, and can be used for diagnosing various infectious and inflammatory diseases and the like.
3. According to the CRP latex enhanced kit, the latex microspheres with different particle sizes are controlled to be combined with different types of C-reactive protein antibodies, so that the large-scale detection is realized, the highest detection concentration is 320mg/L, the detection sensitivity is improved to be 0.15mg/L, the kit has the characteristics of flexibility and universality, and can be used for the detection of CRP in serum, plasma and whole blood; by adjusting the proportion of the CRP latex microspheres to the antibody, the requirements of immunity transmission and scattering detection can be met, and reliable raw materials are provided for kit production.
4. On the basis of simplifying the process for preparing the reagent R2, the CRP latex enhanced kit reduces the feeding amount of antibodies and latex microspheres, effectively reduces the probability of R2 aggregation, increases the stability and sensitivity of the reagent R2, and greatly reduces the cost of the reagent.
5. According to the CRP latex enhanced kit, a certain amount of surfactant Tween 20 is added into R1, so that the interference of the reagent in whole blood detection is effectively reduced, and the detection accuracy is improved.
Drawings
FIG. 1 is a flow chart of the preparation process of the reagent R2 according to the invention.
FIG. 2 is a graph of linear range validation data for inventive example 1 and comparative example.
FIG. 3 is a graph of LOQ validation data for inventive example 1 and comparative example.
Detailed Description
The following examples may assist those skilled in the art in a more complete understanding of the present invention, but are not intended to limit the invention in any way. Raw materials of a buffering agent, a stabilizing agent, a sealing agent, latex microspheres, a C-reactive protein antibody 1 (namely a C-reactive protein antibody 101) and a C-reactive protein antibody 2 (namely a C-reactive protein antibody 102) used in the invention are purchased from chemical and biological raw material companies, and the latex microspheres have the particle size of 80nm and the solid content of 10 percent; the latex microsphere 2 has the particle size of 240nm and the solid content of 10 percent and can be directly used.
Example 1
A kit for CRP latex enhancement comprising reagent R1 and reagent R2,
the reagent R1 comprises the following components in concentration: 15.0g/L of Na2HPO4·12H2O, 1.2g of NaH2PO4·2H2O, 5.0g/L NaCl, 5.0g/L polyethylene glycol 6000, 1g/L Tween 20 and 1ml Proclin 300; the reagent R1 is prepared according to a conventional method;
the reagent R2 comprises the following components in concentration: 50mmol/L Tris-HCl buffer solution, 50 mmol/L2- (N-morpholine) ethanesulfonic acid buffer solution, 2g/L latex microspheres (80nm), 1g/L latex microspheres (240nm), 0.15g/L EDC, 0.08g/L C reactive protein antibody 1, 0.05g/L C reactive protein antibody 2, 40g/L trehalose, 2.5g/L bovine serum albumin;
preparation method of every two liters of the reagent R2: the rotating speed of the stirring is 200 rpm;
(1) and (3) activation: taking a container A and a container B, adding 500ml of 2- (N-morpholine) ethanesulfonic acid buffer solution, adding latex microspheres (80nm) and trehalose into the container A, adding latex microspheres (240nm) and trehalose into the container B, and stirring for 15min in a shaking table at 37 ℃ to uniformly mix; respectively adding EDC, stirring for 35min, and mixing to obtain activated latex microspheres;
(2) marking: adding the C-reactive protein antibody 1 into the activated latex microspheres in the container A while stirring, adding the C-reactive protein antibody 2 into the activated latex microspheres in the container B while stirring, and uniformly mixing in a shaker at 37 ℃ for 60min to obtain marked latex microspheres;
(3) and (3) sealing: adding bovine serum albumin into the marked latex microspheres in the two containers, fully dissolving and mixing for 40min, and sealing to obtain sealed latex microspheres;
(4) and respectively adding 500ml of Tris-HCl buffer solution into the closed latex microspheres, fully and uniformly mixing, and mixing the substances in the container A and the container B to obtain the reagent R2.
Comparative example
A CRP kit comprising reagent R1 and reagent R2 wherein R1 is in accordance with example 1; the R2 materials are shown in Table 1 below;
TABLE 1
The formulation of the reagent R2 per liter was as follows:
(1) and (3) activation: adding 500ml of 2- (N-morpholine) ethanesulfonic acid buffer solution into two containers A and B, adding latex microspheres (80nm) and trehalose into the container A, adding latex microspheres (240nm) and trehalose into the container B, and uniformly stirring at 200rpm in a shaking table at 37 ℃ for 15 min; adding EDC respectively, and stirring and mixing uniformly at 200rpm for 35 min;
(2) centrifuging: placing the activated latex microspheres in a centrifuge for centrifugation at the rotation speed of 18000rpm for 1.5 hours, removing the supernatant after the centrifugation is finished, and leaving the bottom sediment;
(3) ultrasonic: adding 500ml of 2- (N-morpholine) ethanesulfonic acid buffer solution into the centrifuged precipitate, and placing the precipitate in an ultrasonic crusher for ultrasonic resuspension; setting parameters: a horn phi 6; frequency: 20 HZ; time: 20 min; clearance: 30 s;
(4) marking: stirring and adding the C-reactive protein antibody into the resuspended materials, and uniformly stirring and mixing for 120min at 200rpm in a shaker at 37 ℃;
(5) and (3) sealing: adding bovine serum albumin into the marked material, fully dissolving and mixing, and sealing for 60 min;
(6) centrifuging: placing the completely sealed latex microspheres in a centrifuge for centrifugation at the rotation speed of 18000rpm for 1.5 hours, removing the supernatant after the centrifugation is finished, and leaving the bottom precipitate;
(7) ultrasonic: adding 500ml Tris-HCl buffer solution (50mmol/L, pH7.5) into the centrifuged precipitate, and placing the mixture in an ultrasonic crusher for ultrasonic resuspension; setting parameters: a horn phi 6; frequency: 20 HZ; time: 20 min; clearance: 30 s;
(8) and adding 500ml of Tris-HCl buffer solution (50mmol/L, pH7.5) into the completely resuspended latex microspheres, and fully and uniformly mixing.
Application example 1
The kit comprises: the kit of example 1, the kit of the comparative example.
An experimental instrument: hitachi 7180 full-automatic biochemical analyzer.
Test samples: obtained by dilution of high concentration samples with low concentration samples in a multiple ratio, the formulation scheme is shown in table 2 below. The concentration and volume unit of each sample are unified, and a liquid transfer device with good precision and accuracy is selected when liquid is sucked; the sample is mixed thoroughly during preparation and evaporation or other conditions that degrade the sample are avoided.
TABLE 2
| Sample number | 1 | 2 | 3 | 4 | 5 |
| Low concentration serum (mL, concentration 0.15mg/L) | 1.00 | 0.75 | 0.50 | 0.25 | 0.00 |
| High concentration serum (mL, concentration 320.0mg/L) | 0.00 | 0.25 | 0.50 | 0.75 | 1.00 |
The experimental scheme is as follows: the kit was subjected to linear range analysis and LOQ validation analysis and the data was recorded in a chart. Wherein, table 3 shows the results of the linear range detection of the comparative example kit and the kit of example 1 of the present invention. Table 4 shows the results of the calibration tests of the comparative example kit and the kit of example 1 of the present invention. Table 5 shows the results of LOQ-validated analysis of the kit of example 1 of the present invention. Table 3 is the LOQ validation analysis of the comparative example kit. FIG. 2 is a graph of linear range validation data for inventive example 1 and comparative example. FIG. 3 is a graph of LOQ validation data for inventive example 1 and comparative example.
TABLE 3
TABLE 4
From the linear range verification data in fig. 2, tables 3 and 4, it can be seen that the comparative example and the example 1 of the present invention can satisfy the performance requirements of the reagent when testing the linear range of the CRP kit; when the reagent is calibrated, the background value of the reagent is lower than that of the comparative example when the concentration of 0 is tested in the example 1 of the invention, which shows that the technology of the invention can better control the aggregation of the latex microspheres and has better stability when the reagent R2 is prepared.
TABLE 5
TABLE 6
As can be seen from the LOQ verification data in FIG. 3, tables 5 and 6, when the sensitivity of the CRP kit is tested by comparing the comparative example with the example 1 of the invention, the LOQ of the example 1 of the invention can reach 0.15mg/L, and the LOQ of the comparative example is 0.5mg/L, which means that the technology of the invention has higher sensitivity.
Example 2
A kit for CRP latex enhancement comprising reagent R1 and reagent R2,
the reagent R1 comprises the following components in concentration: 20.0g/L of Na2HPO4·12H2O, 4.8g/L NaH2PO4·2H2O, 20.0g/L NaCl, 15.0g/L polyethylene glycol 6000, 5g/L Tween 20 and 1ml Proclin 300; the reagent R1 is prepared according to a conventional method;
the reagent R2 comprises the following components in concentration: 5mmol/L Heppes buffer solution, 5 mmol/L2- (N-morpholine) ethanesulfonic acid buffer solution, 1g/L latex microspheres (80nm), 0.5g/L latex microspheres (240nm), 0.08g/L EDC, 0.03g/L C-reactive protein antibody 1, 0.05g/L C reactive protein antibody 2, 15g/L trehalose, 1.5g/L bovine serum albumin;
preparation method of every two liters of the reagent R2: the rotating speed of the stirring is 200 rpm;
(1) and (3) activation: taking a container A and a container B, adding 500ml of 2- (N-morpholine) ethanesulfonic acid buffer solution, adding latex microspheres (80nm) and trehalose into the container A, adding latex microspheres (240nm) and trehalose into the container B, and stirring for 12min in a shaking table at 37 ℃ to uniformly mix; respectively adding EDC, stirring for 30min, and mixing to obtain activated latex microspheres;
(2) marking: adding the C-reactive protein antibody 1 into the activated latex microspheres in the container A while stirring, adding the C-reactive protein antibody 2 into the activated latex microspheres in the container B while stirring, and uniformly mixing in a shaker at 37 ℃ for 120min to obtain marked latex microspheres;
(3) and (3) sealing: adding bovine serum albumin into the marked latex microspheres in the two containers, fully dissolving and mixing for 60min, and sealing to obtain sealed latex microspheres;
(4) and respectively adding 500ml of Heppes buffer solution into the closed latex microspheres, fully and uniformly mixing, and mixing the A container and the B container together to obtain the reagent R2.
Example 3
A kit for CRP latex enhancement comprising reagent R1 and reagent R2,
the reagent R1 comprises the following components in concentration: 16.0g/L of Na2HPO4·12H2O, 3.2g/L NaH2PO4·2H2O, 15.0g/L NaCl, 8.0g/L polyethylene glycol 6000, 3g/L Tween 20 and 1ml Proclin 300; the reagent R1 is prepared according to a conventional method;
the reagent R2 comprises the following components in concentration: 25mmol/L PBS buffer solution, 25mmol/L glycine buffer solution, 1.2g/L latex microspheres (80nm), 0.7g/L latex microspheres (240nm), 0.25g/L EDC, 0.08g/L C reactive protein antibody 1, 0.08g/L C reactive protein antibody 2, 20g/L sucrose, 3g/L bovine serum albumin;
preparation method of every two liters of the reagent R2: the rotating speed of the stirring is 200 rpm;
(1) and (3) activation: adding 500ml of glycine buffer solution into two containers A and B, adding latex microspheres (80nm) and sucrose into the container A, adding latex microspheres (240nm) and sucrose into the container B, and stirring for 13min in a shaking table at 37 ℃ to mix uniformly; respectively adding EDC, stirring for 34min, and mixing to obtain activated latex microspheres;
(2) marking: adding the C-reactive protein antibody 1 into the activated latex microspheres in the container A while stirring, adding the C-reactive protein antibody 2 into the activated latex microspheres in the container B while stirring, and uniformly mixing in a shaker at 37 ℃ for 70min to obtain marked latex microspheres;
(3) and (3) sealing: adding bovine serum albumin into the marked latex microspheres in the two containers, fully dissolving and mixing for 45min, and sealing to obtain sealed latex microspheres;
(4) and respectively adding 500ml of PBS buffer solution into the closed latex microspheres, fully and uniformly mixing, and mixing the two containers A and B together to obtain the reagent R2.
Example 4
A kit for CRP latex enhancement comprising reagent R1 and reagent R2,
the reagent R1 comprises the following components in concentration: 18.0g/L of Na2HPO4·12H2O, 2.8g/L NaH2PO4·2H2O, 10.0g/L NaCl, 10.0g/L polyethylene glycol 6000, 2g/L Tween 20 and 1ml Proclin 300; the reagent R1 is prepared according to a conventional method;
the reagent R2 comprises the following components in concentration: 40mmol/L glycine buffer solution, 40mmol/L Heppes buffer solution, 1.8g/L latex microspheres (80nm), 1.5g/L latex microspheres (240nm), 0.2g/L EDC, 0.07g/L C reactive protein antibody 1, 0.09g/L C reactive protein antibody 2, 30g/L glucose, 5.5g/L casein;
preparation method of every two liters of the reagent R2: the rotating speed of the stirring is 200 rpm;
(1) and (3) activation: adding 500ml of Heppes buffer solution into two containers A and B, adding latex microspheres (80nm) and glucose into the container A, adding latex microspheres (240nm) and glucose into the container B, and stirring for 14min in a shaker at 37 ℃ to mix uniformly; respectively adding EDC, stirring for 32min, and mixing to obtain activated latex microspheres;
(2) marking: adding the C-reactive protein antibody 1 into the activated latex microspheres in the container A while stirring, adding the C-reactive protein antibody 2 into the activated latex microspheres in the container B while stirring, and uniformly mixing in a shaker at 37 ℃ for 80min to obtain marked latex microspheres;
(3) and (3) sealing: adding casein into the marked latex microspheres in the two containers, fully dissolving and mixing for 50min, and sealing to obtain sealed latex microspheres;
(4) and respectively adding 500ml of glycine buffer solution into the closed latex microspheres, fully and uniformly mixing, and mixing the two containers A and B together to obtain the reagent R2.
The data obtained by performing the tests as in example 1 on the kits obtained in examples 2-4 are similar to those of example 1, which indicates that the kits obtained by the method of the present invention have good reproducibility.
Although the invention has been described in detail hereinabove with respect to a general description and specific embodiments thereof, it will be apparent to those skilled in the art that modifications or improvements may be made thereto based on the invention. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.
Claims (10)
1. A CRP latex-reinforced kit comprising reagent R1 and reagent R2, characterized in that the reagent R2 comprises the following ingredients in the concentrations: 5-50mmol/L buffer solution 1, 5-50mmol/L buffer solution 2, 0.5-2g/L latex microsphere 1, 0.5-2g/L latex microsphere 2, 0.08-0.25g/L EDC, 0.03-0.08g/L C reactive protein antibody 1, 0.03-0.05g/L C reactive protein antibody 2, 15-40g/L stabilizer, 1.5-5.5g/L blocking agent.
2. The CRP latex enhanced kit according to claim 1, characterized in that the reagent R2 comprises the following ingredients in the following concentrations: 50mmol/L buffer solution 1, 50mmol/L buffer solution 2, 2g/L latex microsphere 1, 1g/L latex microsphere 2, 0.15g/L EDC, 0.08g/L C reactive protein antibody 1, 0.05g/L C reactive protein antibody 2, 40g/L stabilizer, 2.5g/L blocking agent.
3. The CRP latex enhanced kit according to claim 1 or 2, wherein the buffer 1 is any one of Tris-HCl buffer, glycine buffer, PBS buffer, Heppes buffer;
the buffer solution 2 is any one of 2- (N-morpholine) ethanesulfonic acid buffer solution, glycine buffer solution and Heppes buffer solution.
4. The CRP latex-enhanced kit of claim 1 or 2, wherein the stabilizer is trehalose or sucrose or glucose; the sealant is bovine serum albumin or casein.
5. The CRP latex reinforced kit of claim 1, wherein the latex microspheres 1 have a particle size of 80nm and a solids content of 10%; the latex microspheres 2 have a particle size of 240nm and a solid content of 10%.
6. The CRP latex enhanced kit according to claim 1, characterized in that the reagent R1 comprises the following ingredients in the following concentrations: 15.0g/L-20.0g/L of Na2HPO4·12H2O, 1.2g-4.8g/L NaH2PO4·2H2O, 5.0-20.0 g/L NaCl, 5.0-15.0 g/L polyethylene glycol 6000, 1-5 g/L Tween 20 and 1ml Proclin 300.
7. A method for the preparation of a kit for the enhancement of CRP latex according to any one of claims 1 to 6, wherein said reagent R1 is formulated according to the conventional method, characterized in that the method for the preparation of R2 per two liters comprises the following steps:
(1) and (3) activation: taking two containers A and B, then respectively adding 500ml of buffer solution 2, adding latex microspheres 1 and a stabilizer into the container A, adding latex microspheres 2 and a stabilizer into the container B, and stirring for 12-15min in a shaking table at 37 ℃ to mix uniformly; respectively adding EDC, stirring for 30-35min, and mixing to obtain activated latex microspheres;
(2) marking: adding the C reactive protein antibody 1 into the activated latex microspheres in the container A while stirring, adding the C reactive protein antibody 2 into the activated latex microspheres in the container B while stirring, and uniformly mixing in a shaking table at 37 ℃ for 60-120min to obtain marked latex microspheres;
(3) and (3) sealing: respectively adding a sealing agent into the marked latex microspheres in the two containers, fully dissolving and mixing for 40-60min, and sealing to obtain sealed latex microspheres;
(4) and respectively adding 500ml of buffer solution 1 into the closed latex microspheres in the two containers, fully and uniformly mixing, and then mixing the substances in the container A and the container B to obtain the reagent R2.
8. The method for preparing the CRP latex-reinforced kit according to claim 7, wherein the stirring speed is 180-200 rpm.
9. Use of a kit for the CRP latex enhancement according to any one of claims 1 to 6 for the detection of the CRP content.
10. The use of the CRP latex-enhanced kit of claim 9, wherein the kit has a CRP detection concentration ranging from 0.15mg/L to 320 mg/L.
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| CN112763725A (en) * | 2020-12-29 | 2021-05-07 | 中生北控生物科技股份有限公司 | Preparation and application of detection reagent for determining reverse protein C by latex turbidimetry |
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| CN106324239A (en) * | 2016-07-28 | 2017-01-11 | 武汉景川诊断技术股份有限公司 | preparation method and application of C-reactive protein immuno latex microsphere |
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