A low-density lipoprotein LDL-C assay kit with reduced cross contamination

Abstract

The present invention discloses a low-density lipoprotein LDL-C detection kit for reducing cross contamination, which includes reagent R1 and reagent R2, wherein the reagent R1 includes a buffer, cholesterol oxidase, cholesterol esterase, 4-aminoantipyrine, surfactant 1 and surfactant 2; the reagent R2 includes a buffer, peroxidase, TOOS, surfactant 3, surfactant 4 and protein. The present invention is by adding surfactant 1 and surfactant 2 to reagent R1, and adding surfactant 3, surfactant 4 and protein in reagent R2. On the one hand, the detection performance of the kit is significantly improved, including indicators such as detection precision and linear range, and on the other hand, the introduction of bile salt analogs in the reagent is avoided, and the cross contamination between different detection items is reduced to the greatest extent, ensuring the detection accuracy of subsequent detection items.

Description

Technical Field

[0001] The invention relates to the field of medical testing, and in particular to a low-density lipoprotein (LDL-C) determination kit capable of reducing cross contamination. Background Art

[0002] At present, biochemical test kits and fully automatic biochemical analyzers have been widely used in the laboratories of hospitals at all levels. However, when conducting sample tests, the accuracy and reliability of the test results are affected by cross contamination, which is increasingly valued by biochemical test personnel. There are two main sources of cross contamination in tests: contamination of reagent needles, stirring rods and colorimetric cups; contamination between reagents. Common types include: ① Direct contamination of reagent components: the reagent of the previous test contains the substance to be tested in the next test; ② Reagent components participate in the reaction: the components contained in the previous reagent will react with a certain component of the next test reagent; ③ The same reaction process: the reaction guided by the previous reagent indirectly interferes with the reaction process of the next project, and the next project measures the sum of the reactions of the previous and next projects; ④ Influence on reaction conditions: The previous test affects the reaction conditions of the next project, such as pH, and thus changes the reaction rate.

[0003] In order to eliminate the cross-interference in the inspection process, on the one hand, we must continuously improve the professional level and operational skills of inspectors, and on the other hand, we must start from the reagent research and development itself, optimize the reagent composition, and replace components that have direct or indirect contamination to other projects. Bile salt substances are a class of anionic steroidal biosurfactants. Due to their special amphiphilic skeleton structure, unique physicochemical properties, good biocompatibility and environmental friendliness, they play the role of solubilizing, dissociating protein-encapsulated substances and oxidizing agents in biochemical reagents. They are excellent stabilizers and dissociating agents. For example, most biochemical detection reagents such as total cholesterol (CHOL), triglycerides (TG), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), lipase (LPS) and lipoprotein a will add bile salt analogs. However, at the same time, since bile salts are zwitterionic substances that are extremely difficult to elute, if bile salts or their analogs are added to the test kit, they are very likely to remain on the test cup and stirring rod, which will cause serious interference with the subsequent TBA (total bile acid) test results, resulting in the appearance of false positive results, which will cause serious over-medicalization and bring unnecessary economic losses to patients, or even cause medical accidents due to misprescription of clinical drugs. Summary of the invention

[0004] Based on the above requirements, the inventors unexpectedly discovered that by adding surfactant 1 and surfactant 2 to reagent R1, and adding surfactant 3, surfactant 4 and a protein to reagent R2, on the one hand, the detection performance of the kit (including precision and linear range, etc.) can be guaranteed, and on the other hand, the introduction of bile salt analogs into the reagent is avoided, thereby minimizing cross contamination between different detection items.

[0005] The present invention provides a low-density lipoprotein (LDL-C) determination kit capable of reducing cross contamination, and adopts the following technical means: a low-density lipoprotein (LDL-C) detection kit capable of reducing cross contamination, comprising a reagent R1 and a reagent R2, characterized in that the reagent R1 comprises a buffer, cholesterol oxidase, cholesterol esterase, 4-aminoantipyrine, a surfactant 1 and a surfactant 2; the reagent R2 comprises a buffer, a peroxidase, TOOS, a surfactant 3, a surfactant 4 and a protein.

[0006] Preferably, the surfactant 1 and the surfactant 2 are selected from two of Triton X-100, A90, Tween-80, B-66 or LS-114, preferably a combination of B-66 and LS-114; the surfactant 3 and the surfactant 4 are selected from two of Triton X-100, A90, Tween-20, Triton305 or Brij 58, preferably a combination of Triton305 and Brij 58; the protein is selected from at least one of casein, BSA and hemocyanin, preferably hemocyanin.

[0007] Preferably, the content of surfactant 1 in the reagent R1 is 2-10 g / L, and the content of surfactant 2 is 2-10 g / L; the content of surfactant 3 in the reagent R2 is 1-3 g / L, the content of surfactant 4 is 1-3 g / L, and the content of hemocyanin is 2-10 g / L;

[0008] Preferably, the reagent R1 further comprises a preservative and an accelerator; and the reagent R2 further comprises a preservative and ascorbic acid oxidase.

[0009] Preferably, the reagent R1 contains 30-100 mmol / L buffer, 300-600 U / L cholesterol oxidase, 800-1200 U / L cholesterol esterase, 10-20 mmol / L 4-aminoantipyrine, 0.5-1 g / L preservative and 10-20 g / L accelerator.

[0010] Preferably, the reagent R2 comprises 30-80 mmol / L of buffer, 1500-2000 U / L of peroxidase, 0.7-2 mmol / L of TOOS, 1-2 g / L of preservative and 2500-3500 U / L of ascorbic acid oxidase.

[0011] Preferably, the buffer in the reagent R1 is selected from at least one of Tris buffer, phosphate buffer and GOOD'S buffer, the preservative is selected from at least one of Proclin-300, sodium azide or gentamicin; the accelerator is selected from at least one of dextran, PEG6000 and PEG8000.

[0012] Preferably, the buffer in the reagent R2 is selected from at least one of Tris buffer, phosphate buffer and GOOD'S buffer, the preservative is selected from at least one of Proclin-300, sodium azide or gentamicin; the accelerator is selected from at least one of dextran, PEG6000 and PEG8000.

[0013] Preferably, the pH value of the reagent R1 is 5.5-6.5, preferably 6.0, and the pH value of the reagent R2 is 7.0-8.0, preferably 7.5.

[0014] A method for reducing cross contamination between biochemical reagents, characterized in that surfactant 1 and surfactant 2 are added to reagent R1, and surfactant 3, surfactant 4 and protein are added to reagent R2, wherein the surfactant 1 and surfactant 2 are selected from two of Triton X-100, A90, Tween-80, B-66 or LS-114, the surfactant 3 and surfactant 4 are selected from two of Triton X-100, A90, Tween-20, Triton305 or Brij 58, and the protein is selected from at least one of hemocyanin, casein or BSA, preferably hemocyanin.

[0015] The beneficial effect of the present invention lies in that: by adding surfactant 1 and surfactant 2 to reagent R1, and adding surfactant 3, surfactant 4 and a protein to reagent R2, for example, surfactant 1 is B-66, surfactant 2 is LS-114, surfactant 3 is Brij58, surfactant 4 is Triton305, and the protein is hemocyanin, on the one hand, the detection performance of the kit is significantly improved, including indicators such as detection precision and linear range, and on the other hand, the introduction of bile salt analogs into the reagent is avoided, cross contamination between different detection items is reduced to the greatest extent, and the accuracy of subsequent detection is ensured. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 A linear relationship diagram of the theoretical concentration of the low-density lipoprotein (LDL-C) assigned sample provided in Example 2 of the present invention and the measured value of the kit of group A;

[0017] Figure 2 A linear relationship diagram of the theoretical concentration of the low-density lipoprotein (LDL-C) assigned sample provided in Example 2 of the present invention and the measured value of the kit of group B;

[0018] Figure 3 A linear relationship diagram of the theoretical concentration of the low-density lipoprotein (LDL-C) assigned sample provided in Example 2 of the present invention and the measured value of the C group test kit;

[0019] Figure 4 A correlation diagram between the low-density lipoprotein (LDL-C) measurement value and the clinical value provided in Example 2 of the present invention; Specific embodiments

[0020] The following examples are included herein to more clearly and specifically illustrate the technical solutions of the present invention in an illustrative manner. It should be understood by those skilled in the art based on the disclosure herein that many changes can be made in the disclosed specific embodiments but still similar or similar results can be obtained without departing from the idea and scope of the present invention. The specific embodiments of the present invention are only used to explain the present invention and are not intended to limit the present invention in any way.

[0021] Example 1 Preparation of low-density lipoprotein (LDL-C) detection kit

[0022] The LDL-C detection kit of the present invention comprises two liquid components, namely, a reagent R1 and a reagent R2, which are independent of each other.

[0023] 1. Preparation of Reagent R1

[0024] Prepare according to the following formula, stir thoroughly and store at 2-8℃.

[0025] Reagent R1:

[0026]

[0027] 2. Preparation of Reagent R2

[0028] Prepare according to the following formula, stir thoroughly and store at 2-8℃.

[0029] Reagent R2

[0030]

[0031]

[0032] 3. How to use the kit

[0033] In this embodiment, a fully automatic biochemical analyzer (Hitachi 7180) is used in conjunction with the kit of the present invention to perform sample testing.

[0034] (1) Instrument parameter setting

[0035]

[0036] (2) Measurement scheme

[0037]

[0038] (3) Calculation method

[0039] Using multi-point nonlinear / semi-logarithmic calibration mode, with spline function as calculation mode, according to the calibration

[0040]

[0041]

[0042] A dose / response curve is drawn by comparing the absorbance change value with the absorbance value, and the content of low-density lipoprotein (LDL-C) in the sample can be calculated on the dose / response curve according to its absorbance change value.

[0043] The detection principle of the present invention is to use an enzymatic method to determine the content of low-density lipoprotein (LDL-C) in human serum.

[0044] ① The surfactant in reagent R1 inhibits the low-density lipoprotein in serum, while the high-density lipoprotein and very low-density lipoprotein are consumed by the reaction of cholesterol enzymes.

[0045]

[0046]

[0047]

[0048] ② After the surfactant in reagent R2 releases low-density lipoprotein, only LDL-C is measured.

[0049] The absorbance value of the quinone dye is proportional to the cholesterol content. By measuring the change in absorbance at 546nm, the LDL-C content in the sample can be calculated.

[0050] Example 2 Performance Test of LDL Detection Kit

[0051] In order to verify the various properties of the kit of the present invention, two sets of kits were set up for performance verification:

[0052] Group A: the kit prepared in Example 1 of the present invention;

[0053] Group B: Reagent R1: cholesterol esterase 1000U / L, cholesterol oxidase 1000U / L, catalase 30000U / L, MOPS buffer pH7.035mmol / L, NaOH 0.5g / L, dextran sulfate 2.0g / L, sodium azide 0.5g / L, magnesium chloride 2.0g / L, 4-aminoantipyrine 0.12g / L, CHAPS 4.0g / L, BSA 1.5g / L, Triton X-100 1500mg / L; Reagent R2: peroxidase 3000U / L, MOPS buffer pH7.0 35mmol / L, NaOH 0.5g / L, EDTA 0.2g / L, BSA 1.5g / L, sodium azide 0.5g / L, TOOS 1.8g / L, Triton X-100 1500mg / L, sodium dodecyl sulfate 2000mg / L;

[0054] Detection method: endpoint method, determination points: 16, 34; detection wavelength: 570nm (primary) / 700nm (secondary); reaction temperature: 37℃; sample volume: 3.0μl; reagent R1: 210μl; reagent R2: 70μl; reaction direction: ascending; calibration mode: linear mode, 2-point calibration; unit: mmol / L. The instrument automatically performs the following operations: first add 3μl of the serum sample to be tested into the cuvette, then add 1210μl of the reagent, mix well, keep warm for 5 minutes in a constant temperature environment of 37℃, and read the absorbance A1; then add reagent 2 to start the second enzyme reaction, continue to keep warm, and read the absorbance A2 when the reaction reaches the endpoint. Calculation formula: LDL-C (mmol / L) = (ΔA determination / ΔA calibration) × Cs (LDL-C concentration of calibration solution), where ΔA determination = (A2-A1) of sample determination, ΔA calibration = (A2-A1) of calibration substance determination;

[0055] Group C: Reagent R1: sodium cholate 100.0mmol / L, cholesterol esterase 2.0KU / L, cholesterol oxidase 5.0KU / L, peroxidase 15.0KU / L, 4-aminoantipyrine 0.6mmol / L, magnesium chloride 10.0mmol / L and 3-morpholinepropanesulfonic acid buffer 20.0mmol / L with a pH value of 7.0; Reagent R2 includes: Triton X-100 100.0mmol / L, 2-hydroxy-3-m-toluidine propanesulfonic acid sodium 4.0mmol / L, magnesium chloride 10.0mmol / L, 3-morpholinepropanesulfonic acid buffer 20.0mmol / L with a pH value of 7.0, melezitose 1.0g / L and lactulose 0.5g / L;

[0056] Detection method: endpoint method, determination points: 16, 34; detection wavelength: 546nm (primary) / 660nm (secondary); reaction temperature: 37℃; sample volume: 3.0μl; reagent R1: 300μl; reagent R2: 100μl; reaction direction: ascending; calibration mode: linear mode, 2-point calibration; unit: mmol / L. The instrument automatically performs the following operations: first add 3μl of the serum sample to be tested into the cuvette, then add 1300μl of the reagent, mix well, keep warm for 5 minutes in a constant temperature environment of 37℃, and read the absorbance A1; then add reagent 2 to start the second enzyme reaction, continue to keep warm, and read the absorbance A2 when the reaction reaches the endpoint. Calculation formula: LDL-C (mmol / L) = (ΔA determination / ΔA calibration) × Cs (LDL-C concentration of calibration solution), where ΔA determination = (A2-A1) of sample determination, ΔA calibration = (A2-A1) of calibration substance determination;

[0057] (1) Accuracy verification

[0058] Three sets of kits were used to test the accuracy of the Beckman LDL test kit (enzymatic method) assignment samples, with 2 replicates, and the signals were read by a fully automatic biochemical analyzer (Hitachi 7180), and the relative deviation between the measured mean and the target value was calculated to verify the accuracy. The test results are shown in the following table:

[0059] Table 1 Accuracy verification

[0060]

[0061] From the above experimental results, it can be seen that the relative deviations of the test value 1 and the target value 1 of the three kits are -0.68%, -8.56% and -2.93% respectively, and the relative deviations of the test value 2 and the target value 2 are 0.75%, -1.69% and -3.20% respectively. Among them, the detection accuracy of the kit prepared in Example 1 of the present invention (Group A) is better than that of the comparative kit-1 (Group B) and the comparative kit-2 (Group C).

[0062] (2) Precision verification

[0063] Clinical LDL-C low-value samples, median-value samples and high-value samples were selected, and the samples were tested using two sets of kits. Each sample was tested 10 times, and the signal was read by a fully automatic biochemical analyzer (Hitachi 7180). The mean and standard deviation of the measurement were calculated, and the coefficient of variation was calculated for precision inspection. The test results are shown in the following table:

[0064] Table 2 Precision verification

[0065]

[0066] From the above experimental results, it can be seen that the coefficients of variation of the three groups of kits in the detection of low-value samples are 0.92%, 2.43% and 2.47% respectively, the coefficients of variation in the detection of median samples are 0.88%, 3.90% and 1.02% respectively, and the coefficients of variation in the detection of high-value samples are 0.52%, 3.51% and 1.30% respectively. The experimental results show that in the detection of low-value, median and high-value samples, the CV values ​​of the three groups of kits are all less than 10%, and the precision is good, and the precision of group A and group C kits is higher than that of group B.

[0067] (3) Linear range verification

[0068] The Beckman LDL-C test kit (enzymatic method) was selected to assign values ​​to the ultra-high value samples in the fully automatic biochemical analyzer (Hitachi 7180), where the theoretical concentration value of the high value sample was 9.56 mg / L, and the theoretical concentration value of the low value sample was 0.16 mg / L. Then, the high value sample and the low value sample were used to configure each concentration gradient sample in proportion, and the samples were tested using three sets of test kits. Each sample was repeated twice, and the signal was read by the fully automatic biochemical analyzer (Hitachi 7180), and the mean value was calculated for linear range inspection. The test results are shown in the following table:

[0069] Table 3 Linear range verification

[0070]

[0071] From the above experimental results, it can be seen that the relative deviations between the test values ​​and the theoretical values ​​of the kit (Group A) prepared in Example 1 of the present invention are small (less than 3%) when the sample concentration is in the range of 0.10-17.26 mg / L, while the relative deviations between the test values ​​and the theoretical values ​​of the control kit-1 (Group B) and the control kit-2 (Group C) are greater than 20% when the sample concentration is 17.26 mg / L. At the same time, the correlation analysis between the test results of the three groups of kits and the theoretical values ​​of the sample concentrations is performed (see Appendix Figure 1-3 The correlation between the test values ​​and the theoretical values ​​of group A and group C was significantly better than that of group B, and the correlation between the test values ​​and the theoretical values ​​of group A was R 2 is 0.9999, Group B R 2 is 0.9491, and R 2 The experimental results show that when an appropriate amount of bile salt analogs or the composition of the present invention is added to the low-density lipoprotein (LDL-C) detection reagent, the detection performance of the kit can be significantly optimized.

[0072] (4) Clinical evaluation

[0073] 100 clinical samples were collected and tested by the kit of Example 1 of the present invention and the Beckman LDL-C test kit (enzymatic method) for comparison. The test results of the two methods were subjected to linear analysis. The results are as follows: Figure 4 As shown, its R 2 =0.9992, therefore, the low-density lipoprotein (LDL-C) detection kit of the present invention has a good correlation with the Beckman LDL-C detection kit, meets the clinical analysis requirements, and is suitable for clinical detection.

[0074] (5) Kit residue test

[0075] In order to verify the interference of bile salt substances on TBA detection items, a total of 3 groups of experiments were set up for verification:

[0076] ① TBA detection reagent (Chongqing Zhongyuan Huiji Biotechnology Co., Ltd., 200901) was used to detect 15 clinical samples on a fully automatic biochemical analyzer (Hitachi 7180); after the test, the A group kit was used to detect low-density lipoprotein (LDL-C) on the same sample on the same instrument and at the same detection position; after the test, TBA detection reagent (Chongqing Zhongyuan Huiji Biotechnology Co., Ltd., 200901) was used again for the same sample on the same instrument and at the same detection position

[0077] ②After thoroughly cleaning the instruments (reaction cup, stirring rod) used in experimental group ①, the method described in experimental group ① was used. The only difference was that in the second step, the low-density lipoprotein (LDL-C) test kit of group B was used.

[0078] ③After thoroughly cleaning the instruments (reaction cup, stirring rod) used in experimental group ②, the method described in experimental group ① was used. The only difference was that in the second step, the low-density lipoprotein (LDL-C) test kit of group C was used.

[0079] The test results of the TBA test reagent for 30 samples in the three groups of experiments were summarized, the absolute deviation was calculated, and the number of positive and negative samples before and after was counted. The experimental results are shown in the following table:

[0080] Table 4 Residue test

[0081]

[0082]

[0083] The experimental results show that the kit (Group ①) and the control kit-1 (Group ②) prepared in Example 1 of the present invention will not interfere with the subsequent TBA test results, while the control kit-2 (Group ③) causes serious interference to the subsequent TBA test results, making the test results seriously biased, which is very likely to cause false positives in clinical practice. It is speculated that this is caused by the inclusion of bile salt analogs in the control kit-2. Although the instrument will be cleaned after each test, the bile salt analogs are easy to remain and difficult to clean, which is very likely to interfere with subsequent test items, especially TBA. The present invention uses a special component ratio to replace the function of bile salt analogs in biochemical reagents on the one hand, while maintaining the good performance of the reagent (such as precision and linear range), on the other hand, it avoids the disadvantages of bile salt analogs being easy to remain and having strong interference.

[0084] Example 4

[0085] In this embodiment, a total of 5 groups of experiments were set up, wherein the difference between the kit used in each group of experiments and that in Example 1 was only that the concentrations of the surfactant 1B-66 and the surfactant 2LS-114 in the reagent R1 were different, and the preparation methods of the remaining kits were the same as those in Example 1. The five kits were used to perform precision tests on the median samples, and the test results were shown in the following table:

[0086] Table 5

[0087]

[0088] Note: In this embodiment, the surfactant 1 in the reagent R1 of group A is 1 g / L B-66, and the surfactant 2 is 1 g / L LS-114; the surfactants of group B are 2 g / L B-66 and LS-114; the surfactants of group C are 5 g / L B-66 and LS-114; the surfactants of group D are 10 g / L B-66 and LS-114; the surfactants of group E are 15 g / L B-66 and LS-114. The remaining components and preparation processes are the same as those in Example 1.

[0089] The experimental results show that when the concentration range of surfactant 1B-66 in reagent R1 is 2-10 g / L, and the concentration range of surfactant 2LS-114 is 2-10 g / L, the detection precision of the kit is higher.

[0090] Example 5

[0091] In this embodiment, a total of 5 groups of experiments were set up, wherein the kit used in each group of experiments differed from that in Example 1 only in that the types of surfactant 1 and surfactant 2 in reagent R1 were different, and the preparation methods of the remaining kits were the same as those in Example 1. The five kits were used to perform precision tests on the median samples, and the test results are shown in the following table:

[0092] Table 6

[0093]

[0094] Note: In this embodiment, the surfactant 1 in the reagent R1 of group A is 5g / L TritonX-100, and the surfactant 2 is 5g / L A90; group B is 5g / L A90 and Tween-80; group C is 5g / L Tween-80 and B-66; group D is 5g / L B-66 and LS-114; group E is 5g / L LS-114 and TritonX-100. The remaining components and preparation process are the same as those in Example 1.

[0095] The experimental results show that when surfactant 1 and surfactant 2 in reagent R1 are a combination of the above five surfactants, the detection precision of the kit is relatively high, especially when surfactant 1 is B-66 and surfactant 2 is LS-114, the detection precision is the highest, reaching 0.84%.

[0096] Example 6

[0097] In this embodiment, a total of 5 groups of experiments were set up, wherein the difference between the kit used in each group of experiments and that in Example 1 was only that the concentrations of surfactant 3 Triton 305 and surfactant 4 Brij 58 in reagent R1 were different, and the preparation methods of the remaining kits were the same as those in Example 1. The five kits were used for testing at the same time, and the test results were shown in the following table:

[0098] Table 7

[0099]

[0100] Note: In this embodiment, the surfactant 3 in the reagent R2 of group A is 0.5 g / L Triton305, and the surfactant 2 is 0.5 g / L Brij 58; the surfactant 3 in group B is 1 g / L Triton305 and Brij 58; the surfactant 2 in group C is 2 g / L Triton305 and Brij 58; the surfactant 3 in group D is 5 g / L Triton305 and Brij 58; the surfactant 2 in group E is 10 g / L Triton305 and Brij 58, and the other components and preparation process are the same as those in Example 1.

[0101] The experimental results show that when the concentration range of surfactant 3Triton305 in reagent R2 is 1-3g / L, and the concentration range of surfactant 4Brij 58 is 1-3g / L, the detection precision of the kit is higher.

[0102] Example 7

[0103] In this embodiment, a total of 5 groups of experiments were set up, wherein the kit used in each group of experiments differed from that in Example 1 only in that the types of surfactant 3 and surfactant 4 in reagent R2 were different, and the preparation methods of the remaining kits were the same as those in Example 1. The five kits were used to perform precision tests on the median samples, and the test results were shown in the following table:

[0104] Table 8

[0105]

[0106] Note: In this embodiment, the surfactant 3 in the reagent R1 of group A is 2g / L TritonX-100, and the surfactant 4 is 2g / L A90; group B is 2g / L A90 and Tween-20; group C is 2g / L Tween-20 and Triton305; group D is 2g / L Triton305 and Brij 58; group E is 2g / L Brij 58 and Triton X-100. The remaining components and preparation process are the same as those in Example 1.

[0107] The experimental results show that when surfactant 3 and surfactant 4 in reagent R2 are a combination of the above five surfactants, the detection precision of the kit is relatively high, especially when surfactant 3 is Triton 305 and surfactant 4 is Brij 58, the detection precision is the highest, reaching 0.69%.

[0108] Example 8

[0109] In this embodiment, a total of 5 groups of experiments were set up, wherein the difference between the kit used in each group of experiments and that in Example 1 was only that the protein (hemocyanin) concentration in the reagent R2 was different, and the preparation methods of the remaining kits were the same as those in Example 1. The five kits were used for testing at the same time, and the test results were shown in the following table:

[0110] Table 9

[0111]

[0112] Note: In this embodiment, the hemocyanin concentration in reagent R2 of group A is 1 g / L; that of group B is 2 g / L; that of group C is 5 g / L; that of group D is 10 g / L; and that of group E is 20 g / L. The remaining components and preparation process are the same as those of embodiment 1.

[0113] The experimental results show that when the concentration range of protein (hemocyanin) in reagent R2 is 2-10g / L, the detection precision of the kit is higher.

[0114] Example 9

[0115] In this embodiment, a total of 4 groups of experiments were set up, wherein the kit used in each group of experiments differed from that in Example 1 only in the type of protein in reagent R2, and the preparation methods of the remaining kits were the same as those in Example 1. The three kits were used to perform precision tests on the median samples, and the test results were shown in the following table:

[0116] Table 10

[0117]

[0118] Note: In this embodiment, the protein in reagent R2 of group A is 5 g / L hemocyanin; the protein in group B is 5 g / L casein; the protein in group C is 5 g / L BSA; no protein is added to group C, and the other components and preparation process are the same as those in Example 1.

[0119] The experimental results show that when the protein in reagent R2 is hemocyanin, the detection precision of the kit is significantly improved, reaching 0.81%.

[0120] Although the embodiments of the present invention have been shown and described above, it is to be understood that the above embodiments are exemplary and are not to be construed as limitations on the present invention. A person skilled in the art may change, modify, replace and deform the above embodiments within the scope of the present invention without departing from the principles and purpose of the present invention.

Claims

1. A low-density lipoprotein (LDL-C) detection kit for reducing cross contamination, comprising reagent R1 and reagent R2, characterized in that: The reagent R1 includes 30-100 mmol / L buffer, 300-600U / L cholesterol oxidase, 800-1200U / L cholesterol esterase, 10-20 mmol / L 4-aminoantipyrine, 2-10g / L B-66 and 2-10g / L LS-114; the reagent R2 includes 30-80 mmol / L buffer, 1500-2000U / L peroxidase, 0.7-2 mmol / L TOOS, 1-3g / LBrij 58, 1-3g / L Triton305 and hemocyanin.

2. A low-density lipoprotein (LDL-C) detection kit for reducing cross contamination according to claim 1, characterized in that: The reagent R1 also includes a preservative and an accelerator; the reagent R2 also includes a preservative and ascorbic acid oxidase.

3. A low-density lipoprotein (LDL-C) detection kit for reducing cross contamination according to claim 2, characterized in that: The reagent R1 contains 0.5-1 g / L of preservative and 10-20 g / L of accelerator.

4. A low-density lipoprotein (LDL-C) detection kit for reducing cross contamination according to claim 2 or 3, characterized in that: The reagent R2 contains 1-2 g / L of a preservative and 2500-3500 U / L of ascorbic acid oxidase.

5. A low-density lipoprotein (LDL-C) detection kit for reducing cross contamination according to claim 4, characterized in that: The buffer in the reagent R1 is selected from at least one of Tris buffer, phosphate buffer and GOOD'S buffer; the preservative is selected from at least one of Proclin-300, sodium azide or gentamicin; and the accelerator is selected from at least one of dextran, PEG6000 and PEG8000.

6. A low-density lipoprotein (LDL-C) detection kit for reducing cross contamination according to claim 5, characterized in that: The buffer in the reagent R2 is selected from at least one of Tris buffer, phosphate buffer and GOOD'S buffer; the preservative is selected from at least one of Proclin-300, sodium azide or gentamicin; and the accelerator is selected from at least one of dextran, PEG6000 and PEG8000.

7. A low-density lipoprotein (LDL-C) detection kit with reduced cross contamination according to claim 5 or 6, characterized in that: The pH value of the reagent R1 is 5.5-6.5, and the pH value of the reagent R2 is 7.0-8.

0.

8. A low-density lipoprotein (LDL-C) detection kit for reducing cross contamination according to claim 7, characterized in that: The pH value of the reagent R1 is 6.0, and the pH value of the reagent R2 is 7.

5.

9. A method for reducing cross contamination of low-density lipoprotein (LDL-C) detection reagents, characterized in that: The reagent R1 is added with 30-100 mmol / L buffer, 300-600 U / L cholesterol oxidase, 800-1200 U / L cholesterol esterase, 10-20 mmol / L 4-aminoantipyrine, 2-10 g / L B-66 and 2-10 g / L LS-114; the reagent R2 is added with 30-80 mmol / L buffer, 1500-2000 U / L peroxidase, 0.7-2 mmol / L TOOS, 1-3 g / L Brij 58, 1-3 g / L Triton305 and hemocyanin.

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