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计算溶液所需的质量、体积或浓度。
尿酸 (UA) 含量检测试剂盒 (尿酸酶, 微量法)
有货
为什么选择此级别
BioReagent,标曲法,比色法,适用于分析 级 ，适用于对基线干扰要求严格的色谱和分析工作流程。
储存与运输
-20°C储存。超低温运输 。请查阅批次 COA 获取详细规格。
质量文档
SDS、COA、产品数据表及规格说明书均可下载。可通过批号查询获取批次 COA。
文献证明
在色谱分析、有机合成和交叉偶联反应领域已被 0 篇同行评审文献引用。
概述
产品介绍
尿酸(UA) 是鸟类和爬行类动物的主要代谢产物，正常人体尿液中产物主要为尿素，含少量尿酸。此外，UA还是重要的抗氧化剂，能清除超氧化物，羟自由基等。体内UA生成量和排泄量不平衡会导致多种疾病的发生。例如，血中UA升高会引起痛风、肾功能损害和动脉硬化，相反UA降低会引起恶性贫血，在临床诊断上具有重要的意义。
本试剂盒检测原理：尿酸酶催化尿酸(UA) 生成尿囊素、CO₂及 H₂O₂，H₂O₂ 在过氧化物酶作用下进一步与酚和 4-氨基安替比林反应生成红色醌类化合物，在 510 nm下有特征吸收峰，测定反应体系510 nm的吸收值，可计算尿酸的含量。
U1516007 | 组件 | 物理外观 | 100 T | 500 T | 储存条件 | 单次测试使用量 |
U1516007A | 尿酸酶 | 无色液体 | 0.4 mL | 2.0 mL | -20℃.避光. | 4 µL |
U1516007B | 过氧化物酶 | 无色液体 | 0.2 mL | 1.0 mL | -20℃.避光. | 2 µL |
U1516007C | 提取液 | 无色液体 | 20 mL | 100 mL | -20℃. | 按需 |
U1516007D | 反应缓冲液 | 无色液体 | 30 mL | 150 mL | -20℃. | 100 µL |
U1516007E | 显色液1 | 无色液体 | 5 mL | 25 mL | -20℃.避光. | 50 µL |
U1516007F | 显色液2 | 无色液体 | 1 mL | 5 mL | -20℃.避光. | 10 µL |
U1516007G | 标准品 | 粉末 | 4.2 mg | 4.2 mg | -20℃.避光. | 按需 |
注意：U1516007F (显色液2)有毒，建议规范穿戴手套和实验服，避免皮肤接触。
使用方法
1. 试剂准备
(1) 将提取液，反应缓冲液，显色液1和显色液2室温解冻，完全平衡至室温后再使用。
(2) 尿酸酶、过氧化物酶冰上解冻。
(3) 标准品配制：将标准品粉末用10 mL反应缓冲液完全溶解 (如不能完全溶解，可50℃水浴加热至完全溶解)，配制成终浓度2.5 mM的标准品溶液，然后分装成小管，-20℃避光保存，避免反复冻融。如果短时间内多次使用，也可暂放4℃保存。
2. 样品准备
(1) 细胞 (菌) 样本：离心收集细胞 (菌)后，建议每百万个细胞 (菌) 用100-200 μL提取液重悬，然后在4℃或冰浴等低温条件下进行超声破碎细胞 ( 200 W，超声2 S，间隔3 S，5 min)。4ºC, 12,000×g离心5分钟，取上清用于后续检测。
(2) 对于组织样品，每10 mg组织加入100 -200 μL 提取液，在4℃或冰浴等低温条件下进行匀浆破碎。4ºC约12,000×g离心5分钟，取上清用于后续检测。
(3) 对于血清 (血浆) 样本：离心后可直接取上清检测，如超出检测上限，可用反应缓冲液稀释后再检测。
(4) 对于尿液样本：建议先用反应缓冲液稀释合适倍数后再检测，直接原液检测可能会导致结果偏低。
3. 显色工作液的配制
等反应缓冲液平衡至室温后，按照每个检测反应166 μL的体积配制显色工作液。根据待检测样品(包括标准品)的数量，配制适量的显色工作液。显色工作液的各个组份的具体配制方法参考下表。建议尽量现配现用，如果不能及时检测，请置于4℃或者冰浴避光保存且不要过夜，以免影响检测效果。
样本数 | 1 | 10 | 100 | 500 |
反应缓冲液 | 100 μL | 1.0 mL | 10 mL | 50 mL |
尿酸酶 | 4 μL | 40 μL | 400 μL | 2000 μL |
过氧化物酶 | 2 μL | 20 μL | 200 μL | 1000 μL |
显色液1 | 50 μL | 500 μL | 5.0 mL | 25.0 mL |
显色液2 | 10 μL | 100 μL | 1.0 mL | 5.0 mL |
4. 样本测定
(1) 标准曲线绘制：取100 μL前面配制好的尿酸标准品(2.5 mM) +150 μL 超纯水稀释至1 mM，然后用超纯水分别梯度稀释至0.5 mM，0.25 mM，0.125 mM，0.0625 mM，0.03125 mM，0 mM（空白孔，仅含超纯水）。取50 μL稀释后的不同浓度标准品溶液至96孔板中，然后每孔分别添加166 μL显色工作液，混匀后，于37℃反应30 min后读取A510。
(2) 样品测定：同样取50 μL样品或稀释后的样品至96孔板中，然后每孔分别添加166 μL显色工作液(对照孔设置为将显色工作液中的尿酸酶替换为同体积的反应缓冲液)，混匀后，于37℃反应30 min后读取A510。
5. 结果计算
(1) 标准曲线：y=0.965*X - 0.0054(仅供参考)；x是尿酸浓度(mM)，y 是在510 nm对应的吸光值。
(2) 建立标准曲线时，需要减去空白孔对照。标准曲线建立完毕后，可根据标准曲线计算出样品中的尿酸浓度（C），计算公示如下：
C(mM)=A×n
注：A为根据标准曲线计算出的尿酸浓度(mM)；n为样品稀释倍数。
注意事项
1. 显色工作液建议现用现配，且注意避光。
2. 检测尿液等液体样本时，建议先稀释合适倍数后看哪个稀释倍数信号值落在检测范围之内，然后选择该稀释倍数稀释样品后再检测，不建议直接用原液检测。
3. 本试剂盒仅用于科研用途，不得用于临床检测或其它用途。
4. 在实验过程中，请规范穿戴手套和实验服，避免皮肤直接接触试剂盒组分。
Product Introduction
Uric acid (UA) is the major metabolic end-product in birds and reptiles. In normal human urine, urea predominates with only a small amount of uric acid present. Furthermore, UA acts as a critical antioxidant capable of scavenging superoxide anions, hydroxyl radicals and other reactive species. Imbalance between endogenous UA production and excretion triggers a variety of diseases. For instance, elevated serum UA may lead to gout, renal impairment and arteriosclerosis, whereas reduced UA level is associated with pernicious anemia. Therefore, UA detection bears great significance in clinical diagnosis.
In this kit: Uricase catalyzes the conversion of uric acid (UA) into allantoin, CO₂ and H₂O₂. In the presence of peroxidase, H₂O₂ further reacts with phenol and 4-aminoantipyrine to produce red quinonoid compounds, which exhibit a characteristic absorption peak at 510 nm. The uric acid content can be calculated by measuring the absorbance of the reaction system at 510 nm.
U1516007 | Components | Appearance | 100T | 500T | Storage | volume per test |
U1516007A | Uricase | Colorless liquid | 0.4 mL | 2.0 mL | -20℃.Store in the dark. | 4 µL |
U1516007B | Peroxidase | Colorless liquid | 0.2 mL | 1.0 mL | -20℃.Store in the dark. | 2 µL |
U1516007C | Extraction Buffer | Colorless liquid | 20 mL | 100 mL | -20℃. | As need |
U1516007D | Reaction Buffer | Colorless liquid | 30 mL | 150 mL | -20℃. | 100 µL |
U1516007E | Chromogenic Solution 1 | Colorless liquid | 5 mL | 25 mL | -20℃.Store in the dark. | 50 µL |
U1516007F | Chromogenic Solution 2 | Colorless liquid | 1 mL | 5 mL | -20℃.Store in the dark. | 10 µL |
U1516007G | Standard | Powder | 4.2 mg | 4.2 mg | -20℃.Store in the dark. | As need |
Note: U1516007F (Chromogenic Solution 2) is toxic. Standard laboratory attire including gloves and lab coats is recommended to avoid skin contact.
Usage Method
1. Reagent Preparation
(1) Thaw Extraction Buffer, Reaction Buffer, Chromogenic Solution 1 and Chromogenic Solution 2 at room temperature. Allow all reagents to fully equilibrate to room-temperature prior to use.
(2) Thaw uricase and peroxidase on ice.
(3) Standard preparation: Completely dissolve the standard powder with 10 mL Reaction Buffer. If full dissolution cannot be achieved, heat in a 50 °C water-bath until completely dissolved to prepare the standard solution with a final concentration of 2.5 mM. Aliquot into small tubes and store at −20 °C; avoid repeated freeze-thaw cycles. For frequent short-term use, temporary storage at 4 °C is acceptable.
2. Sample Preparation
(1) Cell (bacterial) samples: Collect cells (bacteria) by centrifugation. Resuspend every one million cells (bacteria) in 100-200 μL Extraction Buffer. Perform sonication for cell lysis under low-temperature conditions (4 °C or ice bath; 200 W, sonicate for 2 s, interval 3 s, total 5 min). Centrifuge at 12 000×g for 5 min at 4 °C, and collect the supernatant for subsequent detection.
(2) Tissue samples: Add 100-200 μL Extraction Buffer per 10 mg tissue. Homogenize for tissue lysis under low-temperature conditions (4 °C or ice bath). Centrifuge at approximately 12 000×g for 5 min at 4 °C, and collect the supernatant for subsequent detection.
(3) Serum (plasma) samples: The supernatant can be directly used for detection after centrifugation. If the value exceeds the upper detection limit, dilute the sample with Reaction Buffer prior to assay.
(4) Urine samples: It is recommended to dilute urine with Reaction Buffer to an appropriate fold-dilution before measurement. Direct testing of undiluted urine may result in underestimated results.
3. Preparation of Chromogenic Working Solution
After the Reaction Buffer has equilibrated to room temperature, prepare the chromogenic working solution at a volume of 166 μL per reaction. Prepare an appropriate amount according to the total number of samples to be tested (including standards). Refer to the table below for detailed formulation of each component of the chromogenic working solution. It is recommended to prepare fresh working solution immediately before use. If assay cannot be performed promptly, store the solution at 4 °C or on ice in the dark; do not store overnight to avoid compromising assay performance.
Number of sample | 1 | 10 | 100 | 500 |
Reaction Buffer | 100 μL | 1.0 mL | 10 mL | 50 mL |
Uricase | 4 μL | 40 μL | 400 μL | 2000 μL |
Peroxidase | 2 μL | 20 μL | 200 μL | 1000 μL |
Chromogenic Solution 1 | 50 μL | 500 μL | 5.0 mL | 25.0 mL |
Chromogenic Solution 2 | 10 μL | 100 μL | 1.0 mL | 5.0 mL |
4. Sample Measurement
(1) Standard curve preparation: Dilute 100 μL of the previously-prepared 2.5 mM uric acid standard with 150 μL ultrapure water to obtain a 1 mM working standard. Further perform serial dilution with ultrapure water to generate concentrations of 0.5 mM, 0.25 mM, 0.125 mM, 0.0625 mM, 0.03125 mM and 0 mM (blank well containing only ultrapure water). Pipette 50 μL of each diluted standard solution into corresponding wells of a 96-well plate. Add 166 μL chromogenic working solution to each well. Mix well, incubate at 37 °C for 30 min, then record absorbance at A510.
(2) Sample measurement: Pipette 50 μL of sample or diluted sample into wells of a 96-well plate, and add 166 μL chromogenic working solution to each well. For control wells, replace uricase in the chromogenic working solution with an equal volume of Reaction Buffer. Mix thoroughly, incubate at 37 °C for 30 min, and read A510.
5. Calculation of Results
(1) Standard curve: (y = 0.965*X - 0.0054) (for reference only); where x represents uric acid concentration (mM), and y denotes the corresponding absorbance at 510 nm.
(2) When establishing the standard curve, the blank well control needs to be subtracted. After the standard curve is established, the uric acid concentration (C) in the sample can be calculated according to the standard curve, and the calculation formula is as follows:
C (mM) = A × n
Note: A represents the uric acid concentration (mM) calculated from the standard curve; n is the sample dilution factor.
Precautions
1. The chromogenic working solution should preferably be prepared freshly before use and protected from light.
2. When testing liquid samples such as urine, dilute the sample by appropriate fold-dilutions first to identify which dilution yields signals within the assay detection range. Perform the assay using this optimized dilution. Testing undiluted raw sample is not recommended.
3. This kit is for research‑only use. It shall not be applied for clinical diagnosis or any other purposes.
4. During the experiment, wear gloves and lab coats as standard laboratory practice to avoid direct skin contact with kit components.
规格
规格或纯度
BioReagent, 比色法, 适用于分析, 标曲法
英文名称
Uric Acid (UA) Content Assay Kit (Uricase, Microplate Method)
应用
细胞代谢
样品类型
动物组织, 植物组织, 细胞, 细菌, 血浆, 血清
检测范围
0.0315 mM-1 mM
检测方法
尿酸酶，微量法
定量&半定量
定量
检测仪器
酶标仪
检测波长
510 nm
敏感性
对光敏感
检测时长
30 min
样本体积
50 μL
储存与运输
储存条件
-20°C储存
运输条件
超低温运输
稳定性与储存
各组分在相应的储存条件下保质期1年。
组分与储存
U1516007 | 组件 | 物理外观 | 100T | 500T | 储存 | 单次测试使用量 |
U1516007A | 尿酸酶 | 无色液体 | 0.4 mL | 2.0 mL | -20℃.避光 | 4 µL |
U1516007B | 过氧化物酶 | 无色液体 | 0.2 mL | 1.0 mL | -20℃.避光 | 2 µL |
U1516007C | 提取液 | 无色液体 | 20 mL | 100 mL | -20℃. | 按需 |
U1516007D | 反应缓冲液 | 无色液体 | 30 mL | 150 mL | -20℃. | 100 µL |
U1516007E | 显色液1 | 无色液体 | 5 mL | 25 mL | -20℃.避光 | 50 µL |
U1516007F | 显色液2 | 无色液体 | 1 mL | 5 mL | -20℃.避光 | 10 µL |
U1516007G | 标准品 | 粉末 | 4.2 mg | 4.2 mg | -20℃.避光 | 按需 |
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