BioReagent 级 ，适用于对基线干扰要求严格的色谱和分析工作流程。
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肌酐（creatinine，Cr）是人体或动物肌肉内代谢的产物，每20g肌肉代谢可产生约1mg肌酐，由肾小球滤过排出体外，外源性肌酐是肉类食物在体内代谢后的产物，内源性肌酐是体内肌肉组织代谢的产物。
检测原理：血清、血浆、尿液中的肌酐与苦味盐反应，生成橘红色的苦味盐肌酐复合物的反应速率，选择适宜的速率监测时间，避开干扰物质对肌酐与苦味盐反应的干扰，通过酶标仪测定510nm处吸光度，可用于测定人体、动物的血浆、血清、尿液样品中肌酐含量。该试剂盒仅用于科研领域，不宜用于临床诊断或其他用途。
| C1505879 | Component | 100T | Storage |
| C1505879A | 肌酐标准 (10mmol/L) | 1 mL | 2-8℃. Store in the dark. |
| C1505879B | 肌酐标准稀释液 | 2 mL | RT |
| C1505879C | Cr显色液 | 10 mL | RT |
| C1505879D | Cr Assay Buffer | 10 mL | RT |
自备仪器和试剂
1、蒸馏水
2、酶标仪、96孔板、恒温箱或水浴锅
实验流程
1、准备样品
血浆、血清按照常规方法制备，-20℃冻存。如果尿液中肌酐含量较高，可用蒸馏水作 1：10~100 稀释后再测定。一般不建议稀释倍数过高，否则导致吸光度差值较小。
2、配制标准品工作液
按肌酐标准 (10mmol/L)：肌酐标准稀释液=1:49的比例混合，使浓度达到200μmol/L，即为标准品工作液-肌酐标准(200μmol/L)。4℃保存1周有效。
3、配制Cr显色工作液
取Cr显色液和Cr Assay Buffer等量混合，室温放置20min，即为Cr显色工作液。
4、Cr加样
按照下表设置标准孔、测定孔，溶液应按照顺序依次加入，并注意避免产生气泡。如果样品中的Cr浓度过高，可以适当稀释10~100倍后再进行测定。
| 加入物 (μl) | 标准孔 | 测定孔 |
| 肌酐标准 (200μmol/L) | 20 | — |
| 血清、血浆、稀释尿液 | — | 20 |
| Cr显色工作液 | 200 | 200 |
注意：对于某些干扰物较多的样品，可以提高肌酐标准的工作浓度以减少误差，如配制浓度为 300~400μmol/L 的肌酐标准进行测定，其计算公式中应相应×300~400，而不是×200。一般先加标准品和待测样品，再加入Cr显色工作液后准确计时、读数。
5、Cr测定
反应温度37℃，充分混匀后准确反应20s，酶标仪测定510nm吸光度，读取各孔吸光度（A标准1、A测定1）；待反应进行至准确60s，再读取各孔吸光度（A标准2、A测定2）。注意：采用酶标仪测定时，一般建议用临床生化分析仪，手工或半手工操作有可能不准确，或者测定效果不佳。
6、计算
肌酐 (μmol/L)=(A测定2-A测定1)/(A标准2-A标准1)×200×N
参数说明：
A测定1：反应20s测定孔的吸光度；
A测定2：反应60s测定孔的吸光；
A标准1：反应20s标准孔的吸光度；
A标准2：反应60s标准孔的吸光度；
N：样本稀释倍数
参考区间：
| 成年人男性血肌酐 | 62~115μmol/L (0.7-1.3mg/dl) |
成年人女性血肌酐 | 53~97μmol/L(0.6-1.1mg/dl) |
参考标准曲线范围：测定肌酐标准在200μmol/L时，通过酶标仪505nm测定60s与20s的吸光度差值大多在0.02~0.05之间。测定肌酐标准在0、50、100、200、300、400μmol/L的差值，据此作出其标准曲线：
注意：由于检测仪器和操作手法等条件的不同，参考值范围会有波动，该值仅供参考，对于要求精确计算肌酐含量的，可以采用标准曲线进行多点测定。
注意事项
1、由于本产品检测方法属于速率法，其全程反应时间在2~3min 左右，反应较快，因此测定的关键点在检测时间，一定注意操作时间，否则结果会显示不出差异。
2、该 PA 速率法线性范围可达2000μmol/L，样本浓度过高时应稀释后再测定。
3、测定各孔时，待测样品、标准品温度均需达到室温，否则影响结果。
4、轻度溶血样本对肌酐测定无影响。
5、一次性不宜同时测定过多样本，避免因延长加入试剂时间导致测定结果的不准确。建议在加入样品后，最多一次性加入2~3个管的Cr显色工作液，如采用排枪，可适当增加一次性加入的管数。
6、一般尿液肌酐含量较高，如果显色后吸光度仍超过本法的线性范围 (即2000μmol/L)，可用蒸馏水稀释 10~100 倍后再重新进行测定。
7、为了您的安全和健康，请穿实验服并戴一次性手套操作。
8、试剂开封后请尽快使用，以防影响后续实验效果。
Creatinine (Cr) is a metabolic product of human or animal muscle. Approximately 1 mg of creatinine is produced for every 20 g of muscle metabolized. It is filtered and excreted by the glomeruli. Exogenous creatinine originates from the metabolism of meat in the diet, while endogenous creatinine is derived from the metabolism of muscle tissue within the body.
Assay Principle
This assay measures the reaction rate between creatinine in serum, plasma, or urine and picrate, forming an orange-red creatinine-picrate complex. By selecting an appropriate rate monitoring time, interference from substances that affect the creatinine-picrate reaction is minimized. The absorbance at 510 nm is measured using a microplate reader. This kit can be used to determine creatinine content in human or animal plasma, serum, and urine samples. This product is for research use only and is not suitable for clinical diagnosis or other purposes.
| C1505879 | Component | 100T | Storage |
| C1505879A | Creatinine Standard (10 mmol/L) | 1 mL | 2-8℃. Store in the dark. |
| C1505879B | Creatinine Standard Diluent | 2 mL | RT |
| C1505879C | Cr Chromogenic Solution | 10 mL | RT |
| C1505879D | Cr Assay Buffer | 10 mL | RT |
Required Materials Not Provided
Distilled water
Microplate reader, 96-well plate, Incubator or water bath
Experimental Procedure
1. Sample Preparation
Prepare plasma and serum by conventional methods. Store at -20°C.
If the creatinine concentration in urine is high, dilute it 1:10 to 1:100 with distilled water before assay. Excessively high dilution factors are generally not recommended as they may lead to small absorbance differences.
2. Preparation of Standard Working Solution
Mix the Creatinine Standard (10 mmol/L) with the Creatinine Standard Diluent at a 1:49 ratio to achieve a concentration of 200 μmol/L. This is the Creatinine Standard Working Solution (200 μmol/L).
Store at 4°C; stable for 1 week.
3. Preparation of Cr Chromogenic Working Solution
Mix equal volumes of Cr Chromogenic Solution and Cr Assay Buffer.
Allow the mixture to stand at room temperature for 20 minutes. This is the Cr Chromogenic Working Solution.
4. Cr Assay Setup
Set up Standard and Test wells according to the table below.
Add reagents in the specified order, avoiding bubbles.
If the sample Cr concentration is too high, dilute it appropriately (10-100 fold) before assay.
| Reagent (μL) | Standard Well | Test Well |
| Creatinine Standard (200 μmol/L) | 20 | — |
| Serum, Plasma, Diluted Urine | — | 20 |
| Cr Chromogenic Working Solution | 200 | 200 |
Note: For samples with many interfering substances, increasing the concentration of the creatinine standard (e.g., 300-400 μmol/L) can reduce error. In this case, adjust the calculation formula accordingly (multiply by 300-400 instead of 200).
Generally, add the standard and test samples first, then add the Cr Chromogenic Working Solution, starting accurate timing and reading immediately after addition.
5. Cr Measurement
React at 37°C.
Mix thoroughly immediately after adding the working solution.
Precisely at 20 seconds, measure the absorbance at 510 nm (recorded as Astandard1 and Atest1).
Precisely at 60 seconds, measure the absorbance again (recorded as Astandard2 and Atest2).
Note: When using a microplate reader, a clinical chemistry analyzer is recommended. Manual or semi-manual operation may be inaccurate or yield suboptimal results.
6. Calculation
Creatinine (μmol/L) = [(Atest2 - Atest1) / (Astandard2 - Astandard1)] × 200 × N
Parameter Definitions:
Atest1: Absorbance of Test well at 20 seconds
Atest2: Absorbance of Test well at 60 seconds
A<sub>standard1</sub>: Absorbance of Standard well at 20 seconds
A<sub>standard2</sub>: Absorbance of Standard well at 60 seconds
N: Sample dilution factor
Reference Interval:
| Adult Males | 62~115μmol/L (0.7-1.3mg/dl) |
| Adult Females | 53~97μmol/L(0.6-1.1mg/dl) |
Typical ΔA for 200 μmol/L Creatinine Standard: The difference in absorbance between 60s and 20s (ΔA) for a 200 μmol/L creatinine standard typically ranges from 0.02 to 0.05 when measured at 505 nm.
A standard curve can be constructed by measuring the ΔA values for creatinine standards at 0, 50, 100, 200, 300, and 400 μmol/L.
Note: Reference ranges may vary due to differences in instruments and operational techniques. For precise creatinine quantification, use a multi-point standard curve.
Precautions
This assay is based on the rate method; the entire reaction is fast (approx. 2-3 min). Precise timing is critical; failure to adhere to timing may result in undetectable differences.
The linear range of this rate method extends up to 2000 μmol/L. Dilute samples with high concentrations before assay.
Ensure both samples and standards reach room temperature before assay, as temperature affects results.
Slight hemolysis does not affect the creatinine assay.
Avoid processing too many samples simultaneously to prevent inaccuracies from prolonged reagent addition times. After adding samples, add the Cr Chromogenic Working Solution to a maximum of 2-3 tubes at once. A multi-channel pipette can slightly increase this number.
Urine creatinine levels are generally high. If the absorbance after color development still exceeds the linear range (2000 μmol/L), dilute the urine sample 10-100 fold with distilled water and repeat the assay.
For your safety and health, wear lab coats and disposable gloves during operation.
Use reagents promptly after opening to avoid affecting subsequent experimental results.
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| 批号(Lot Number) | 证书类型 | 货号 |
|---|---|---|
 ZJ26F0131809 |
分析证书 | C1505879 |
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