BioReagent 级 ，适用于对基线干扰要求严格的色谱和分析工作流程。
2-8°C储存,避光。低温运输 。请查阅批次 COA 获取详细规格。
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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。
无机磷主要指无机磷酸根，参与生物体内多种代谢，包括能量代谢、核酸代谢、蛋白质磷酸化和去磷酸化等，此外还可以促进碳水化合物的合成、转化和转运。
检测原理：钼酸铵与无机磷酸根生成在660nm有特征吸收峰的物质，通过测定660nm吸收度，即可计算无机磷含量。
适用样本：动植物组织
产品组分表
| O1506749 | Component | 96T | Storage |
| O1506749A | Reagent Ⅰ | 120 mL | 2-8℃ |
| O1506749B | Reagent Ⅱ | 3 mL | 2-8℃ |
| O1506749C | Reagent Ⅲ | 1 EA | 2-8℃. Store in the dark. |
| O1506749D | Reagent Ⅳ | 1 EA | 2-8℃. Store in the dark. |
| O1506749E | Standard | 1 mL | 2-8℃ |
注意：正式检测前，建议选择 2-3 个预期差异较大的样本进行预实验。
自备仪器和试剂
1、酶标仪或可见分光光度计（能测 660 nm 处的吸光度）
2、96孔板或微量玻璃比色皿、可调节式移液枪及枪头
3、可调式水浴锅、台式离心机
4、去离子水、匀浆器
实验流程
1、试剂准备
| 试剂名称 | 试剂准备 | 注意事项 |
| Reagent Ⅰ | 即用型；使用前平衡到室温 | 4℃保存；有刺激性气味，建议在通风橱进行实验 |
| Reagent Ⅱ | 即用型；使用前平衡到室温 | 4℃保存；有腐蚀性，建议在通风橱进行实验 |
| Reagent Ⅲ | 临用前配制，加入1.5mL去离子水充分溶解后使用（可置于37℃水浴5min促进溶解） | 4℃避光保存一周 |
| Reagent Ⅳ | 临用前配制，加入6mL去离子水充分溶解后使用 | 4℃避光保存一周 |
| 定磷剂 | 按照去离子水: Reagent Ⅱ: Reagent Ⅲ: Reagent Ⅳ=10:5:2:8 的比例配制 | 请根据实际需要进行配制，仅当天使用 |
| Standard | 即用型；使用前平衡到室温 | 4℃保存 |
2、样本制备
组织样本：称取约0.1g样本，加入1mL Reagent Ⅰ（按照组织质量 (g)： Reagent Ⅰ体积 (mL) 为1：10 的比例）进行冰浴匀浆，10,000g，4℃离心10min，取上清液，置冰上待测。
注意：如需测定蛋白浓度，推荐使用阿拉丁B665595 BCA蛋白定量试剂盒或R1491648 即用型BCA蛋白定量试剂盒。
3、实验步骤
3.1 酶标仪或可见分光光度计预热30min，调节波长到660nm。可见分光光度计用去离子水调零。
3.2 操作表（下述操作在1.5mL离心管中操作）：
| 试剂（μL） | 空白管 | 标准管 | 测定管 |
| 样本上清 | 0 | 0 | 10 |
| 去离子水 | 100 | 90 | 90 |
| Standard | 0 | 10 | 0 |
| 定磷剂 | 100 | 100 | 100 |
3.3 混匀后置于40℃水浴保温10min，室温冷却10min后取200μL 上清液于微量玻璃比色皿或96孔板中660nm处测定吸光度， 记为A空白、A标准、A测定，计算ΔA测定=A测定-A空白、ΔA标准=A标准-A空白。
注意：空白孔和标准孔只需测定1次。实验之前建议选择2-3个预期差异大的样本做预实验。如果ΔA测定过小可适当加大样本量。如果ΔA测定大于1.0，样本可用Reagent Ⅰ进一步稀释，计算结果乘以稀释倍数，或减少提取用样本量。
4、结果计算
注意：我们为您提供的计算公式，包括推导过程计算公式和简洁计算公式。两者完全相等。建议以加粗的简洁计算公式为最终计算公式。
4.1 按照蛋白含量计算
无机磷含量 (μmol/mg prot)=(CStandard×ΔA测定÷ΔA标准)×V总÷Cpr=1×ΔA测定÷ΔA标准÷Cpr
4.2 按照样本质量计算
无机磷含量 (μmol/g 鲜重)=(CStandard×ΔA测定÷ΔA标准)×V总÷W=1×ΔA测定÷ΔA标准÷W
参数说明：
CStandard：1 mmol/L；
V总：上清液总体积，1 mL=0.001 L；
W：样品质量，g。
5、结果展示（实验实例）
5.1 取0.1g绿萝加入1mL Reagent Ⅰ，离心取上清之后按照测定步骤操作，使用96孔板测得A测定=0.334，A空白=0.043，A标准=0.311，按样本质量计算无机磷含量得：无机磷含量 (μmol/g 样本)=1×ΔA测定÷ΔA标准÷0.1=10.86 μmol/g 样本。
5.2 取0.04g小鼠肌肉加入0.4mL Reagent Ⅰ，离心取上清之后按照测定步骤操作，使用96孔板测得A测定=1.201，A空白=0.042，A标准=0.29，按样本质量计算无机磷含量得：无机磷含量 (μmol/g 样本)=0.4×ΔA测定÷ΔA标准÷0.04=46.73 μmol/g 样本。
注意事项
1、定磷剂需临用前配制，限当天使用，建议 40 min 内完成检测。
2、本产品仅供科学研究使用，不适用于临床诊断。为了您的安全和健康，请穿实验服并戴一次性手套操作。
Inorganic phosphorus primarily refers to inorganic phosphate ions, which participate in various metabolic processes within organisms, including energy metabolism, nucleic acid metabolism, and protein phosphorylation/dephosphorylation. Furthermore, it promotes the synthesis, transformation, and transport of carbohydrates.
Assay Principle:Ammonium molybdate reacts with inorganic phosphate ions to form a complex with a characteristic absorption peak at 660 nm. The inorganic phosphorus content is calculated by measuring the absorbance at 660 nm.
Applicable Samples: Animal and plant tissues
| O1506749 | Component | 96T | Storage |
| O1506749A | Reagent I | 120 mL | 2-8℃ |
| O1506749B | Reagent II | 3 mL | 2-8℃ |
| O1506749C | Reagent III | 1 EA | 2-8℃. Store in the dark. |
| O1506749D | Reagent IV | 1 EA | 2-8℃. Store in the dark. |
| O1506749E | Standard | 1 mL | 2-8℃ |
Note: It is recommended to perform a pilot experiment with 2-3 samples expected to have significant differences before formal testing.
Required Materials Not Provided
Microplate reader or visible spectrophotometer (capable of measuring absorbance at 660 nm)
96-well plate or micro glass cuvettes, Adjustable pipettes and tips
Adjustable water bath, Benchtop centrifuge
Deionized water, Homogenizer
Experimental Procedure
1. Reagent Preparation
| Reagent Name | Preparation Steps | Notes & Storage |
| Reagent I | Ready-to-use; equilibrate to room temperature before use. | Store at 4°C. Pungent odor - recommended to use in a fume hood. |
| Reagent II | Ready-to-use; equilibrate to room temperature before use. | Store at 4°C. Corrosive - recommended to use in a fume hood. |
| Reagent III | Before use, dissolve the vial contents in 1.5 mL deionized water (incubation in a 37°C water bath for 5 min may aid dissolution). | Stable at 4°C protected from light for 1 week. |
| Reagent IV | Before use, dissolve the vial contents in 6 mL deionized water. | Stable at 4°C protected from light for 1 week. |
| Phosphorus Assay Reagent | Prepare by mixing Deionized Water : Reagent II : Reagent III : Reagent IV at a ratio of 10 : 5 : 2 : 8. | Prepare according to actual need, use on the same day. |
| Standard | Ready-to-use; equilibrate to room temperature before use. | Store at 4°C. |
2. Sample Preparation
Tissue Samples: Weigh approximately 0.1 g of tissue.
Add 1 mL of Reagent I (using a ratio of tissue mass (g) : Reagent I volume (mL) = 1:10).
Homogenize in an ice bath.
Centrifuge at 10,000 g, 4°C for 10 minutes.
Collect the supernatant and keep on ice for assay.
Note: If determining protein concentration is required, the use of the Aladdin B665595 BCA Protein Quantification Kit or R1491648 Ready-to-Use BCA Protein Quantification Kit is recommended.
3. Assay Procedure
3.1 Instrument Preparation
Preheat the microplate reader or visible spectrophotometer for 30 minutes. Set wavelength to 660 nm.
Zero the visible spectrophotometer with deionized water.
3.2 Assay Setup (Perform in 1.5 mL centrifuge tubes):
| Reagent (μL) | Blank Tube | Standard Tube | Test Tube |
| Sample Supernatant | 0 | 0 | 10 |
| Deionized Water | 100 | 90 | 90 |
| Standard | 0 | 10 | 0 |
| Phosphorus Assay Reagent | 100 | 100 | 100 |
3.3 Measurement
Mix well after addition.
Incubate in a 40°C water bath for 10 minutes.
Cool at room temperature for 10 minutes.
Transfer 200 μL of the supernatant to a micro glass cuvette or 96-well plate.
Measure the absorbance at 660 nm.
Record the values as ABlank, AStandard, and ATest. Calculate ΔAtest = ATest - ABlank and ΔAstandard = AStandard - ABlank.
Note: The Blank and Standard tubes need only be measured once.
A pilot experiment with 2-3 variable samples is recommended.
If ΔAtest is too small, consider increasing the sample amount.
If ΔAtest is greater than 1.0, dilute the sample further with Reagent I and multiply the result by the dilution factor, or reduce the sample amount used for extraction.
4. Calculation of Results
Note: We provide both the derived formula and the simplified formula for your convenience. They are mathematically equivalent. The bolded simplified formula is recommended for final calculations.
4.1 Based on Protein Content:
Derived Formula: Inorganic Phosphorus Content (μmol/mg prot) =(CStandard × ΔAtest ÷ ΔAstandard) × Vtotal ÷ Cpr
Simplified Formula: Inorganic Phosphorus Content (μmol/mg prot) = 1 × ΔAtest ÷ ΔAstandard ÷ Cpr
4.2 Based on Sample Mass:
Precautions
1.The Phosphorus Assay Reagent must be prepared fresh before use and used on the same day. It is recommended to complete the measurements within 40 minutes.
2.This product is for research use only. Not for use in diagnostic procedures. For your safety and health, please wear lab coats and disposable gloves during operation.
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| 批号(Lot Number) | 证书类型 | 货号 |
|---|---|---|
 ZJ26F0131307 |
分析证书 | O1506749 |
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