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- 技术资料
- 保存条件：
2-8℃
- 保质期：
6个月
- 英文名：
Soil Alkaline Phosphatase(S-AKP/ALP) Activity Assay Kit
- 库存：
99
- 供应商：
北京索莱宝科技有限公司
- CAS号：
BC0285-100T/96S
- 规格：
100管/96样
产品组成：使用前请认真核对试剂体积与瓶内体积是否一致，有疑问请及时联系索莱宝工作人员。
| 试剂名称 | 规格 | 保存条件 |
| 试剂一 | 液体42 mL×1瓶 | 2-8℃保存 |
| 试剂二 | 粉剂×1瓶 | 2-8℃保存 |
| 试剂三 | 液体2.5 mL×1瓶 | 2-8℃保存 |
| 试剂四 | 粉剂×2支 | 2-8℃保存 |
| 标准品 | 液体1 mL×1支 | 2-8℃保存 |
溶液的配制：
- 试剂二：临用前加100 mL蒸馏水充分溶解，2-8℃保存8周；
- 试剂四：临用前取1支加576 μL无水乙醇（自备），24 μL蒸馏水充分溶解。用不完的试剂2-8℃保存一周（变褐色后不能再使用，一瓶即可以实验100T，为延长反应时间故多给一瓶）；
- 标准品：0.5 μmol/mL苯-酚标准液。
产品说明：
土壤磷酸酶是一类催化土壤有机磷矿化的酶，其活性的高低直接影响着土壤中有机磷的分解转化及其生物有效性，是评价土壤磷素生物转化方向与强度的指标。土壤磷酸酶受到土壤碳、氮含量、有效磷含量和pH显著影响，根据最适pH范围，通常分为酸性、中性和碱性三种类型。
碱性环境中，S-AKP/ALP催化磷酸苯二钠水解生成苯-酚和磷酸氢二钠，通过测定酚的生成量即可计算出S-AKP/ALP活性。
注意：实验之前建议选择2-3个预期差异大的样本做预实验。如果样本吸光值不在测量范围内建议稀释或者增加样本量进行检测。
需自备的仪器和用品：
分光光度计/酶标仪、微量玻璃比色皿/96孔板、台式离心机、水浴锅/恒温培养箱、分析天平、可调式移液器、冰、研钵、蒸馏水、30-50目筛、乙醇和甲苯（不允许快递）。
操作步骤：
一、样本处理（可适当调整待测样本量，具体比例可以参考文献）
1. 新鲜土样自然风干或37℃烘箱风干，过30~50目筛。
2. 称取风干混匀土壤约0.1g，加入0.05mL甲苯（自备），轻摇15min；加0.4mL试剂一并且摇匀后，置于37℃水浴锅/恒温培养箱，开始计时，催化反应24h；到时后迅速加入1mL试剂二充分混匀，以终止酶催化的反应。
10000rpm室温离心10min，取上清液置于冰上待测。
二、测定步骤
- 分光光度计/酶标仪预热30min以上，调节波长到660nm，分光光度计蒸馏水调零。
- 空白管：取微量比色皿/96孔板，加入10μL试剂一，20μL试剂三，4μL试剂四，充分混匀，显色后再加蒸馏水166μL，混匀后室温静置30min，于660nm测定吸光度，记为A空白管。（空白管只需要测定1-2次）
- 标准管：取微量比色皿/96孔板，加入10μL标准液，20μL试剂三，4μL试剂四，充分混匀，显色后再加蒸馏水166μL，混匀后室温静置30min，于660nm测定吸光度，记为A标准管。（标准管只需要测定1-2次）
- 测定管：取微量比色皿/96孔板，加入10μL上清液，20μL试剂三，4μL试剂四，充分混匀，显色后再加蒸馏水166μL，混匀后室温静置30min，于660nm测定吸光度，记为A测定管。
三、S-AKP/ALP活性计算
活性单位定义：37℃中每克土壤每天释放1nmol酚为1个酶活单位。
S-AKP/ALP活性（U/g 土样）=[C标准液×(A测定管-A空白管)÷(A标准管-A空白管)]×V总÷W÷T×1000
=725×(A测定管-A空白管)÷(A标准管-A空白管)÷W
C标准液：0.5μmol/mL；V总：催化体系总体积，1.45mL；W：土壤样本质量，g；T：催化反应时间，24 h=1d；1000：单位换算系数，1μmol=1000nmol。
相关发表文献：
- Liu B, Wang S, Wang J, et al. The great potential for phytoremediation of abandoned tailings pond using ectomycorrhizal Pinus sylvestris[J]. Science of The Total Environment, 2020, 719: 137475.
- Shao T, Zhao J J, Liu A, et al. Effects of soil physicochemical properties on microbial communities in different ecological niches in coastal area[J]. Applied Soil Ecology, 2020: 103486.
参考文献：
- Guan S. Soil Enzyme and Its Research Method [M]. Beijing: Science Press, 1982.
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