Soil Inorganic Phosphate (S-PHOS) Content Assay Kit (MB, Micro Method)

Cat. No.: S1506787

GRADE & PURITY BioReagent

Overview

  Phosphorus is an essential macronutrient for plants. Plants primarily absorb phosphorus from the soil through their root systems. Soil phosphorus exists mainly in organic and inorganic forms. Organic phosphorus must be mineralized and converted into inorganic phosphorus before it can be absorbed and utilized by plants.

  Assay Principle:Inorganic phosphorus is extracted from the soil. In an acidic environment, phosphorus is determined by the molybdenum blue method. The absorbance at 660 nm is measured to calculate the inorganic phosphorus content in the soil.

  Applicable Samples: Soil samples

S1506787	Component	48T	96T	Storage
S1506787A	Extraction Buffer	60 mL	120 mL	2-8℃
S1506787B	Reagent Ⅰ	1 EA	1 EA	2-8℃. Store in the dark.
S1506787C	Reagent Ⅱ	1 EA	1 EA	2-8℃. Store in the dark.
S1506787D	Reagent Ⅲ	2.5 mL	5 mL	RT
S1506787E	Standard	1 mL	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

1. Microplate reader or visible spectrophotometer (capable of measuring absorbance at 660 nm)

2. 96-well plate or micro glass cuvettes, Adjustable pipettes and tips

3. Analytical balance, Adjustable water bath, Benchtop centrifuge, 100-mesh sieve

4. Deionized water

Experimental Procedure

1. Reagent Preparation

Reagent Name	Preparation Steps	Notes & Storage
Extraction Buffer	Ready-to-use; equilibrate to room temperature before use.	Store at 4°C.
Reagent Ⅰ	Before use, dissolve the vial contents in 2.5 mL (48T) or 5 mL (96T) deionized water. Mix thoroughly.	Stable at 4°C protected from light for 1 month.
Reagent Ⅱ	Before use, dissolve the vial contents in 2.5 mL (48T) or 5 mL (96T) deionized water. Mix thoroughly.	Stable at 4°C protected from light for 1 month.
Reagent Ⅲ	Ready-to-use.	Store at Room Temperature.
Standard	Ready-to-use; equilibrate to room temperature before use.	Store at 4°C.
Phosphorus Assay Reagent	Prepare by mixing Deionized Water : Reagent I : Reagent II : Reagent III at a ratio of 2 : 1 : 1 : 1. The prepared reagent should be light yellow.	Prepare fresh before use. Discard if colorless (reagent degraded) or blue (phosphorus contaminated). Prepare as needed.

2. Sample Preparation

• Soil Sample: Take an appropriate amount of air-dried soil, remove impurities like stones and twigs, and pass through a 100-mesh sieve.

• Accurately weigh 0.01 g of the sieved soil.

• Add 1 mL of Extraction Buffer.

• Shake or vortex to mix thoroughly at room temperature.

• Incubate in a 40°C water bath for 1 hour for extraction.

• Centrifuge at 8,000 g, 25°C for 10 minutes.

• Collect the supernatant for assay.

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	40
Phosphorus Assay Reagent	40	0	0
Phosphorus Assay Reagent	200	200	200
Standard	0	40	0

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 mixture to a 96-well plate or micro glass cuvette.

Measure the absorbance at 660 nm.

Record the values as A_Blank, A_Standard, and A_Test. 

Calculate ΔA_test = A_Test - A_Blank and ΔA_standard = A_Standard - A_Blank. 

Note: The Blank and Standard wells/tubes only need to be measured once. 

A pilot experiment with 2-3 variable samples is recommended.

If ΔA_test is less than 0.01, consider increasing the sample amount. If ΔA_test is greater than 1.0, dilute the sample further with Extraction Buffer and multiply the result by the dilution factor, or reduce the soil amount used for extraction.

4. Calculation of Results

Soil Inorganic Phosphorus Content Calculation

(Using a 96-well plate or micro glass cuvette):

S-PHOS (μmol/g dry weight) = C_standard × ΔA_test ÷ ΔA_standard × V_total ÷ W = 0.5 × ΔA_test ÷ ΔA_standard × V_total ÷ W = 50 × ΔA_test ÷ ΔA_{standard
Parameter Definitions:}
C_standard: Standard solution concentration, 500 μmol/L 

V_total: Total volume of the supernatant, 1 mL = 0.001 L 

W: Soil sample mass, g

Precautions

The Phosphorus Assay Reagent must be prepared fresh before use and used on the same day.

Complete the measurement within 40 minutes after starting the incubation step (3.3).

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.

Storage and Shipping

Storage

Store at 2-8°C,Protected from light,Room temperature

Shipped In

Wet ice

Stability And Storage

Each component has a shelf life of 1 year under corresponding storage conditions.