Glutamate (Glu) Content Assay Kit (Colorimetric Method)

Cat. No.: G1515930

GRADE & PURITY BioReagent

Overview

Glutamic acid (Glu) is widely distributed in animals, plants, microorganisms and cultured cells. As one of the 20 proteinogenic amino acids, it participates in the synthesis of various amino acids via transamination and serves as a major source of amino groups in organisms. In addition, glutamic acid is the principal active ingredient of monosodium glutamate and is commonly applied as a food additive and for spice production.

Assay Principle:

Samples are extracted with dedicated extraction solution followed by chromogenic reaction with color-developing reagent; absorbance is measured at 570 nm after color development.

Note: This kit is for research purpose only and shall not be used for clinical diagnosis or other applications.

Contents & Storage:

Item No.	Appearance	Components	50T	Storage
G1515930A	Liquid	Reagent 1: Liquid	100 mL	2-8℃.
G1515930B	Solid	Reagent 2: Powder	1 EA	2-8℃.

Add 10 mL of distilled water to Reagent 2 before use, mix thoroughly until completely dissolved. Unused reagent shall be stored at 4°C.

Self-prepared Instruments and Reagents:

⁕Spectrophotometer (capable of absorbance measurement at 570 nm)

⁕Benchtop centrifuge, adjustable pipettes, 1 mL glass cuvettes, mortar

⁕Ice, distilled water

Glutamic Acid Extraction:

1. Bacteria or cultured cell samples: Collect bacteria or cells into centrifuge tubes and discard supernatant after centrifugation. Add Reagent Ⅰ at a volume ratio of 500–1000 : 1 (10⁴ cells : mL Reagent Ⅰ; recommended: add 2 mL Reagent Ⅰ per 10 million cells). Lyse cells by sonication on ice (power: 20% or 200 W, sonicate for 3 s, pause for 10 s, repeat 30 times). Centrifuge at 8000 g for 10 min at ambient temperature, collect supernatant and keep on ice pending assay.

2. Tissue samples: Homogenize tissue on ice with Reagent Ⅰ at a mass-volume ratio of 1 g tissue : 5–10 mL Reagent Ⅰ (recommended: weigh ~0.2 g tissue and add 2 mL Reagent Ⅰ). Centrifuge at 8000 g for 10 min at ambient temperature, collect supernatant and keep on ice pending assay.

3. Serum (plasma) or cell culture medium samples: Mix sample with Reagent Ⅰ on ice at a volume ratio of 1 : 5–10 (recommended: add 2 mL Reagent Ⅰ into 0.2 mL serum/plasma or cell culture medium). Centrifuge at 8000 g for 10 min at ambient temperature, collect supernatant and keep on ice pending assay.

Assay Procedures:

1. Preheat spectrophotometer for no less than 30 min, set wavelength to 570 nm and zero the instrument with distilled water.

2. Add listed reagents into capped EP tubes as follows:

Reagent name (μl)	Assay Tube	Control Tube
Sample	1000	/
Reagent 1	/	1000
Reagent 2	200	200

Mix thoroughly, incubate in a 90 °C water bath for 20 min with tubes tightly capped to avoid water evaporation. Cool under running tap water, then measure absorbance at 570 nm. Calculate ΔA = A_assay − A_control. Only one control tube is required. . Only one control tube is required.

Calculation of Glutamic Acid Content:

1. The regression equation obtained under standard calibration: y=0.0074x-0.5255; Where: x= glutamic acid concentration (μg/ml); y= absorbance value.

2. Calculation based on serum (plasma) or cell culture medium volume

Glutamic acid (μg/ml)=[(ΔA+0.5255)÷0.0074×V1]÷(V3×V1÷V2）=1351×(ΔA+0.5255)

3. Calculation based on protein concentration

Glutamic acid (μg/mg prot)=[(ΔA+0.5255)÷0.0074×V1]÷(V1×Cpr)=135.1×(ΔA+0.5255)÷Cpr

4. Calculation based on fresh sample weight

Glutamic acid (μg/g fresh weight)=[(ΔA+0.5255)÷0.0074×V1]÷(W×V1÷V2)=270.2×(ΔA+0.5255)÷W

5. Calculation based on bacterial/cell density

Glutamic acid (μg/10⁴ cell)=[(ΔA+0.5255)÷0.0074×V1]÷(1000×V1÷V2)=0.27×(ΔA+0.5255)

V1: Sample volume added into reaction system, 1ml

V2: Volume of extraction solution (Reagent Ⅰ), 2ml

V3: Initial volume of serum/plasma or cell culture medium, 0.2ml

Cpr: Sample protein concentration, mg/ml

W: Fresh weight of sample, g

1000: Total number of bacteria/cells (ten million cells)

Notes:

1. This kit is only used for glutamic acid quantification in fermentation broth or tissue samples; the lower limit of detection is 100μg/ml。

2. Linear range of standard curve: 100μg/ml-600μg/ml。

3. Valid linear range of ΔA: 0.01-2; If measured ΔA>2, dilute the supernatant with Reagent 1 appropriately before testing, and multiply the final result by the corresponding dilution factor in calculation.

Storage and Shipping

Storage

Store at 2-8°C

Shipped In

Wet ice

Stability And Storage

Store at 2-8℃ long term (6 months).

Contents & Storage

Item No.	Appearance	Components	50T	Storage
G1515930A	Liquid	Reagent 1: Liquid	100 mL	2-8℃.
G1515930B	Solid	Reagent 2: Powder	1 EA	2-8℃.

Add 10 mL of distilled water to Reagent 2 before use, mix thoroughly until completely dissolved. Unused reagent shall be stored at 4°C.