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Cat. No.: HPGWB-500 (for 500T)

Cat. No.: HPGWB-2k (for 2000T)

Description

HPG Protein Synthesis Kit by Western Blot, also known as the HPG Nascent Protein Assay Kit by Western Blot (WB method), is based on the incorporation of HPG (L-homopropargylglycine), an analog of methionine, during protein synthesis. Subsequently, through a click reaction, HPG is labeled with Biotin-azide, enabling simple, rapid, and highly sensitive detection of newly synthesized proteins via Western blot.

This assay kit allows for quantitative detection of newly synthesized proteins in cultured live cells [1-2]. The detection process requires no radioactive isotopes (such as 35S-methionine) or antibodies, only a straightforward two-step reaction. Newly synthesized proteins, also known as nascent proteins or newly generated proteins, can be quantitatively detected.

HPG (L-homopropargylglycine) is an analog of methionine. HPG substitutes methionine during protein synthesis, and its alkyne group can undergo a copper-catalyzed covalent reaction with fluorescently labeled small molecule azide probes (such as Azide Alexa Fluor 488, Azide Alexa Fluor 555, Azide Alexa Fluor 594, Azide Alexa Fluor 647), forming a stable triazole ring. This highly efficient reaction, termed click reaction, enables the labeling of newly synthesized proteins with corresponding fluorescent probes, detectable using appropriate fluorescence detection equipment.

Features

• The assay kit features simple reaction procedures and high sensitivity. It relies on a straightforward and efficient click reaction, requiring no special temperature conditions, and only a small amount of Biotin Azide to effectively label HPG incorporated into newly synthesized proteins.

• The kit offers convenience and excellent compatibility. Following cell lysis, it facilitates a click reaction with Biotin Azide without generating additional unwanted by-products. It is compatible with subsequent antibody incubation and mass spectrometry detection.

Storage Conditions

Store at -20ºC, valid for one year. Biotin Azide must be stored protected from light.

Precautions

• When preparing the Click Additive solution, properly aliquot and freeze. If a white precipitate forms after dissolving, invert the container several times until fully dissolved before use. If the solution turns brown, the component has become ineffective and should be discarded.
• If you need to use anisomycin as a control, you can order Anisomycin (JNK Activator).
• If you need more HPG, you can order L-Homopropargylglycine (HPG) (≥98%, BioReagent).
• Due to the presence of endogenous methionine in amino acid-containing media and serum, which competitively affects the incorporation of HPG, thus influencing the intensity of Western blot bands, we recommend using DMEM high glucose medium (without methionine) or RPMI 1640 medium (without methionine).
• Since endogenous methionine cannot be removed from animal tissues, this assay kit is not recommended for the detection of animal tissue slice samples.
• This product is for scientific research by professionals only and is not to be used for clinical diagnosis or treatment, food, or drugs. Do not store in a residential area.
• For your safety and health, wear a lab coat and disposable gloves during operation.