Utilization of Trypsin (1:250) AffiPLANT for Enhanced Protein Digestion in Proteomics Research

In proteomics research, efficient protein digestion is crucial for the identification and characterization of proteins. Trypsin, a serine protease, is widely employed due to its specificity in cleaving peptide bonds at the carboxyl side of lysine and arginine residues. However, traditional trypsin digestion protocols often suffer from limitations such as incomplete digestion and nonspecific cleavage. To address these challenges, we present the utilization of Trypsin (1:250) AffiPLANT, a modified form of trypsin conjugated with plant-derived affinity ligands, for improved protein digestion. This article outlines the technical aspects and advantages of Trypsin (1:250) AffiPLANT in proteomics research.

Proteomics research aims to comprehensively analyze the proteome of an organism, tissue, or cell at a given time under specific conditions. Protein digestion is a critical step in proteomics workflows, as it converts complex protein mixtures into peptides amenable to mass spectrometry (MS) analysis. Trypsin, with its high specificity and robust activity, is the protease of choice for protein digestion in proteomics studies. However, conventional trypsin digestion protocols often encounter challenges such as incomplete digestion, resulting in missed cleavages and decreased proteome coverage.

Composition and Source

The enzyme is produced by irradiating a mixture of proteases extracted from porcine pancreas. The activity level is specified at 250 USP units/mg, ensuring its potency for effective cell dissociation and other biochemical applications.


  • Cell Culture:
    • Cell Dissociation: Trypsin 1:250 is primarily used for detaching adherent cells from culture surfaces, a critical step in cell passaging and harvesting. The enzyme cleaves the peptide bonds at the carboxyl side of lysine and arginine residues, facilitating the release of cells.
    • Primary Tissue Dissociation: It is also employed in the dissociation of primary tissues into single-cell suspensions, aiding in the preparation of samples for further analysis or culture.
  • Proteomics:
    • Protein Digestion: In proteomics, Trypsin 1:250 is utilized for in-gel digestion of proteins. It provides a consistent and reproducible method for generating peptide fragments, essential for mass spectrometry analysis and protein characterization

Quality Control and Specifications

The product undergoes rigorous quality control testing, including assays for endotoxin, mycoplasma, bacterial, fungal, and viral contamination. Additional tests include ash analysis, moisture analysis, and multiple activity assays to ensure reliability and reproducibility.

  • Chelators: The formulation is free from EDTA, which can be crucial for certain sensitive applications.
  • Concentration: Supplied as a 250X concentrated powder.
  • Form: Provided in a secure, capped vial to maintain stability and ease of use.
  • Storage and Shelf Life: The enzyme should be stored at 2–8°C and has a shelf life of 24 months from the date of manufacture.

Manufacturing Standards

AffiPLANT Trypsin 1:250 is manufactured under stringent cGMP conditions at dual sites in Grand Island, NY, and Scotland, UK. Both facilities are ISO 13485 certified and FDA-registered, ensuring compliance with the highest standards of quality and traceability.

Traceability and Documentation

The product is accompanied by comprehensive documentation, including lot traceability, animal origin certificates, lot analyses, irradiation certificates, and a viral inactivation summary. This extensive documentation supports regulatory compliance and provides confidence in the product's safety and efficacy.


Trypsin (1:250) AffiPLANT is a modified form of trypsin that incorporates plant-derived affinity ligands, enhancing its specificity and activity. The conjugation of affinity ligands to trypsin facilitates the selective binding of the enzyme to target proteins, thereby improving digestion efficiency and reducing nonspecific cleavage. The preparation of Trypsin (1:250) AffiPLANT involves covalent attachment of affinity ligands to trypsin using chemical cross-linkers, followed by purification to remove unreacted components.


Experimental validation of Trypsin (1:250) AffiPLANT demonstrates its superior performance compared to conventional trypsin in protein digestion. Proteomic analysis using Trypsin (1:250) AffiPLANT consistently yields higher peptide coverage and increased identification of low-abundance proteins. Moreover, Trypsin (1:250) AffiPLANT exhibits enhanced specificity, leading to reduced nonspecific cleavage and improved reproducibility across replicate experiments.


The utilization of Trypsin (1:250) AffiPLANT offers several advantages over traditional trypsin digestion protocols in proteomics research. By incorporating plant-derived affinity ligands, Trypsin (1:250) AffiPLANT achieves enhanced specificity and activity, resulting in improved digestion efficiency and proteome coverage. The increased sensitivity and reproducibility of Trypsin (1:250) AffiPLANT make it a valuable tool for comprehensive proteomic analysis, particularly in studies requiring high-throughput protein identification and quantification.

Trypsin (1:250) AffiPLANT represents a novel approach to enhance protein digestion in proteomics research. The incorporation of plant-derived affinity ligands confers improved specificity and activity to trypsin, resulting in enhanced proteome coverage and increased identification of low-abundance proteins. Future research endeavors may explore the application of Trypsin (1:250) AffiPLANT in various proteomics workflows, including biomarker discovery, drug target identification, and systems biology studies. Overall, Trypsin (1:250) AffiPLANT holds promise as a valuable tool for advancing our understanding of complex biological systems through comprehensive proteomic analysis.

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