Recombinant Protein Information

What is a recombinant protein?

A recombinant protein is a manufactured protein that has been produced using cells engineered to carry a gene of interest. Recombinant proteins can have the same amino acid sequence as the corresponding naturally occurring (wild-type) protein, or have their sequence modified for improved properties such as solubility or production yield.

What are recombinant proteins used for?

There are myriad applications for recombinants proteins in biomedical research, as well as diagnostics and therapeutics. Recombinant proteins have become an important tool in various areas of science. Laboratory uses for recombinant proteins include:

Cell culture supplements, including growth factors for primary cell expansion, differentiation,or stem cell reprogramming
Generation of organoids and other 3D cell cultures for disease modeling and drug discovery
Expansion of CAR-T cells by stimulation with recombinant cytokines
Investigational tools for probing cellular function and cell signaling pathways
Antigens for immunization and production of antibodies
Components of functional cell-based assays such as cell migration and angiogenesis

Biologically active recombinant proteins

The above applications require that the recombinant protein be biologically active; that is, the protein is recognized by cells and cause changes in those cells. Bioactivity can be measured in an appropriate cell-based assay and quantified as a specific activity or EC50.

Look up international units for PeproTech proteins
Search catalog of bioactive recombinant proteins

Non-bioactive recombinant proteins may be used as standards in protein analysis studies including immunoassays such as western blotting, ELISA, gel shift, and immunohistochemistry.

Types of recombinant proteins

Many biologically important classes of recombinant proteins are available commercially, obviating the need to express, purify, and validate commonly used bioactive recombinant protein(s).

Major types of recombinant proteins include:

Growth factors: a group of intercellular signaling proteins that cells produce and use to communicate. They stimulate a variety of cellular processes including cell division, differentiation, migration and survival. Prominent growth factor families include:
- FGF: fibroblast growth factors
  - Learn more about the FGF family
  - Explore FGF signaling pathways
- VEGF: vascular endothelial growth factors
  - Learn more about VEGF
  - Explore the VEGF signaling pathway
- Neurotrophins: a subset of growth factors that act primarily on cells involved in the brain and nervous system
Cytokines: including interferons, interleukins, pro-inflammatory cytokines, and chemokines critical to immune cell function and communication
Enzymes: bioactive proteins that can perform chemical transformations; includes recombinant proteases, kinases, and nucleases

Learn more about chemokine structure and function
Explore cytokine and growth factor pathways

Recombinant protein manufacturing categories

When choosing a recombinant protein for a specific application, it helps to know which levels of quality control and documentation are required.

Research use only (RUO)

Research-grade recombinant proteins are suitable for a wide variety of life science research applications including development of cell models, maintaining cell phenotypes, and in vitro assays.

See specifications for Gibco PeproTech RUO proteins

Animal-free

As an additional safeguard against animal pathogen contamination, animal-free recombinant proteins are made using raw materials that are not of animal origin.

Learn more about PeproTech animal free recombinant proteins

GMP recombinant proteins

GMP-manufactured recombinant proteins undergo extensive testing and are accompanied by documentation to support projects/applications that require this level of rigor.

Learn more about PeproGMP recombinant proteins

FAQs about recombinant proteins

How is the biological activity of a recombinant protein determined?

Bioassays are intended to measure the biological activity of a given growth factor or cytokine. In most of the cases, the bioassays are cell-based tests using different indicator cells such as primary cells or cell lines. Commonly used bioassays include cell proliferation assay, chemotaxis assay, cytokine production assay, and cytotoxicity assay. The biological activity of a given cytokine is expressed as ED₅₀, which represents the concentration of the cytokine that induces 50% of the maximum response.

What is the relationship between the specific activity expressed as an ED50 and as units/mg?

The specific activity of a biologically active protein can be determined using the following equation:
1 x 10⁶/ ED₅₀ (ng/mL) = specific activity (units/mg)

The ED₅₀ (AKA EC₅₀) can be found on the CoA for the recombinant protein, but we advise determining the ED₅₀ of a given recombinant protein in your functional assay system.

For additional information on ED₅₀ and its relationship with specific activity, please refer to our Tech Tip.

How should recombinant proteins be reconstituted?

First, centrifuge the container to concentrate the powder at the bottom of the tube. In general, we recommend reconstitution to a concentration of 0.1 to 1.0 mg/mL. Most proteins can be reconstituted with the addition of sterile, distilled water. However, the product data sheet or CoA will indicate when a diluent other than water is required. Recommended solutions, carrier protein concentrations and extended storage conditions can also be found on these documents.

How should recombinant proteins be stored?

In general, we recommend that lyophilized recombinant proteins be stored at –20°C, but short-term storage at 4°C or room temperature is permissible. For reconstituted protein solutions, we recommend that you make working aliquots containing at least 10 μL of protein solution and store at –20°C to –80°C. Do not allow multiple freeze-thaw cycles. Product-specific instructions for storage of lyophilized and reconstituted recombinant proteins are provided on either the product manual, data sheet, or certificate of analysis (CoA).

Why can’t I see the protein pellet in the vial for my lyophilized recombinant protein?

Recombinant proteins are usually formulated without carrier proteins or additives (e.g., BSA, HSA, sucrose, etc.). As a result, during lyophilization the protein product may be deposited on the vial as a thin, and sometimes invisible, film instead of a pellet. The size of the pellet, if any, is not directly related to the quantity of the recombinant protein in the vial.

To ensure complete recovery of protein product, before opening a vial of lyophilized recombinant protein, we recommend centrifuging it for 20–30 seconds to drive any protein that may be lodged in the cap or on the side to the bottom of the vial. After reconstitution, you can confirm the presence of product protein by running a small amount on SDS-PAGE. In general, a protein band with expected size should be visible with as little as 10 ng of protein loaded on an acrylamide gel.

See all recombinant protein FAQs

Learn more about recombinant proteins and their applications

General

Technical note: Converting ED50 (ng/mL) to specific activity (units/mg)

3D cell culture

Application note: Generation of MCF7 spheroids in serum-free conditions
Peer-reviewed methods for organoid generation with recombinant growth factors
Flyer: Recombinant proteins for organoid research

Immunology

Application note: Integrated generation and characterization of CAR-T cells
Protocol: In vitro differentiation of macrophages from monocytes via M-CSF

Neurobiology

Application note: Enhanced proliferation of primary NSCs and sustained differentiation into precursors using heat-stable bFGF
Protocol: Differentiating neural stem cells into neurons and glial cells

Stem cells

Flyer: Recombinant cytokines and growth factors for differentiation of embryonic and induced pluripotent stem cells

Recombinant proteins learning center

Learn what recombinant proteins are, what they are used for, and what types are available

About PeproTech proteins

Learn about the PeproTech brand of recombinant proteins

Gibco cell culture basics

Learn about all aspects of cell culture

3D cell culture protocols

See which recombinant proteins are used to generate organoids

Western transfer protocols and recipes

Procedures and buffers for performing western blots

Cytokine and growth factor signaling pathways

Explore the role of proteins and cytokines in various signaling pathways

Support

Recombinant Proteins Support Center

Find Getting Started guides, protocols, troubleshooting, and other resources

Recombinant proteins FAQs

Search the database to find answers to your questions

Contact general support

Get technical or customer support for any recombinant protein; find SDS, CoA, and phone numbers based on a specific product

Contact GMP support

Get in touch with GMP technical and regulatory experts

Recombinant Protein Information - US (2024)