Invitrogen™

MembranePro™ Functional Protein Expression Kit

For customers studying GPCRs and other membrane proteins, MembranePro™ is the ready-to-use system that delivers enriched, functional membrane proteins efficientlyRead more

Catalog Number	Quantity
A11667	10 reactions kit

Catalog number A11667

Price (EUR)

Quantity:

10 reactions kit

For customers studying GPCRs and other membrane proteins, MembranePro™ is the ready-to-use system that delivers enriched, functional membrane proteins efficiently and reliably. Benefits of MembranePro™ include:

• Proteins are displayed on human (or other mammalian) cell membranes
• Cellular quality control and mammalian posttranslational processing helps produce functional protein
• Proteins bud off as lipoparticles and are easily collected from the culture medium
• Lipoparticles are enriched with expressed receptors

MembranePro™ workflow is less labor intensive in comparison to traditional production of membrane fractions. Briefly, the MembranePro™ protocol allows you to:

1. Clone your gene of interest using a TOPO™ vector
2. Transfect with Lipofectamine™ 2000 and MembranePro™ Reagent into 293FT cells
3. Harvest lipoparticles 48h post-transfection, concentrate with MembranePro™ Precipitation Mix
4. Store lipoparticles at -80 C or proceed to downstream assays

MembranePro™ products are offered in two configurations – as an expression kit (TOPO™ vector, Lipofectamine™ 2000, MembranePro™ Precipitation Mix, MembranePro™ Reagent, 293FT cells) and as a support kit (Lipofectamine™ 2000, MembranePro™ Precipitation Mix, MembranePro™ Reagent). The expression kit is only available in a 10 reaction size; the support kit is available in 10, 60 and 600 reaction sizes.

For Research Use Only. Not for use in diagnostic procedures.

Specifications

Cell Line293FT

Delivery TypeVirus-Like Particles (VLPs)

Expressed Protein TypeMembrane Protein

Expression MechanismCell-Based Expression

For Use With (Application)Membrane Protein Expression

High-throughput CompatibilityHigh-throughput Compatible

Product LineMembranePro

Quantity10 reactions kit

Selection Agent (Eukaryotic)Blasticidin

Selection Marker EukaryoticBsdR

Expression SystemMammalian

PromoterEF-1α

TypeMembranePro Functional Protein Expression Kit

Unit SizeEach

Contents & Storage

The kit contains:
•pEF6 V5-His TOPO™ TA Vector Kit for at least 10 TOPO™ cloning reactions (Store at -20°C)
•3 x10^6 293FT Cells (Store in liquid nitrogen)
•1 x 75 ml MembranePro™ Precipitation Mix (Store at room temperature)
•1 x 0.3 mg (1 mg⁄ml) MembranePro™ Reagent (Store at -20°C)
•3 x 0.75 ml Lipofectamine™ 2000 (Store at +4°C)
•Manuals

Frequently asked questions (FAQs)

I had a problem growing up the 293T cells provided in the MembranePro Functional Protein Expression Kit. Can I use a different cell line?

While other cell lines can be tested, we recommend using the supplied 293FT cells, as the performance of the kit has been optimized for use with this cell line (i.e., transfection efficiency).

Find additional tips, troubleshooting help, and resources within our Protein Expression Support Center.

Your MembranePro Functional Protein Expression Kit/Expi293 MembranePro Expression System does not include a positive control. How do I tell if MembranePro particles were formed?

You can visually tell if MembranePro particles formed via a pellet at the bottom of the tube following precipitation. You can also test the function of your MembranePro particles via receptor-ligand binding studies.

Find additional tips, troubleshooting help, and resources within our Protein Expression Support Center.

A GPCR has 2 polypeptides - a ligand-binding polypeptide (receptor chain) and the G-protein (signal transduction chain). Can I use a G-protein coupling receptor (RG-form) specific ligand as a tracer?

If a ligand-binding chain is transfected alone, there may not be coupling of G-protein. We have demonstrated agonist ligand binding to 5HT1a and muscarinic M1 receptors captured on MembranePro particles. Different agonists compete ligand off both muscarinic and 5HT1a receptors in MembranePro particles, and they do so in order of affinity, indicating physiologically relevant interaction with the receptors on MembranePro particles. In direct binding studies, agonist binds in the low-affinity form (higher Kd), indicating little or no G-protein is coupled to the receptor. However, the binding is still specific and competitive. You may see increased binding by coexpressing the cognate G-protein.

Find additional tips, troubleshooting help, and resources within our Protein Expression Support Center.

What is the size range of proteins produced with the MembranePro Functional Protein Expression Kit and the Expi293 MembranePro Expression System?

The GPCRs we produced and tested are approximately 50 kDa. In theory, there is no limit on the size of the extracellular or transmembrane domains. However, there may be packaging issues if the C-terminal domain is too large.

Find additional tips, troubleshooting help, and resources within our Protein Expression Support Center.

Does the expression vector in the MembranePro Functional Protein Expression Kit offer a His tag?

The MembranePro Functional Protein Expression System is optimized for use with the pEF6 vector. However, cloning your gene into the pEF6/V5-His-TOPO vector without a stop codon and in frame with the polylinker will result in a fusion protein with V5 and polyhistidine (6×His) tags on the C-terminus of your protein. As the C-terminus of your transmembrane protein will likely be inside the VLP, these tags will be inaccessible to purification resins and antibodies. In theory, these tags could be used to identify and isolate a fusion membrane protein after denaturing the VLP; however, the MembranePro Functional Protein Expression System does not support using the tags for extraction and purification.

Find additional tips, troubleshooting help, and resources within our Protein Expression Support Center.

View all MembranePro™ Functional Protein Expression Kit FAQs