Anti-mCherry Nanobody Magnetic Beads – Catalog #B2025472

Anti-mCherry Nanobody Magnetic Beads (Catalog #B2025472) are affinity purification reagents designed for one-step capture and isolation of mCherry-tagged proteins. The beads combine a superparamagnetic core with nanobodies specific to the red fluorescent protein mCherry, enabling efficient immunomagnetic separation directly compatible with standard workflows.

Scientific Overview

mCherry is a bright monomeric red fluorescent protein widely used as an affinity epitope in recombinant protein work. Nanobodies are single-domain antibodies derived from camelids, offering compact size (~15 kDa), high thermal stability, and exceptional binding affinity. Anti-mCherry nanobodies recognize conformational epitopes on the mCherry protein body with Kd values in the nM range. When coupled to magnetic particles, they create a robust affinity matrix for immunomagnetic capture.

Compared to conventional anti-mCherry antibodies, nanobodies offer superior resistance to denaturants and proteolysis, making them ideal for challenging purification conditions. The magnetic particles allow rapid, equipment-free separation: bind protein complexes in minutes, wash away contaminants, and elute the target.

Key applications include:

• One-step affinity purification of mCherry-fusion proteins from cell lysates
• Protein complex co-immunoprecipitation (co-IP)
• Rapid pull-down assays and binding kinetics studies
• Interactome mapping and multi-protein isolation workflows

Usage & Handling Guidance

Equilibrate the beads to room temperature before use. Resuspend gently (do not vortex) and add to your sample or lysis buffer. Incubate at 4°C or room temperature with gentle rotation (minutes to hours, depending on target concentration). Apply a magnet to pellet the beads, remove supernatant, wash 3–5 times with PBS or binding buffer, and elute with low pH or denaturing buffer.

• Binding: Use physiological buffers (PBS, TBS) or mild lysis buffers; extreme pH or high salt may reduce binding.
• Washing: Include detergent (0.1% Triton X-100 or Tween-20) in wash buffer to remove non-specific binding.
• Elution: Low pH (pH 2–3, 0.1 M glycine–HCl), 6 M urea, or SDS (1%) will elute bound protein; choose based on downstream analysis.

What You Get

• 0.1 mL of anti-mCherry nanobody magnetic beads, ready to use
• High-affinity, specific capture of mCherry-tagged proteins
• Fast separation using a standard magnet
• For research use only (RUO)

Why Researchers Choose It

• Nanobodies provide higher affinity and stability than conventional antibodies
• Magnetic format enables hands-on, equipment-free purification
• Rapid protocol suitable for time-sensitive work
• Minimal sample handling reduces loss of protein complexes

Frequently Asked Questions (FAQ)

• What is the Kd of anti-mCherry nanobodies?
  High-affinity anti-mCherry nanobodies typically bind with Kd values in the low nM range, though the exact value may vary by nanobody clone. Contact us for the lot-specific characterization.
• Can I use these beads with fluorescence detection?
  Yes. The magnetic core does not interfere with fluorescence from mCherry or co-expressed tags; however, measure fluorescence before magnet application to avoid quenching by the iron oxide.
• Are these beads suitable for native or denaturing conditions?
  Primarily optimized for native conditions (physiological pH and ionic strength). Nanobodies tolerate moderate denaturants (e.g., 1% SDS) but are best used under conditions that preserve protein-protein interactions.
• How much protein can the beads capture?
  Binding capacity depends on target size and nanobody density. Start with 10–50 µL of bead suspension per 1–5 µg of target; scale up or down based on recovery.
• Can I reuse the beads?
  Beads can be reused after thorough washing and regeneration, though repeated rounds may reduce binding efficiency. For most applications, fresh beads are recommended.
• How do I know if binding was successful?
  Perform a positive control by spiking your sample with a known mCherry-tagged protein, or measure absorbance at 587 nm (mCherry excitation wavelength) before and after binding.

References

• Cong ATQ, Witter TL, Schellenberg MJ. High-efficiency recombinant protein purification using mCherry and YFP nanobody affinity matrices. Protein Sci. 2022;31(9):e4383.
• Liang H, Ma Z, Wang Z, Zhong P, Li R, Jiang H, Zong X, Zhong C, Liu X, Liu P, Liu J, Zhu H, Liu R, Ding Y. Structural Insights into the Binding of Red Fluorescent Protein mCherry-Specific Nanobodies. Int J Mol Sci. 2023;24(8).
• Wang Z, Li L, Hu R, Zhong P, Zhang Y, Cheng S, Jiang H, Liu R, Ding Y. Structural insights into the binding of nanobodies LaM2 and LaM4 to the red fluorescent protein mCherry. Protein Sci. 2021;30(11):2298-2309.
• Fridy PC, Li Y, Keegan S, Thompson MK, Nudelman I, Scheid JF, Oeffinger M, Nussenzweig MC, Fenyö D, Chait BT, Rout MP. A robust pipeline for rapid production of versatile nanobody repertoires. Nat Methods. 2014;11(12):1253-60.