Nano-CaptureLigand mouse IgG2a

BLI binding kinetics of a mouse IgG2a anti-V5-tag antibody to a V5-tagged protein. A monoclonal mouse IgG2a anti-V5-tag antibody was immobilized using Nano-CaptureLigand mouse IgG2a, Fc-specific VHH, biotinylated on FortéBio Streptavidin (SA) Biosensors and assayed with different concentrations of a V5-tagged protein.

Immobilization of the Nano-CaptureLigand (green) on Biosensor (white) for biolayer interferometry (BLI) followed by capture of mouse IgG2a antibody (orange). The setup is ready for characterization of mouse IgG2a antibodies’ binding kinetics and affinity to substrate (red), mouse IgG2a antibody discovery and screening in crude liquids like hybridoma supernatant, serum, and plasma.

ELISA capture of mouse IgG2a antibody using Nano-CaptureLigand mouse IgG2a, Fc-specific VHH, biotinylated. 50 nM Nano-CaptureLigand mouse IgG2a, Fc-specific VHH, biotinylated was used for coating on an avidin-coated MaxiSorp plate. Mouse IgG2a antibody was titrated in a 1:2 dilution series and detected with an alkaline phosphatase-conjugated detection antibody.

Sandwich ELISA: Immobilization of the Nano-CaptureLigand on streptavidin coated microplates for ELISA. No biotinylation of the antibody is required. Coating of the streptavidin microplate with Nano-CaptureLigand (top left). Site-directed antibody immobilization via Nano-CaptureLigand (top right). Addition of sample including target (red, bottom left). Addition of detection and secondary antibody for read-out (bottom right).

Description
The Nano-CaptureLigand mouse IgG2a captures and immobilizes mouse IgG2a antibodies and Fc-fragments to avidin/streptavidin coated surfaces used in BLI, SPR, and ELISA assays. It consists of a biotinylated Nanobody that specifically binds to the mouse IgG2a subclass with high affinity and in a site-directed manner.

Specificity: Fc-fragment of mouse IgG2a antibodies
Does not bind to: Goat, human, macaque, and sheep serum and to mouse IgG1, IgG2b, and IgG3
Cross-reactivity to mouse IgG2c
Nano-CaptureLigands specificity overview

Conjugates: Biotin

Applications
Mouse IgG2a capture to biosensors used in BLI and SPR
Mouse IgG2a immobilization for ELISA

Advanced immobilization of antibodies
Specific, stable, and site-directed immobilization of non-biotinylated mouse IgG2a antibodies to streptavidin/avidin

Product	Size	Code	Price	Buy
Product Nano-CaptureLigand™ mouse IgG2a, Fc-specific VHH, biotinylated	Size 10 µL	Code smsG2aB-1-10	Price $ 130	Buy +
Product Nano-CaptureLigand™ mouse IgG2a, Fc-specific VHH, biotinylated	Size 100 µL	Code smsG2aB-1-100	Price $ 480	Buy +

Product Type
Capture Nanobody (V_HH)

Applications
Immobilization of mouse IgG2a antibodies to avidin and streptavidin surfaces for Bio-Layer Interferometry (BLI), Surface Plasmon Resonance (SPR), and ELISA

Target/ Specificity
Fc-fragment of mouse IgG2a

Cross-reactivity
No cross-reactivity to mouse IgG1, Ig2b, IgG3; human IgG1, IgG2, IgG3, IgG4, IgA; rat IgG1, IgG2a, IgG2b; sera from goat, human, macaque (cynomolgus monkey), sheep
Cross-reactivity to mouse IgG2c

Affinity (Kd) of monovalent (1:1) binding mode
1.5 nM
Apparent affinity may be higher for full IgGs due to avidity effects (1 antibody captured by 2 Nanobodies).

Concentration
1 g/L (68 μM)

Conjugate
Biotin

Degree of biotinylation
On average 1-2 biotin molecules per Nanobody

Format
Alpaca single domain antibody, monovalent

Host
Alpaca-derived, recombinantly produced in bacteria

Clonality
Monoclonal

Clone
CTK0108 (V_HH0249)

How to cite this product
Nano-CaptureLigand™ mouse IgG1, Fc-specific V_HH, biotinylated
RRID: AB_2848188

Synonyms
Alpaca single domain antibody, V_HH, Nanobody, binding domain of single domain antibody, Nano-antibody

Validation
Application validated for ELISA and BLI (FortéBio Octet^® systems)
Determination of cross-reactivity, subclass specificity, sequence, affinity, and melting temperature

Purity
Recombinantly expressed and purified via His-tag

Form
Buffered aqueous solution

Storage buffer
25 mM TAPS pH 8.5, 500 mM NaCl, 5 mM EDTA, Preservative: 0.09 % sodium azide

Storage conditions
Upon receipt store at +4°C/+40°F. Optional: Aliquot upon arrival and store at -20°C/-4°F

Stability
Stable for 1 year at +4°C/+40°F

Shipment
Shipped at ambient temperature

Protocol
Protocol Nano-CaptureLigand mouse IgG2a, Fc-specific VHH, biotinylated (PDF)

SDS
SDS Nano-CaptureLigand mouse IgG2a, Fc-specific VHH, biotinylated (PDF)

Nano-CaptureLigand Specificity

Nano-CaptureLigands specificity overview (PDF)

Application note

Nano-CaptureLigands for COVID-19 antibodies capture in BLI binding kinetics assay (PDF)

Comparison of immobilization methods

Comparison of BLI/SPR immobilization methods (PDF)

Comparison of ELISA immobilization methods (PDF)

Applications

Detection and quantitation of mouse IgG2a antibodies and their antigens
Characterization of mouse IgG2a antibodies’ binding kinetics and affinity
Mouse IgG2a antibody discovery and screening in crude liquids like hybridoma supernatant, serum, and plasma
Epitope binning

The next level of antibody immobilization to BLI and SPR biosensors

Gentle, site-directed and Fc-specific immobilization of mouse IgG2a antibodies and Fc-fragments to streptavidin coated biosensors without biotinylation of the antibody
Compatible to surface plasmon resonance (SPR) and biolayer interferometry (BLI) assays (e.g. FortéBio Octet^® systems)
High affinity for stable baseline without significant dissociation
>10x regeneration for multiple reuse
High specificity enables mouse IgG2a antibody discovery and screening in crude liquids like hybridoma supernatant, serum, and plasma samples

Comparison of BLI/SPR immobilization methods

The next level of antibody immobilization for ELISA

Compatible to enzyme-linked immunosorbent assays (ELISA)
Site-directed immobilization of mouse IgG2a capture antibodies to streptavidin/avidin coated surfaces of sandwich ELISAs without biotinylation of antibody
Combine the site-specificity of protein A, G, and L with the stability of the biotin/streptavidin interaction
Mouse IgG2a subclass specificity allows the application of a broad range of detection antibodies
High affinity for strong immobilization of mouse IgG2a antibodies even at low antibody concentrations

Comparison of ELISA immobilization methods

Nano-CaptureLigands are comprehensively validated

Characterized: DNA and protein sequences, affinity, melting temperature
Recombinantly produced, virtually without lot-to-lot variation
Manufacturing process is animal free

Only for research applications, not for diagnostic or therapeutic use!