FluoTags®

Fully defined ready-to-go single-domain antibody conjugates

FluoTag® is NanoTag’s trademark for camelid single-domain antibodies conjugated to chemical fluorophores.
All FluoTags® carry a defined number of fluorophores at known positions. Accordingly, they are available as FluoTag®-Q (one fluorophore; Quantitative) or coupled to multiple fluorophores (FluoTag®-X2 or FluoTag®-X4).
Together with their monovalent binding mode, the defined fluorophore conjugation of FluoTags® results in a more quantitative and reliable readout in all fluorescence-based analyses.


Small, Stoichiometric & Reproducible labeling

FluoTag® is NanoTag’s trademark for camelid single-domain antibodies conjugated to chemical fluorophores. Using site-specific coupling chemistry, we ensure a uniform labeling stoichiometry and a defined localization of the fluorophore(s) on the sdAb. After chemical fluorophore attachment, the excess dye is carefully removed from the preparation. These production hallmarks ensure the best possible quality for low background and highest reproducibility. Together with the high-affinity monovalent binding mode of our sdAbs, these features provide a highly linear and reliable signal in all fluorescence-based analyses. In addition, due to the small size of sdAbs, all FluoTag® reagents show much better and more rapid tissue penetration as compared to conventional antibodies. This relates to both, access to crowded cellular environments and fast and deep penetration in 3D tissue sections and whole-mount samples.

Our FluoTags are available in up to three different flavors:

FluoTag®-Q (Quantitative): A single fluorophore reveals target(T)
FluoTag®-X2: Two fluorophores are associated with a single target(T)
FluoTag®-X4 correspond to 2 different sdAbs binding simultaneously to a single target(T), each carrying 2 fluorophores

FluoTag®-Q: One target – One sdAb – One fluorophore (1:1:1)

All FluoTag®-Q reagents are conjugates featuring exactly one nanobody per fluorophore. The fluorophore is placed very near (< 1.5 nm) the antigen-binding surface. 

Features:

  • High affinity monovalent binding + one nanobody per fluorophore – Ideal for epitope counting and highly linear quantitative analyses
  • Small size for optimal access to the target epitope, also in crowded environment or 3D samples.
  • Minimal fluorophore displacement – ideal for super-resolution imaging at highest possible resolution

FluoTag®-X2: One target – One sdAb – Two fluorophores (1:1:2)

FluoTag®-X2 reagents are composed of a single sdAb carrying two fluorophores per protein molecule. The conjugate is therefore up to 2-fold (“X2”) brighter than the corresponding FluoTag®-Q products. The maximal displacement of fluorophores from the antigen-binding surface is ~3 nm. FluoTag®-X2 reagents are only slightly larger than the corresponding FluoTag®-Q products and therefore show similarly good tissue penetration features.

Features:

  • High affinity monovalent binding + two fluorophores per sdAb for brighter signals
  • Reagent is made from small-sized proteins. This ensures optimal access to the respective target protein, also in crowded environment or 3D samples.

Very low fluorophore displacement from the bound target protein – ideal for standard superresolution technologies

FluoTag®-X4: One taget – Two sdAbs – Four fluorophores (1:2:4)

Each FluoTag®-X4 reagent is a blend of two different sdAbs (FluoTag®-X2 and FluoTag®-X2-B) molecules that independently bind the target protein. As each sdAb carries two fluorophores per protein molecule, the blend can direct up to 4 fluorophores to each target protein. The reagent is therefore up to 4-fold (“X4”) brighter than the corresponding FluoTag®-Q products. The maximal displacement of fluorophores from each antigen-binding surface is ~3 nm. FluoTag®-X4 reagents have the same size as the corresponding FluoTag®-X2 products and therefore show similarly good tissue penetration features.

Features:

  • A blend of two high affinity monovalent sdAbs directs four fluorophores to each target protein – ideal for bright signals
  • Small size for optimal access to the respective target protein, also in crowded environment or 3D samples.
  • Very low fluorophore displacement from the bound target protein – ideal for standard super-resolution technologies

Celline
Rapid and efficient affinity selection of target-specific B lymphocytes

NanoTag’s proprietary technology called “Celline” allows for the straightforward discovery of single-domain antibodies by affinity purification of target-specific B lymphocytes from a pool of PBMCs.
Using a single selection step, Celline drastically speeds up sdAb candidate selection while at the same time allowing to set avidity/affinity boundaries.
This method has a proven record for the fast generation of high affinity and low-affinity binders.

The ALFA system
One tag – Three sdAbs – Unlimited applications

A unique, patented technology based on three single-domain antibodies recognizing a rationally designed epitope tag of only 14 amino acids.
Covering a range from pM to nM affinities, this highly specific sdAb ensemble allows for an unmatched spectrum of applications.
High-resolution imaging in life or fixed cells, immobilization and affinity purification of functional proteins and protein complexes, and even therapeutic applications – everything is possible!

Secondary nanobodies
Exploit the full potential of your primary antibodies!

A set of single-domain antibodies recognizing conventional immunoglobulins with exquisite species and isotype specificity is the core of our patented technology.
Our secondary tools provide unique benefits for all antibody-based assays in research and diagnostics by combining reproducible specificity and production with small and high-affinity monovalent binding.
In addition, our technology offers surprising new possibilities like one-step indirect immunofluorescence or easy multiplexing.

FluoTags®
Fully defined ready-to-go single-domain antibody conjugates

FluoTag® is NanoTag’s trademark for camelid single-domain antibodies conjugated to chemical fluorophores.
All FluoTags® carry a defined number of fluorophores at known positions. Accordingly, they are available as FluoTag®-Q (one fluorophore; Quantitative) or coupled to multiple fluorophores (FluoTag®-X2 or FluoTag®-X4).
Together with their monovalent binding mode, the defined fluorophore conjugation of FluoTags® results in a more quantitative and reliable readout in all fluorescence-based analyses.

FX technology
FX technology for reliable post-fixation and expansion microscopy

Our sdAbs can be equipped with fixation (FX) cassettes, short amino acid stretches featuring multiple primary amines.
FX cassettes significantly improve the post-fixation properties of sdAbs, which is beneficial for extended experimental procedures, e.g. STORM and DNA-PAINT microscopy.
In addition, FX cassettes provide multiple anchor points in a ProteinaseK-resistant environment and therefore improve the retention of fluorophores in expansion microscopy (ExM) applications.

Selector resins
Affinity purification made easy!

Selector resins are affinity resins based on controlled immobilization of single-domain antibodies.
For production, high-quality agarose resins are functionalized in-house using propriety chemistry. This results in both a highly specific, stable, and oriented sdAb attachment and an ultra-low non-specific background binding.
The precise and oriented chemistry also ensures optimal functionality and accessibility of the sdAbs and therefore results in beads with a high capacity.

Minibodies
Single-domain antibodies fused to immunoglobulin Fc domains

A fusion of a single-domain antibody to an immunoglobulin Fc domain results in a Minibody. Such divalent recombinant antibodies provide maximal flexibility for technologies based on conventional Fc domains, like, e.g., classical secondary amplification systems.
Minibodies can, however, also be detected using our Secondary nanobodies.

One-step indirect immunofluorescence
One-step indirect immunofluorescence: Combine higher resolution with experimental speed!

Conventional secondary antibodies are multivalent and therefore form large clusters with primary antibodies.
In contrast, our species-specific secondary nanobodes are monovalent binders and therefore form defined stoichiometric complexes with primary antibodies in solution.
This feature efficiently prevents cluster formation. Primary antibodies and secondary nanbodies can therefore be pre-mixed or even co-incubated on the sample and thus enable convenient one-step immunofluorescence or Western assays.

Multiplexing
One-step multi-color detection breaking species boundaries

Due to their monovalent high-affinity binding, NanoTag’s species and isotype-specific secondary nanobodies allow multi-color fluorescent assays using multiple primary antibodies raised in the same species.
Multiplexing thereby drastically expands the possible combinations of primary antibodies and at the same time comes with extra features like a time-saving one-step protocol and an increased imaging resolution.
Discover the new freedom in all multi-color fluorescent assays!

Bi-specific sdAb fusions
Freely combine sdAbs to create new adaptor entities!

Bi-specific sdAb fusions (diabodies) are often used as adaptor molecules bringing two individual target molecules in tight contact.
A fusion of two sdAbs targeting the same molecule at different sites may also be used to increase the apparent binding affinity.
NanoTag has first-hand experience in the design, cloning, and expression of bi-specific sdAb fusions in prokaryotic- and eukaryotic systems and provides ready-to-to vector systems.

Want to learn more on our technologies?