What are VHH antibodies?

Next to conventional IgG antibodies, which are composed of two heavy chains and two light chains, the immune repertoire of camelids such as dromedaries, llamas and alpacas contains so-called heavy chain only IgGs. These antibodies lack the CH1 domain of the heavy chain and are devoid of any light chain1. Therefore, the variable domain of these heavy chain only IgGs, called VHH or Nanobody, represents a stand-alone antigen binding entity and can be produced as a single domain antibody fragment (sdAb).

What makes our technology special?

For more than a decade, ChromoTek generates highly specific and affine VHH antibodies for various applications. By combining our proprietary immunization scheme with phage display2, we are capable to create recombinant sdAbs with tailor-made properties. Using this technique, we get the best of both worlds: Highly specific antibodies that have undergone affinity maturation in vivo, but that are produced recombinantly with known sequence and without any batch-to-batch variations. Typically, affinities are in the picomolar to single-digit nanomolar range. Subsequently, the VHH domains can be further engineered or adapted, depending on the requirements of a certain application.
To date, ChromoTek has generated more than 150 VHH immune libraries.

What are the advantages of VHH antibodies compared to other antibody formats?

Compared to other antibody formats such as IgGs, Fab fragments or scFvs, VHH domains have many unique properties and advantages.
First, with only three CDR loops and a tendency for long CDR3 sequences, VHHs cover a different, yet overlapping, epitope space when compared to conventional IgGs. In general, there seems to be a tendency to bind to native, discontinuous conformational epitopes rather than to linear peptide epitopes. Exceptions are ChromoTek’s anti-Spot-Tag VHH and anti-Myc VHH, which recognize small linear peptide tags, respectively.

VHHs can be produced in a variety of different expression systems, ranging from bacteria via yeast to mammalian cells. Just as conventional IgGs, they can be labelled with a variety of (fluorescent) dyes, biotin or other small molecules. In addition, they can be covalently attached to surfaces, such as agarose beads or other matrices. Also, site-directed functionalization and stoichiometric label attachment is possible.
Due to their small size (approx. 12-15 kDa) and single chain structure, VHHs are suited for applications for which the large size of conventional four peptide chain IgGs (150 kDa) is disadvantageous, such as super-resolution microscopy, tissue penetration or FRET- and ECL-based techniques.
Importantly, VHH domains are perfectly suited to generate bispecific antibody formats, either by genetic fusion or conjugation to other recombinant antibodies. Again, their small size, high stability and good expressability is advantageous for the generation of bi- or even multispecific molecules.
Furthermore, and in contrast to full IgGs, certain VHHs can be genetically expressed intracellularly and bind to their respective intracellular target proteins. When fused to fluorescent proteins such as GFP or RFP, these sdAbs can be used for intracellular visualization of the endogenous target structures in vivo. ChromoTek offers a variety of such VHH-FP fusions, called Chromobodies, e.g. for in vivo visualization of actin, PCNA, the nuclear Lamina and other cytoskeletal or nuclear structures.


1Hamers-Casterman C, Atarhouch T, Muyldermans S, Robinson G, Hamers C, Bajyana Songa E, Bendahman N, Hammers R. Naturally occurring antibodies devoid of light chains. Nature 1993; 363: 446-8.

2Smith GP. Filamentous fusion phage: novel expression vectors that display cloned antigens on the virion surface. Science 1985; 228: 1315-7.