MonoRab™ Anti-VHH Antibodies

Product List

• Anti-Camelid VHH, mAb

  Cat. No.	Product Name	Quantity	Price	Order
  A01860	MonoRab™ Rabbit Anti-Camelid VHH Antibody, mAb		$215.00 200 μl
  A01861	MonoRab™ Rabbit Anti-Camelid VHH Antibody [HRP], mAb		$215.00 200 μl
  A01862	MonoRab™ Rabbit Anti-Camelid VHH Antibody [iFluor488], mAb		$215.00 200 μl
  A01863	MonoRab™ Rabbit Anti-Camelid VHH Antibody [iFluor555], mAb		$215.00 200 μl
  A01994	MonoRab™ Rabbit Anti-Camelid VHH Antibody [iFluor 647], mAb		$215.00 200 μl
  A01995	MonoRab™ Rabbit Anti-Camelid VHH Antibody [Biotin], mAb		$215.00 200 μl
  A02226	MonoRab™ Rabbit Anti-Camelid VHH Antibody [FITC], mAb		$215.00 200 μl
  A02227	MonoRab™ Rabbit Anti-Camelid VHH Antibody [PE], mAb		$215.00 200 μl

• Anti-Humanized VHH, mAb

  Cat. No.	Product Name	Quantity	Price	Order
  A02165	MonoRab™ Rabbit Anti-Humanized VHH Antibody, mAb		$215.00 200 μl
  A02166	MonoRab™ Rabbit Anti-Humanized VHH Antibody [Biotin], mAb		$215.00 200 μl
  A02167	MonoRab™ Rabbit Anti-Humanized VHH Antibody [HRP], mAb		$215.00 200 μl
  A02168	MonoRab™ Rabbit Anti-Humanized VHH Antibody [iFluor 488], mAb		$215.00 200 μl
  A02169	MonoRab™ Rabbit Anti-Humanized VHH Antibody [iFluor 555], mAb		$215.00 200 μl
  A02170	MonoRab™ Rabbit Anti-Humanized VHH Antibody [iFluor 647], mAb		$215.00 200 μl
  A02171	MonoRab™ Rabbit Anti-Humanized VHH Antibody [PE], mAb		$215.00 200 μl
  A02172	MonoRab™ Rabbit Anti-Humanized VHH Antibody [FITC], mAb		$215.00 200 μl

• Anti-Camelid VHH, mAb Cocktail

  Cat. No.	Product Name	Quantity	Price	Order
  A02014	MonoRab™ Rabbit Anti-Camelid VHH Cocktail		$385.00 200 μl
  A02015	MonoRab™ Rabbit Anti-Camelid VHH Cocktail [Biotin]		$385.00 200 μl
  A02016	MonoRab™ Rabbit Anti-Camelid VHH Cocktail [HRP]		$385.00 200 μl
  A02017	MonoRab™ Rabbit Anti-Camelid VHH Cocktail [FITC]		$385.00 200 μl
  A02018	MonoRab™ Rabbit Anti-Camelid VHH Cocktail [PE]		$385.00 200 μl
  A02019	MonoRab™ Rabbit Anti-Camelid VHH Cocktail [iFluor 647]		$385.00 200 μl
  A02020	MonoRab™ Rabbit Anti-Camelid VHH Cocktail [iFluor 555]		$385.00 200 μl
  A02021	MonoRab™ Rabbit Anti-Camelid VHH Cocktail [iFluor 488]		$385.00 200 μl

Selection Guide

• Binding Compatibility with Camelid VHHs
  

  Figure 1. Binding compatibility comparison of MonoRab™ Rabbit Anti-Camelid VHH Antibody, mAb (Cat. No. A01860) and MonoRab™ Rabbit Anti-Camelid VHH Cocktail (Cat. No. A02014) with camelid VHHs. The cocktail antibody (Cat. No. A02014) is broadly reactive, and is ideal to be used for VHH antibody screening. In most cases, the monoclonal antibody (Cat. No. A01860) exhibits stronger affinity because it has low cross-reactivity. For labeling applications, or to minimize background, use A01860.

• Binding Compatibility with Humanized VHHs

  In therapeutic applications, camelid VHH needs to be humanized to reduce the risk of immunogenicity and anti-drug antibody (ADA) development in humans. As a result of humanization, some anti-camelid VHH antibodies can not recognize humanized VHH. GenScript provides antibodies that recognize humanized VHH.

  

  Figure 2. Binding compatibility comparison of MonoRab™ Rabbit Anti-Humanized VHH Antibody, mAb (Cat. No. A02165), MonoRab™ Rabbit Anti-Camelid VHH Antibody, mAb (Cat. No. A01860), and MonoRab™ Rabbit Anti-Camelid VHH Cocktail (Cat. No. A02014) with humanized VHHs. The performance of A02165 is the best. It is ideal to be used for humanized VHH antibody detection.

Applications

• Detection of VHH-based Jurkat-CAR cells by Flow Cytometry

  Due to their small size and high specificity, recombinant VHH constructs are increasingly being used in therapeutic applications, such as CAR-T therapy. To track VHH in patients, animal models, and cell cultures, specific anti-VHH antibodies are required. Techniques such as flow cytometry can then be used to study the modified T cells, and cells can be sorted and used for downstream applications.

  GenScript MonoRab™ Rabbit Anti-VHH antibodies show ultra-high specificity for VHH-based CAR T cells. They are able to accurately detect the amount of VHH-based CAR T cells in samples, as shown in Figure 3. The MonoRab™ Rabbit Anti-VHH antibodies can also be used in CAR-T cell determination and sorting (Figure 4) by magnetic-activated cell sorting (MACS).

  

  Figure 3. The above data shows FACS binding of a series of dilutions of MonoRab™ Rabbit Anti-Camelid VHH Cocktail [PE] (Cat. No. A02018) with Jurkat cells and samples containing 20% of VHH-based Jurkat-CAR cells. With the development of CAR-T therapy, VHH is often used in CAR generation. A02018 shows excellent binding activity on VHH-based Jurkat-CAR cells. It is used for CAR-T cell verification and determination.

  

  Figure 4. VHH-based CAR-T cells were sorted by MACS with MonoRab™ Rabbit Anti-Camelid VHH Cocktail [Biotin] (Cat. No. A02015) and anti-Biotin magnetic nanobeads (Miltenyi) where naïve T cells served as negative control. Then, the CAR-T cells were stained with MonoRab™ Rabbit Anti-Camelid VHH Cocktail [iFluor 555] (Cat. No. A02020) and analyzed by FACS.

  As shown in the figure, the ratio of positive CAR-T cells grows form about 10% to 90% after MACS cell separation. A02015 is ideal to be used for CAR-T cell MACS separation.

• Detection of infiltrated VHH-transduced CAR-T cells in tissue by immunofluorescence (IF) and immunohistochemistry (IHC)

  While CAR-T cell therapies have had success in eradicating some types of cancer, such as advanced leukemia, their efficacy in treating solid tumors is limited. The main reason for that is failure of CAR-T cell infiltration through the physical barriers of solid tumors. To optimize CAR-T cells for greater efficacy, it is important to detect the distribution of CAR-T cells. Additionally, the distribution of CAR-T cells provides necessary information for predicting possible adverse reactions. Non-specific expression of tumor antigen in normal tissue increases the possibility of injury on normal tissue caused by CAR-T cells.

  GenScript MonoRab™ Anti-Camelid VHH Cocktail can specifically bind with VHH-transduced CAR-T cells infiltrated in the NCG mouse spleen (Figure 5) and PDX mouse liver (Figure 6) by immunofluorescence and immunohistochemical analysis, which is helpful in detecting the tissue distribution of CAR-T cells.

  

  Figure 5. Immunofluorescence analysis of VHH-transduced CAR-T cells infiltrated NCG mouse spleen, VHH was labelled with MonoRab™ Rabbit Anti-Camelid VHH Cocktail (Cat. No. A02014) at 0.5 µg/mL, and anti-rabbit IgG (H+L) (Alexa Fluor 488) was used as secondary antibody at 1/500 (4 μg/mL) (Green). APC hCD45 monoclonal antibody was used to define all infused T cells at 1/50 (0.5 μg/mL)(Red). Confocal images showed cytoplasmic staining in infiltrated VHH-transduced CAR-T cells in the NCG mouse spleen. The untransduced infiltrated T cells showed a negative signal with A02014.

  VHH-CART infiltrated liver

  

  Figure 6. Immunohistochemical analysis of VHH-transduced CAR-T cells infiltrated liver tissue of PDX mouse. VHH was labelled with MonoRab™ Rabbit Anti-Camelid VHH Cocktail (Cat. No. A02014) at 1 μg/mL, followed by incubation of biotinylated anti-rabbit IgG (H+L) (2.5 μg/mL). The final signal was amplified by ABC (Avidin-Biotin Complex) Kit. Positive staining was shown in the membrane or cytoplasm of VHH-transduced CAR-T cells infiltrated into liver tissue. Counterstained with hematoxylin.

• VHH antibodies screening or titer determination

  Using an anti Nanobody^® secondary antibody for direct detection, rather than by adding a tag to the nanobody, can provide advantages in many applications. For example, in therapeutic applications, most pharmaceutical companies do not add tag proteins like FLAG^® or His to nanobodies to avoid adding more risk factors. Some researchers may design Fc-fused nanobodies to aid detection and extend the half-life. However, Fc-fused nanobodies would lose their small size advantage and trigger non-specific binding (many cells have Fc receptors) as well as Fc-mediated complement reactions. Besides, anti-VHH antibody can reduce cross-reactivity with endogenous proteins that are homologous to some tag epitopes.

  GenScript MonoRab™ Rabbit Anti-VHH antibodies specifically recognize heavy chain antibodies but not the conventional IgG1 type (Figure 7) found in camelid sera and have low cross-reactivity among other species (Figure 8). As a result, GenScript MonoRab™ Rabbit Anti-VHH antibodies can be used as an anti llama secondary antibody or anti alpaca secondary antibody and accurately detect the amount of VHH in a sample.

  

  Figure 7. MonoRab™ Rabbit Anti-Camelid VHH Antibody, mAb (Cat. No. A01860) and MonoRab™ Rabbit Anti-Camelid VHH Cocktail (Cat. No. A02014) are specific to the variable domain of camelid heavy chain antibodies (IgG2&3).

  

  Figure 8. MonoRab™ Rabbit Anti-Camelid VHH Antibody, mAb (Cat. No. A01860) and MonoRab™ Rabbit Anti-Camelid VHH Cocktail (Cat. No. A02014) are specific to Camelid IgG, while MonoRab™ Rabbit Anti-Humanized VHH Antibody, mAb (Cat. No.A02165) is specific to Humanized VHH. All these three antibodies have no cross-reactivity with mouse, rat, rabbit, goat, and human immunoglobulins, so that they can be used for the detection of different species of blood samples.

• VHH antibodies titer determination

  During the development of VHH drugs, researchers need to study the pharmacokinetics (PK) of the drug, which requires titration of VHH. GenScript MonoRab™ Rabbit Anti-VHH antibodies can detect VHH titers in a sandwich format with high sensitivity (Figure 9), making them ideal for use in blood drug analysis.

  

  Figure 9. Standard curve of VHH Sandwich ELISA. The VHH Sandwich ELISA assay is developed using MonoRab™ Rabbit Anti-Camelid VHH Antibody, mAb (Cat. No. A01860) and MonoRab™ Rabbit Anti-Camelid VHH Cocktail [HRP] (Cat. No. A02016) as the capture and detection antibodies, respectively.

  In this ELISA assay, the detection antibody was labeled with horseradish peroxidase (HRP)*. The sensitivity of detecting VHH is up to 39 pg/mL.

  * If your assay needs an unconjugated antibody, GenScript’s MonoRab™ Rabbit Anti-Camelid VHH Cocktail is the ideal choice (Cat. No. A02014). GenScript provides customized conjugation services for this product per the customer's request.

• Enrichment of heavy chain antibody-expressing B cells in peripheral blood mononuclear cells (PBMC)

  In the development of engineered nanobodies, conventional IgG1 and heavy chain antibodies (IgG2&3) are produced in the immunized animals after immunization of the target protein to alpacas or llamas. Distinguishing heavy chain antibody specific-PBMCs from total PBMCs can exclude interference from non-VHH sequences and optimize library construction and b-cell cloning process. Thus an anti alpaca IgG2&3 antibody or anti llama IgG2&3 antibody would be very important.

  GenScript MonoRab™ Rabbit Anti-VHH antibodies can specifically recognize alpaca or llama IgG2&3 so that they can be used to detect and enrich PMBCs producing VHH from blood samples of immunized animals, as shown in Figure 10. Using these antibodies, researchers can obtain heavy chain antibody-expressing B cells from the samples using FACS or MACS. When combined with specific antigen screening, it can significantly reduce the workload and improve the efficiency of library construction and b-cell cloning in VHH discovery.

  

  Figure 10. VHH-positive PBMCs of non-immunized camel were sorted by FACS using 2 μg MonoRab™ Rabbit Anti-Camelid VHH Cocktail [PE] (Cat.No. A02018) . As shown in the figure, the proportion of VHH-positive PBMC cells was 23.579%, which was consistent with the theoretical value, indicating that A02018 has good specificity for VHH-positive PBMCs. A02018 is ideal to be used for isolation of VHH-positive PBMCs.

Related Products

Resource

MonoRab™ Anti-VHH Antibodies

Unique Tools for Detecting Camelid/Humanized Single Domain Antibodies

Free Download

What is VHH?

Members of the Camelidae family including llamas, alpacas, and camels produce two different types of IgG antibodies. One type is the conventional IgG1 (MW~150 kDa) composed of two heavy chains and two light chains. The other type includes IgG2 and IgG3 (MW~90 kDa) which lack the light chains as well as one constant domain of the heavy chain (CH1 domain). Those antibodies are commonly called heavy chain antibodies (HCAbs). The variable domain of the HCAbs is referred to as single domain antibody, also called sdAb or nanobody or VHH. VHH is considered the smallest antigen-binding entity.

Back to the Top