MonoRab™ Anti-VHH Antibodies

Highlights

Exceptional Specificity

• No cross-reactivity with mouse, rat, rabbit, goat, and human immunoglobulins, enabling their use in the detection of blood samples from different species

Versatile Applicability

• Ability to recognize camelid and humanized VHHs

High Quality and Consistency

• Monoclonal antibody manufacturing process consistently ensures high standards of quality, performance, and reliability

Product List

• Anti-Camelid VHH, mAb

  Cat. No.	Product Name	Quantity	Price	Order
  A01860	MonoRab™ Rabbit Anti-Camelid VHH Antibody, mAb		$209.00 200 μl
  A01861	MonoRab™ Rabbit Anti-Camelid VHH Antibody [HRP], mAb		$209.00 200 μl
  A01862	MonoRab™ Rabbit Anti-Camelid VHH Antibody [iFluor488], mAb		$209.00 200 μl
  A01863	MonoRab™ Rabbit Anti-Camelid VHH Antibody [iFluor555], mAb		$209.00 200 μl
  A01994	MonoRab™ Rabbit Anti-Camelid VHH Antibody [iFluor 647], mAb		$209.00 200 μl
  A01995	MonoRab™ Rabbit Anti-Camelid VHH Antibody [Biotin], mAb		$209.00 200 μl

• Anti-Humanized VHH, mAb

  Cat. No.	Product Name	Quantity	Price	Order
  A02165	MonoRab™ Rabbit Anti-Humanized VHH Antibody, mAb		$209.00 200 μl
  A02166	MonoRab™ Rabbit Anti-Humanized VHH Antibody [Biotin], mAb		$209.00 200 μl
  A02167	MonoRab™ Rabbit Anti-Humanized VHH Antibody [HRP], mAb		$209.00 200 μl
  A02168	MonoRab™ Rabbit Anti-Humanized VHH Antibody [iFluor 488], mAb		$209.00 200 μl
  A02169	MonoRab™ Rabbit Anti-Humanized VHH Antibody [iFluor 555], mAb		$209.00 200 μl
  A02170	MonoRab™ Rabbit Anti-Humanized VHH Antibody [iFluor 647], mAb		$209.00 200 μl

• Anti-Camelid VHH, mAb Cocktail

  Cat. No.	Product Name	Quantity	Price	Order
  A02014	MonoRab™ Rabbit Anti-Camelid VHH Cocktail		$375.00 200 μl
  A02015	MonoRab™ Rabbit Anti-Camelid VHH Cocktail [Biotin]		$375.00 200 μl
  A02016	MonoRab™ Rabbit Anti-Camelid VHH Cocktail [HRP]		$375.00 200 μl
  A02017	MonoRab™ Rabbit Anti-Camelid VHH Cocktail [FITC]		$375.00 200 μl
  A02018	MonoRab™ Rabbit Anti-Camelid VHH Cocktail [PE]		$375.00 200 μl
  A02019	MonoRab™ Rabbit Anti-Camelid VHH Cocktail [iFluor 647]		$375.00 200 μl
  A02020	MonoRab™ Rabbit Anti-Camelid VHH Cocktail [iFluor 555]		$375.00 200 μl
  A02021	MonoRab™ Rabbit Anti-Camelid VHH Cocktail [iFluor 488]		$375.00 200 μl

• Related Products

  Cat. No.	Product Name	Quantity	Price	Order
  A02022	MonoRab™ Rabbit IgG Control (Whole Molecule), mAb		$105.00 200 μl
  A02023	MonoRab™ Rabbit IgG Control [Biotin] (Whole Molecule), mAb		- 200 μl
  A02024	MonoRab™ Rabbit IgG Control [FITC] (Whole Molecule), mAb		$105.00 200 μl
  A02025	MonoRab™ Rabbit IgG Control [PE] (Whole Molecule), mAb		$105.00 200 μl
  A02026	MonoRab™ Rabbit IgG Control [iFluor 647] (Whole Molecule), mAb		$105.00 200 μl
  A02027	MonoRab™ Rabbit IgG Control [iFluor 555] (Whole Molecule), mAb		$105.00 200 μl
  A02028	MonoRab™ Rabbit IgG Control [iFluor 488] (Whole Molecule), mAb		$105.00 200 μl
  L00905	MonoRab™ Anti-Camelid VHH Affinity Resin		$474.99 1 ml Resin (Total 2 ml) 5 ml Resin (Total 10 ml) 10 ml Resin (Total 20 ml) 25 ml Resin (Total 50 ml)

  Find more Control Antibodies

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.

Application Data

• 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 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.

Science of VHH and scFv

• VHH and scFv Applications

• Advantages and Features

• Full Offerings

Single-chain fragment variable (scFv) and nanobody (also known as VHH) are both genetically engineered antibodies. Due to their small molecular weight, strong penetration, short half-life and low immunogenicity, these antibodies play an important role and have broad application prospects in disease prevention, diagnosis and treatment.

Bottlenecks of existing scFv and VHH research tools and reagents

scFv and VHH have broad applications and are attracting increasing interest from research institutions and pharmaceutical companies. However, they lack Fc fragments, making Fc-based detection and purification methods inapplicable. Additionally, pharmaceutical companies avoid adding tag proteins like Flag and His to scFv or VHH to prevent potential risks, meaning tag-based strategies are also unsuitable. For scFv and VHH without tags, current detection and purification methods face several challenges:

• Universal detection methods for scFv, such as Anti-Fab antibodies and Protein L, are not specifically developed for scFv, leading to compatibility issues, high background values, and low detection rates.
• No efficient universal detection and purification reagent exists for VHH.
• Idiotypic detection tools like anti-idiotypic antibodies and antigens are impractical for early-stage screening due to their long development cycles, high costs, and low applicability.

How GenScript Supports scFv and VHH Research with Advanced Solutions?

To support scFv and VHH drug research, GenScript offers a comprehensive solution with tools and reagents for every research stage. This includes antibody discovery, purification, functional identification, lentiviral transduction titer determination, CAR-T cell characterization and separation, and pharmacokinetic detection. Our goal is to provide complete support for researchers to ensure success in scFv and VHH research.

Figure 11. Overall solution for scFv and VHH

Advantages of scFv and VHH Applications

VHH nanobodies and single-chain fragment variables (scFv) are genetically engineered antibodies with advantages such as small size, strong penetration, short half-life, low immunogenicity, and easy engineering. They excel over full-length antibodies in cell therapy, bispecific antibody construction, solid tumor treatment, and drug delivery. Additionally, they are widely used in viral infection, biological monitoring, biosensors, and basic research. As research progresses, scFv and VHH are expected to play an increasingly significant role in medicine, agriculture, and industry.

Figure 12. Application of scFv and VHH

Feature Comparison of scFv and VHH

Nature	scFv	VHH
Size	28-35 KDa	12-15 KDa
Blood half-life	＜1h	≪ 1h
Source	Mice, rats, etc.	Camelidae
Development time	Shorter	Longer
Water Solubility	Good water solubility (+)	Good water solubility (+++)
Gather	Easy to aggregate	Almost no gathering
Stability	Moderate stability (+)	Excellent stability (++)
Resistant to extreme environments	Not resistant to high temperatures or extreme pH	Stable at high temperatures and extreme pH
Tissue penetration	Strong tissue penetration (++)	Better tissue penetration (+++)
Affinity	High affinity (+)	Higher affinity (++)
Cost of production	Low	Lower
Engineering Transformation	Easier to modify (+)	Very easy to modify (+++)
Epitope diversity	Consistent with traditional antibodies	New epitopes available
Hidden epitope recognition	Difficult to identify hidden epitopes	Can identify hidden epitopes
Expression system	Prokaryotes, yeast, mammals	Prokaryotes, yeast, mammals

*Note: “+” represents the degree of the descriptive item. The more “+” there are, the higher the degree is.

Table 1. Comparison of scFv and VHH structures and physicochemical properties²

Product List

Product Type		Species Specificity	Unconjugated	Conjugated
				HRP	Biotin	iFluor 488	iFluor 555	iFluor 647	PE	FITC
Anti-VHH antibody	Anti-Camelid VHH,mAb	Llama, Camel, Alpaca	A01860	A01861	A01995	A01862	A01863	A01994	A02227	A02226
	Anti-Humanized VHH, mAb	Humanized, Llama, Camel, Alpaca	A02165	A02167	A02166	A02168	A02169	A02170	A02171	A02172
	Anti-Camelid VHH, mAb Cocktail	Humanized, Llama, Camel, Alpaca	A02014	A02016	A02015	A02021	A02020	A02019	A02018	A02017
Anti-scFv antibody	Anti-scFv Antibody Cocktail	Humanized, Mouse	A02282	A02289	A02283	A02286	A02287	A02288	A02285	A02284
	Anti-scFv Antibody Cocktail (Min X)*	Humanized, Mouse		A02306	A02303	A02304		A02305	A02315
GS Linker Antibody								A02311	A02314
Application
Flow cytometry			✔		✔	✔	✔	✔	✔	✔
Western blot / Dot blot			✔	✔	✔
ELISA			✔	✔	✔
Immunohistochemistry			✔	✔	✔
Immunofluorescence			✔		✔	✔	✔	✔	✔	✔
Cell sorting			✔		✔	✔	✔	✔	✔	✔

*Anti-scFv antibody Cocktail (Min X) has no cross-reactivity to human IgG, mouse IgG, goat IgG and rabbit IgG.

Anti-VHH and anti-scFv antibody derivatives

Product Number	Product Name
L00905	MonoRab™ Anti-Camelid VHH Affinity Resin
L00946	MonoRab™ Rabbit Anti-Camelid VHH Antibody Plate (Clear, 8×12 strip)
L00951	MonoRab™ Anti-Humanized VHH Affinity Resin FF
L01008	MonoRab™ Rabbit Anti-VHH Microbeads
L01033*	VHH Affinity Ligand ELISA kit
L01034	MonoRab™ Anti-VHH Affinity Magnetic Beads

*The L01033 VHH Affinity Ligand ELISA kit already contains the CaptureSelect™ AAVX standard. If you want to detect Capto AVB、CaptureSelect™ AAV8 or CaptureSelect™ KappaXP，you need to purchase the AVB Affinity Ligand Standard(GenScript, Z03794)、AAV8 Affinity Ligand Standard(GenScript, Z03795) or KappaXP Affinity Ligand Standard (GenScript, Z03796) separately.

Other products

Product	Unconjugated	Conjugated
		HRP	Biotin	iFluor 488	iFluor 555	iFluor 647	PE	FITC
Mouse Anti-Human IgG Fab Antibody (12H3C4A6), mAb		A01855
protein L		M00098	M00097	M00921		M00922		M00920
Mouse Anti-Human IgG Fc Antibody (50B4A9), mAb	A02257	A01854
Streptavidin	Z02043	M00091
MonoRab™ Rabbit IgG Control (Whole Molecule), mAb	A02022			A02028	A02027	A02026	A02025
Human Kappa Light Chain (2F1C1), mAb, Mouse	V06801
Human Lambda Light Chain (2D54), mAb, Mouse	V06901
MonoRab™ Biotin Antibody (53C8), mAb, Rabbit	V90201			A02248		A02249	A02250

Reference

• 1. Van Audenhove I , Gettemans J .Nanobodies as Versatile Tools to Understand, Diagnose, Visualize and Treat Cancer[J].EBioMedicine, 2016, 8(C).DOI:10.1016/j.ebiom.2016.04.028.
• 2. Asaadi Y , Jouneghani F F , Janani S ,et al.A comprehensive comparison between camelid nanobodies and single chain variable fragments[J].Biomarker Research, 2021, 9(1):87-.DOI:10.1186/s40364-021-00332-6.

Resource

MonoRab™ Anti-VHH Antibodies

Unique Tools for Detecting Camelid/Humanized Single Domain Antibodies

Free Download

GenScript’s scFv and VHH Handbook

Comprehensive Applications of Single-Chain Antibodies and Nanobodies for CAR-T Therapy and Antibody Drug Research

Free Download

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