Biotin and avidin or streptavidin (SA) have a strong specific interaction, making it an ideal tool for protein biochemistry. However, generating high-quality biotinylated proteins poses a substantial hurdle.
Scientists frequently encounter technical issues, such as excessive batch-to-batch variance, poor labeling, and unpredictable performance. The choice of biotinylation procedures also significantly influences the final output. Every target protein (assay) has unique biochemical properties and requires a specific solution.
These challenging characteristics make it difficult for laboratories to create and use biotin conjugates, and only a few solutions are available on the market.
ACROBiosystems has developed a unique library of pre-labeled and experimentally verified biotinylated recombinant proteins. The Mabsol® biotinylated protein collection contains over a hundred commonly studied drug targets and biomarker proteins.
The company starts with high-quality recombinant proteins that closely resemble natural conformation and modification. It exhausts all possibilities to achieve maximum bioactivity and detection sensitivity. To ensure the fewest batch-to-batch changes, the company implements the strictest quality control techniques.
The Mabsol® biotinylated protein collection is comprised of two unique and complementary product series: the UltraLys series, which utilizes an in-house-designed chemical labeling approach, and the PrecisionAvi series, which is built using Avitag™ technology. Every aspect is carefully considered when creating these products.
Products
PrecisionAvi series
An Exclusive Collection of Ready-to-Use Avitag™ Biotinylated Proteins
This product line utilizes Avitag™ technology exclusively. In short, the Avi tag, a 15 amino acid peptide, is attached to the recombinant protein during expression vector assembly. The Avi tag’s single lysine residue is enzymatically biotinylated by BirA, an E. Coli biotin ligase.
This single-point enzymatic labeling approach has various advantages for binding assays that are often used.
- The biotinylation only occurs on the lysine residue of the Avi tag
- Even when immobilized on an avidin-coated surface, the protein orientation remains consistent
- There is no disruption to the target protein’s inherent binding properties
All Products
Table 1. Source: ACROBiosystems
. |
2B4 |
4-1BB |
4-1BB Ligand |
ACE2 |
Alpha-Synuclein |
Angiopoietin-2 |
Angiopoietin-like 3 |
ANGPTL7 |
APRIL |
B7-1 |
B7-2 |
B7-H3 |
B7-H3 (4Ig) |
B7-H4 |
B7-H5 |
B7-H6 |
B7-H7 |
BAFF |
BAFFR |
BCMA |
BLAME |
BTLA |
BTN1A1 |
BTN2A1 |
BTN3A1 |
BTN3A2 |
CA125 |
Carbonic Anhydrase IX |
CBLB |
CD133 |
CD14 |
CD155 |
CD161 |
CD19 |
CD2 |
Click here to learn about many more products.
UltraLys Series
A Special Series of Chemically Labeled Biotinylated Proteins with Ultra-Sensitivity
This series of products is manufactured using an in-house-designed chemical labeling procedure. The primary amines in the side chains of lysine residues and the N-terminus of proteins are conjugated with biotins, resulting in many biotin attachments on a single protein molecule.
- Higher detection sensitivity can be obtained
- Includes an in-house designed chemical labeling process
All Products
Table 2. Source: ACROBiosystems
. |
Adalimumab |
B7-H4 |
Bevacizumab |
CD19 |
CD24 |
CD3
epsilon |
CD3E &
CD3D |
Cetuximab |
CX3CL1 |
DNP |
EpCAM |
EphB4 |
ErbB3 |
FcRn
(FCGRT & B2M) |
FMC63 |
GM-CSF |
GPA33 |
Growth
Hormone R |
Her2 |
IL-6 |
Mesothelin |
NKp46 |
Nucleocapsid
protein |
Protein L |
SOST |
Spike RBD |
TFPI |
TNF-alpha |
Transferrin R |
VEGF165 |
Vitronectin |
|
|
|
|
Applications
- FACS
- Enrichment
- Biopanning
- SPR
- ELISA
- BLI
Key Features
Closest to Natural Conformation and Modification
Recombinant proteins, such as biotinylated proteins, are produced using the patented HEKMax® expression platform. As expression hosts, human HEK293 cells have several advantages over other cell types, as shown in the table below.
Table 3. Source: ACROBiosystems
Expression Systems |
Folding |
Phosphorylation |
Proteolytic Processing |
Glycosylation |
E.Coli |
+ |
N/A |
N/A |
N/A |
Insect Cell |
++ |
++ |
++ |
Poor |
Plant Cell |
+++ |
+++ |
+++ |
Poor |
CHO Cell |
++++ |
++++ |
++++ |
Non-Human Like |
Human Cell |
+++++ |
+++++ |
+++++ |
Human Authentic |
High Bioactivity and Detection Sensitivity
The protein's structure determines the bioactivity of biotinylated proteins and how biotinylation is carried out. The company evaluates several tag and biotinylation techniques for each protein while evaluating the results in various binding experiments. However, only those with the best performances are chosen for production. Figure 1 shows the internal evaluation experiments.
![Binding activity of different forms of biotinylated PD1 evaluated in a functional ELISA against rhPD-L1 (Cat. No. PD1-H5258).](https://d1g9yur4m4naub.cloudfront.net/images/equipments/ImageForEquipment_615_17165427053728888.png)
Figure 1. Binding activity of different forms of biotinylated PD1 evaluated in a functional ELISA against rhPD-L1 (Cat. No. PD1-H5258). Image Credit: ACROBiosystems
ACROBiosystems’ biotinylated proteins provide high bioactivity and great detection sensitivity in various applications. In functional ELISA, biotinylated human TACI binds strongly to BAFF (Figure 2).
![Immobilized Human BAFF, Fc Tag, active trimer (Cat. No. BAF-H5261) at 1 μg/mL (100 μL/well) can bind Biotinylated Human TACI, Fc, Avitag (Cat. No. TAI-H82F6) with a linear range of 0.002–0.05 μg/mL.](https://d1g9yur4m4naub.cloudfront.net/images/equipments/ImageForEquipment_615_17165427160463767.png)
Figure 2. Immobilized Human BAFF, Fc Tag, active trimer (Cat. No. BAF-H5261) at 1 μg/mL (100 μL/well) can bind Biotinylated Human TACI, Fc, Avitag (Cat. No. TAI-H82F6) with a linear range of 0.002–0.05 μg/mL. Image Credit: ACROBiosystems
Interestingly, compared to random immobilization of unconjugated CD155, oriented immobilization of biotinylated CD155 offers much higher sensitivity (see Figure 3). Figures 4 and 5 show that the biotinylated PD1 on AlphaLISA can be utilized to measure the binding activity with its ligands, PD-L1 and PD-L2.
ELISA
![Comparison of oriented immobilization (biotinylated CD155 on precoated streptavidin plate, Cat. No. CD5-H82E3) and random immobilization (unconjugated CD155 direct coating on 96-well plate, Cat. No. CD5-H5223) binding to TIGIT (Cat. No. TIT-H5254). Oriented immobilization of biotinylated CD155 offers much higher sensitivity than random immobilization of unconjugated CD155.](https://d1g9yur4m4naub.cloudfront.net/images/equipments/ImageForEquipment_615_1716542722548419.png)
Figure 3. Comparison of oriented immobilization (biotinylated CD155 on precoated streptavidin plate, Cat. No. CD5-H82E3) and random immobilization (unconjugated CD155 direct coating on 96-well plate, Cat. No. CD5-H5223) binding to TIGIT (Cat. No. TIT-H5254). Oriented immobilization of biotinylated CD155 offers much higher sensitivity than random immobilization of unconjugated CD155. Image Credit: ACROBiosystems
AlphaLISA
![Biotinylated Human PD-1 (Cat. No. PD1-H82F1) at 1 μg/mL (5 μL/well) can bind Human PDL1 (Cat. No. PD1-H5229) with a linear range of 0.02–0.625 μg/mL](https://d1g9yur4m4naub.cloudfront.net/images/equipments/ImageForEquipment_615_17165427287267744.png)
Figure 4. Biotinylated Human PD-1 (Cat. No. PD1-H82F1) at 1 μg/mL (5 μL/well) can bind Human PDL1 (Cat. No. PD1-H5229) with a linear range of 0.02–0.625 μg/mL. Image Credit: ACROBiosystems
Biotinylated proteins can easily determine the affinity between a protein and a therapeutic antibody in an SPR test, as shown in Figures 6 and 7. Biotinylated proteins are also frequently used in cell-based assays, such as cytotoxicity assays and CAR expression evaluations (see Figure 8).
![Biotinylated Human PD-1 (Cat. No. PD1-H82F1) at 1 μg/mL (5 μL/well) can bind Human PDL2 (Cat. No. PD2-H5220) with a linear range of 0.02–0.625 μg/mL.](https://d1g9yur4m4naub.cloudfront.net/images/equipments/ImageForEquipment_615_17165427353772351.png)
Figure 5. Biotinylated Human PD-1 (Cat. No. PD1-H82F1) at 1 μg/mL (5 μL/well) can bind Human PDL2 (Cat. No. PD2-H5220) with a linear range of 0.02–0.625 μg/mL. Image Credit: ACROBiosystems
SPR
![Immobilized biotinylated human VEGF165 (Cat. No. VE5-H82Q0) on CM5 Chip via streptavidin, can bind Avastin with an affinity constant of 0.417 nM as determined in SPR assay (Biacore T200).](https://d1g9yur4m4naub.cloudfront.net/images/equipments/ImageForEquipment_615_17165427421024082.png)
Figure 6. Immobilized biotinylated human VEGF165 (Cat. No. VE5-H82Q0) on CM5 Chip via streptavidin, can bind Avastin with an affinity constant of 0.417 nM as determined in SPR assay (Biacore T200). Image Credit: ACROBiosystems
BLI
![Loaded Biotinylated Human PD-1, Avitag, His Tag (recommended for biopanning) (Cat. No. PD1-H82E4) on SA Biosensor, can bind Human PD-L1, His Tag (HPLC verified) (Cat. No. PD1-H5229) with an affinity constant of 2.4 μM as determined in BLI assay (ForteBio Octet Red96e).](https://d1g9yur4m4naub.cloudfront.net/images/equipments/ImageForEquipment_615_17165427484392708.png)
Figure 7. Loaded Biotinylated Human PD-1, Avitag, His Tag (recommended for biopanning) (Cat. No. PD1-H82E4) on SA Biosensor, can bind Human PD-L1, His Tag (HPLC verified) (Cat. No. PD1-H5229) with an affinity constant of 2.4 μM as determined in BLI assay (ForteBio Octet Red96e). Image Credit: ACROBiosystems
Cell-Based Assay
![Recombinant human TNF-alpha (Cat. No. TNA-H82E3) induces cytotoxicity effect on the WEH1-13VAR cells in the presence of the metabolic inhibitor actinomycin D. The EC50 for this effect is 0.014–0.029 ng/mL. The result shows that the biotinylated human TNF-alpha is consistent with naked TNF-alpha in cytotoxicity assay](https://d1g9yur4m4naub.cloudfront.net/images/equipments/ImageForEquipment_615_17165427561251555.png)
Figure 8. Recombinant human TNF-alpha (Cat. No. TNA-H82E3) induces cytotoxicity effect on the WEH1-13VAR cells in the presence of the metabolic inhibitor actinomycin D. The EC50 for this effect is 0.014–0.029 ng/mL. The result shows that the biotinylated human TNF-alpha is consistent with naked TNF-alpha in cytotoxicity assay. Image Credit: ACROBiosystems
Low Batch-to-Batch Variation
ACROBiosystems routinely implements strict quality control methods to maintain the products' functioning. Recently released products are compared side by side with the internal standard in various tests.
Products can only be released if there is a reasonable margin of variance. Three batches of biotinylated hTNF-alpha (Cat. No. TNA-H82E3) were examined and compared using a conventional ELISA assay against Adalimumab, as shown in Figure 9. The findings show barely any batch variation among the examined samples. As demonstrated by the cell-based assay (see Figure 10), there is high batch-to-batch conformity.
![In the above ELISA analysis, three different lots of biotinylated hTNF-alpha (Cat. No. TNA-H82E3) were used to detect immobilized Adalimumab (0.5 μg/mL). The result showed that the batch variation among the tested samples is negligible](https://d1g9yur4m4naub.cloudfront.net/images/equipments/ImageForEquipment_615_17165427622139972.png)
Figure 9. In the above ELISA analysis, three different lots of biotinylated hTNF-alpha (Cat. No. TNA-H82E3) were used to detect immobilized Adalimumab (0.5 μg/mL). The result showed that the batch variation among the tested samples is negligible. Image Credit: ACROBiosystems
![Recombinant biotinylated human TNF-alpha (Cat. No. TNA-H82E3) induces cytotoxicity effect on the WEH1-13VAR cells in the presence of the metabolic inhibitor actinomycin D. The EC50 for this effect is 0.029–0.052 μg/mL](https://d1g9yur4m4naub.cloudfront.net/images/equipments/ImageForEquipment_615_17165427685713567.png)
Figure 10. Recombinant biotinylated human TNF-alpha (Cat. No. TNA-H82E3) induces cytotoxicity effect on the WEH1-13VAR cells in the presence of the metabolic inhibitor actinomycin D. The EC50 for this effect is 0.029–0.052 μg/mL. Image Credit: ACROBiosystems