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AqT™ Biotin Active Ester Antibody Labeling Kit (T2A15)
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| SDS | USER MANUAL CM86140 |
AqT™ Biotin Active Ester Antibody Labeling kit (T2A15) is a proprietary biotinylation kit developed at CellMosaic for obtaining high quality and high loading of biotinylated antibody with minimal or no aggregation. The kit uses a biotin modified with a super-hydrophilic, water-soluble, and charge neutral AqueaTether™ (AqT™) T2A15 linker to improve its water solubility and decrease its non-specific interaction with other proteins or plastic surfaces. Biotinylated antibodies with AqT™ linker can largely retain its native properties. AqT™ T2A15 linker composed of two threitols and 15 atom lengths. This atom length is equivalent to PEG4 (16 atom lengths) and, based on our experience, can largely keep the bivalent molecule function independent. The conjugates we have tried with this atom length include antibody oligo, streptavidin oligo, and HRP oligo, among others.
For a detailed description of the AqT™ technologies, please see our technology section.
The CM86140 kit is designed to biotinylate any antibody via surface amines of the antibody. The user supplies the antibody. Using the kit components, the user biotinylates their antibody in one simple step, followed by filtration-type purification to remove any unreacted AqT™-biotin, exchanging it with PBS or other buffer for downstream application. A quick 10-minute biotin assay (protocol and reagent) is also included in the kit to obtain the biotinylation level immediately after labeling. The biotinylation level can easily be tuned by increasing or decreasing the reaction temperature. The hands-on time is less than 30 minutes, and the whole preparation time varies from 3 h to overnight depending on the reaction time.
The kit has multiple configurations for labeling various amounts of antibody (10 μg to 3 mg) with either one (x1) or three (x3) antibody samples.
| Catalog Number | Number of Reactions in Kit | Antibody Amount per Reaction |
| CM86140.1x1 | 1 | 10 μg |
| CM86140.1x3 | 3 | |
| CM86140.2x1 | 1 | 50 μg |
| CM86140.2x3 | 3 | |
| CM86140.3x1 | 1 | 100 μg |
| CM86140.3x3 | 3 | |
| CM86140.4x1 | 1 | 1 mg |
| CM86140.4x3 | 3 | |
| CM86140.5x1 | 1 | 3 mg |
| CM86140.5x3 | 3 |
CM86140 and Its Product Compared to Other Commercial Antibody Biotinylation Kits
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| AqT™ Biotin Labeling Kit | Other Commercial Kits |
| Biotinylated Product Features
| Chemical structure | Fully Disclosed · Complete knowledge of your product | Varies by vendor, mostly not disclosed. |
| Properties | Retain antibody properties · No changes in hydrophobicity · No increase of apparent MW (hydrodynamic volume) · No increase in aggregation · Minimal heterogeneity after labeling | Change antibody properties · Increased hydrophobicity (non-specific interaction) · Increased apparent MW (hydrodynamic volume) · May increase aggregation · Highly heterogeneous product | |
| Stability | Stable even with high biotin loading Can be easily stored frozen and in lyophilized form without any stabilizer | Decreased stability High biotin loading may cause aggregation and precipitation | |
| Kit Features | Reagent | Proprietary water-soluble pre-activated biotin with 15-atom spacer | Biotin (zero length) or biotin with hydrophobic ethylene- or polyethylene glycol-type linker |
| Labeling buffer | 100% aqueous buffer | Some percentage of organic solvent needed, such as DMSO | |
| Biotinylation level | Predictable · Tunable loading, from low to very high, per your need | Unpredictable Usually target low labeling to avoid aggregation (difficult to tune) | |
| Usability | Easy to use · One step labeling · <30 minutes hands-on time | Comparable quick labeling may use EDC chemistry with antibody crosslinking to each other causing large aggregates | |
| Purification method & components | Included Product is free of any unreacted AqT™ biotin after purification. | Varies by vendor, usually not included | |
| Biotin Assay | Included A quick 10-minute biotin assay protocol with reagents and buffer | Not included |
Requirements for antibody:
- MW: ≥ 100 KDa (if MW is lower, please use an alternative kit)
- Purity: preferably > 90% pure by gel electrophoresis
- Amount: 10 μg to 3 mg protein content as measured by UV
Labeling Chemistry
Overview of the workflow for preparing AqT™ antibody-biotin conjugates
Comparing the hydrophilicity of AqT™ biotin-T2A15 acid with biotin and biotin-PEG4 acid (C18 HPLC Profile)
Comparing the hydrophilicity and heterogenicity of AqT™ biotin-T2A15 labeled antibody with unmodified biotin labeled antibody (HIC HPLC Profile)
Comparing the apparent MW (hydrodynamic volume) of AqT™ biotin-T2A15 labeled antibody with unmodified biotin labeled antibody (SEC HPLC Profile)
