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| Product Name | Biotin Protein Labeling Reagent Kit |
| Catalog No. | PTR-HMM-0068 |
| Description | A reagent kit for the covalent attachment of biotin to proteins via NHS-ester chemistry on primary amines. Biotinylated proteins are widely used as detection reagents in immunoassays, immunohistochemistry, and protein interaction studies. |
| Intended Use | Covalent biotin conjugation to antibodies, enzymes, and other proteins for use as detection probes in ELISA, Western blot, immunohistochemistry, and protein interaction assays. |
| Principle / Technology | N-hydroxysuccinimide (NHS) ester of biotin reacts with primary amine groups on lysine residues and the N-terminus of proteins to form stable amide bonds under mildly alkaline conditions. |
| Detection Method | Biotinylated proteins are subsequently detected using streptavidin- or avidin-conjugated enzymes, fluorophores, or other reporter molecules |
| Sample Type | Purified proteins, antibodies, or peptides dissolved in amine-free buffer (avoid Tris, glycine, or other primary amine-containing buffers) |
| Performance Range / Specifications | Labeling efficiency: typically 3–6 biotin molecules per IgG antibody under standard reaction conditions; degree of labeling can be adjusted by varying the molar ratio of biotin reagent to protein |
| Sensitivity / LOD | Effective labeling of proteins at concentrations as low as 1 mg/mL; amine-containing carrier proteins (BSA, gelatin) must be removed prior to labeling |
| Specificity | NHS-biotin reacts with all accessible primary amines including lysine side chains and protein N-termini; labeling may affect protein activity if amines are present in the active site or binding interface |
| Reaction Conditions / Protocol | Dissolve protein in amine-free buffer at 1–10 mg/mL, add biotin reagent at the desired molar ratio, incubate at room temperature for 30–60 minutes, remove excess unreacted biotin by desalting column or dialysis |
| Components / Formulation | NHS-PEG4-biotin reagent (pre-weighed), anhydrous DMSO for reagent dissolution, Zeba desalting spin columns for unreacted biotin removal, HABA-avidin reagent for biotin incorporation quantification, reaction buffer, detailed protocol |
| Storage Conditions | -20°C for biotin reagent (desiccated and protected from moisture); desalting columns and HABA reagent at 2–8°C; DMSO at room temperature |
| Shelf Life | 24 months for biotin reagent when stored desiccated at -20°C |
| Package Specifications | Sufficient for 5–10 labeling reactions (1 mg protein each) |
| Product Form | Solid biotin reagent (NHS-PEG4-biotin) with liquid accessory reagents |
| Quality Control | Each lot of biotin reagent tested for reactivity by conjugation to a standard IgG and determination of biotin incorporation by HABA assay |
| Key Features | PEG4 spacer arm reduces steric hindrance and improves streptavidin accessibility; spin column format simplifies unreacted biotin removal; HABA assay enables rapid biotin incorporation quantification |
For research use only, not for clinical use.
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